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-Immunology CONTENTS: 1. Water and fluid- compartments
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RENAL PATHOPHYSIOLOGYThese lecture notes accompany my lectures on
pathophysiology in the study module "Kidneys and Urinary Tract" at the
Medical University of Innsbruck. The
English version serves two purposes: as a learning aid for international
students and to encourage German-speaking students to familiarize themselves
with medical English; the lectures are delivered in German. Thanks to Prof. Gert Mayer for critically reading these notes! The
translation from the original German version is my own; I am
afraid it will occasionally sound appalling to native English speakers, but it
should at least be intelligible. There is also a printable pdf-version.
Version
1.5e ©Arno Helmberg 2016-2020
Terms of use: http://www.helmberg.at/terms.htm
Why do we have kidneys? If we produce water-soluble
metabolites which are hard to transport across cells and are toxic beyond a
certain concentration, we need a direct pathway from the extracellular fluid
compartment to the outside to get rid of them. In fact, such metabolites exist:
for example, when we use amino acids to produce energy, what remains are the
amino groups. In the form of either ammonium or the less toxic urea, nitrogen
is hard to secrete. This problem is solved by shuttling it directly out.
Yet, directly pumping out extracellular fluid would
kill us within minutes, unless we succeed in reabsorbing everything we need
with utter efficiency, in a precisely regulated process, starting with water.
That's what kidneys are about. And that helps to understand what's going wrong
in kidney disease. Let's start with water.
1. WATER AND FLUID COMPARTMENTS
Water is the most abundant constituent of our body, at
50-60 % of our body mass (75% in infants). Why this large variation? The wide
range is mainly due to individual differences in fat stores and muscle mass.
Fat excludes water; therefore, fat cells contain very little water. Under the
influence of androgens, males on average have a higher proportion of
"watery" muscle to water-excluding fat, while even lean females have
a lower proportion and hence a lower percentage of water, tending to 50%. Almost
two thirds of body water resides inside cells, the rest is extracellular fluid.
The body of a lean woman of 70 kg thus may contain 35
liters of total body water, of which approximately 21 liters (60%) are
intracellular fluid and 14 liters extracellular fluid, including interstitial
fluid and blood plasma (about 3 l). In comparison, a 70 kg non-obese male may
contain 42 liters of total body water, with 25 l of intracellular and 17 l of
extracellular fluid. For both sexes, blood volume is the sum of three liters of
plasma plus the volume of cells contained in the blood, adding up to 5 l in
females and 5.5 l in males: hematocrit, the cellular fraction of the blood, is
a little higher in males.
Let's review a few necessary definitions:
Osmole is a unit of measurement expressing the number of
moles of solute that contribute to the osmotic pressure of a solution
Osmotic
concentration (Osmolarity)
is expressed in osmol/l. In medicine, it is usually not determined precisely
but approximated by calculation, as in:
Osmotic concentration of serum =
= 2 Na + Urea + Glucose (all in mmol/l)
or, in case urea and glucose are given in mg/dl:
=(2
x serum sodium [mEq/l]) + (BUN [mg/dl]/2.8) + (glucose [mg/dl]/18)
Osmolality, on the other hand, relates not to a volume but
rather to a kg of solvent (osmoles/kg solvent). Osmolality is measured directly
by either freezing-point depression of water or vapor pressure techniques.
For a given solution, osmotic concentration is
slightly lower than osmolality, because the volume used in calculating osmotic
concentration includes the solutes, while osmolality is based on 1 kg of
solvent excluding the weight of any solute. In other words, to get from
osmolarity to osmolality, you have to "top up" your solution,
including more particles. In practice, with the relatively small amounts of
solutes per volume or mass of solvent in our body, the difference is almost negligible.
Osmolality of intracellular volume, interstitial
volume as well as blood plasma is equal at 290 mosmol/kg; otherwise water would
shift until a new equilibrium is reached. The composition of solutes is very
different, however. Life started in the oceans, and multicellular organisms
migrating elsewhere took their "inner ocean" with them. Therefore,
our extracellular compartment is rich in salt, NaCl, while within cells, our
main cation is K+. Cell membranes constitute the border between
intracellular and extracellular compartments. The main factor maintaining this
asymmetry between the two compartments is Na-K-ATPase, which is pumping three
Na+ ions out of the cell in exchange for 2 K+ ions. Regarding
extracellular compartments, the main asymmetry between blood plasma and
interstitial fluid is due to protein content. The liver constantly releases
proteins into the blood, many of which are too big to pass the endothelial
barrier as long as there is no inflammation.
Thus, our main fluid compartments differ as follows:
Albumin is our most abundant extracellular protein. It
is synthesized by the liver at a rate of 9-12 g per day. It is a 66 kDa
protein of triangular shape, with a strong negative net charge of -15.
Therefore, it lines up at the anode-near end of all proteins in serum electrophoresis.
Although we think of albumin as a plasma protein, there is actually more interstitial
albumin than plasma albumin: albumin concentration is four times higher in
plasma, but interstitial volume is five times plasma volume. The rate at which
albumin leaves the plasma depends on the tightness of capillaries in the
respective organs. While very little albumin passes the wall of brain
capillaries, capillaries in gut, muscle or fat tissue are much more permeable,
not to speak of the fenestrated endothelia of liver and glomeruli. For example,
albumin concentration has been reported to be 1.3 g/dl in muscle and 0.7 g/dl
in fat interstitial fluid. Albumin is recycled to the blood via the lymphatics.
If there were no lymphatic removal, interstitial protein concentration would
eventually equal that in the plasma.
If water is the most abundant constituent of our body,
how do we regulate its volume? Imagine a barrel: it's easy to determine when
it's full. Yet, our body lacks the rigid structure of a barrel; it's more like
a balloon: for its elasticity, it's hard to determine its "correct"
volume.
In fact, our body
regulates water volume in a two-step procedure.
1.
First,
it regulates osmolality very tightly: it adjusts water to keep osmotic pressure
constant, which in practice means it adjusts water to keep extracellular Na+ concentration constant. That still leaves us with the balloon problem: we may
put more or less isotonic saline into the balloon of our body. That is actually
the case: depending on our Na+ intake, our extracellular volume
varies a lot more than our osmolality. Put another way: our osmoregulation is stricter
than our volume regulation.
2.
Yet,
it is clear we also need to measure and regulate volume in a second step, if
with much less precision. For this purpose, we use pressure sensors in
arteries, cardiac atria and the vasa afferentia of renal glomeruli. Yet, our
ability to gauge extracellular fluid volume is restricted by these measuring
devices: we do not have an ability to measure excess extracellular volume in
the form of interstitial edema, ascites or pleural effusion.
Note that we have a single, precise device to measure
osmolality which does a perfect job and several devices to gauge effective
circulatory volume, which leave much to be desired because of their inability
to determine total extracellular volume.
2. REGULATION OF RENAL BLOOD FLOW
The kidneys have a combined weight of less than 0.5%
of the entire body, yet receive approximately 20% of cardiac output. While
resting, cardiac output may be near 5 l/min, of which 1 l/min goes to the
kidneys. This is primarily necessary to be able to excrete solutes that cannot
be secreted by cells. In addition, it reflects high metabolic activity: the
kidneys account for 7-10 % of total O2 consumption of our body in a
resting state.
Renal plasma flow can be measured with the help of
para-aminohippurate (PAH). While traces of PAH may occur in our body by
metabolic processes, for the purpose of determining plasma flow, large
quantities are infused intravenously. PAH is freely filtrated at the glomerulus
and in addition actively secreted by the proximal tubule. Almost all of PAH
reaching the kidney is eliminated in a single pass (92%), so that PAH clearance
approximates renal plasma flow and, with the help of the hematocrit, can be
used to estimate renal blood flow.
About 90% of the blood leaving the glomeruli perfuses
the cortex; only 10%, from a subpopulation of juxtamedullary glomeruli, go to
the medulla via the vasa recta. Because of the high fluid resistance of these
long capillaries, very little of this blood reaches the papilla. This is
important: otherwise, the high osmotic gradient in the medulla would wash out
and we would not be able to concentrate urine. On the other hand, it puts cells
in the medulla in a precarious position: as soon as there is a problem with
blood flow, these cells are prone to suffer damage from lack of oxygen.
Regulation of
glomerular filtration rate
A selective increase of afferent arteriole resistance
decreases glomerular filtration rate (GFR). In contrast, increasing the
resistance of the efferent arteriole has a biphasic response. At the lower end
of the resistance range, an increase in resistance increases GFR as a result of
increased filtration pressure. In addition, filtration fraction goes up: a
higher fraction of blood entering the glomerulus is filtrated, meaning somewhat
less blood is available to perfuse renal parenchyma "behind" the vas
efferens. Yet with further increases of resistance, renal plasma flow declines,
causing GFR to first level off and then decline.
Two levels of regulation affect the glomerular
arterioles. Autoregulation relies on feedback within an individual nephron and
affects only the afferent arteriole. Systemic regulation adds to this via the
autonomous nervous system, as well as via chemical mediators.
Autoregulation is
the result of two independent mechanisms:
1.
Myogenic
response: An increase in arterial pressure opens stretch-activated nonselective
cation channels in smooth muscle cells of afferent arterioles, depolarizing the
membrane and opening voltage-dependent Ca2+ channels, leading to
contraction.
2.
Tubuloglomerular
feedback: an increase of single nephron GFR brings more NaCl to the macula densa in the thick ascending
limb. This is sensed by the cell via its Na-K-2Cl-cotransporter (NKCC2,
SLC12A1). Higher uptake
of Cl− indirectly causes
membrane depolarization of the macula densa cell: uptake via the
Na-K-2Cl-cotransporter is electroneutral, but basolateral trickling out of Cl− acts depolarizing. Ca2+ influx then leads to the release of paracrine agents, like adenosine or
thromboxane, which trigger contraction of smooth muscle cells in the afferent
arteriole.
Pharmacology cross
reference: by blocking the Na-K-2Cl-cotransporter, furosemide not only blocks the uptake
of Na+ and Cl−,
it also inhibits tubuloglomerular feedback. GFR thus remains high in the face
of sodium chloride losses, which contributes to diuresis. The opposite is true
for thiazide diuretics.
A high protein diet increases glomerular capillary
pressure and glomerular filtration rate. By an unknown mechanism, increased
protein intake enhances reabsorption of NaCl in the thick ascending limb of the
loop of Henle. With less NaCl reaching the macula densa, tubuloglomerular
feedback opens the vas afferens and increases GFR. This mechanism helps to get
rid of incremental nitrogen from amino groups, but increases the risk of
pressure damage to the glomerulus. Thus, a high-protein diet on the margin
increases the risk of renal damage.
Sympathetic
stimulation increases both afferent and efferent resistances. Intense sympathetic
stimulation leads to drastic reductions in both renal blood flow and GFR.
Sympathetic stimulation also increases renin release via β1-receptors.
Renin- angiotensin
II: A decrease
in arterial pressure is sensed by the afferent arteriole, which acts as a
baroreceptor. It directs neighboring granular cells to release renin, to bring
up arterial pressure and to maintain extracellular volume as far as possible. Angiotensin
converting enzyme (ACE), anchored to the plasma membrane of glomerular
endothelial cells, locally produces angiotensin II, which thus constricts the
efferent arteriole more than the afferent one, with the net effect of
maintaining GFR while minimizing loss of volume.
ANP. High extracellular volume is sensed by atrial
myocytes, which release ANP. ANP vasodilates afferent more than efferent arterioles
and lowers the sensitivity of tubuloglomerular feedback. Together, it increases
GFR, lowers osmotic pressure in the medulla and increases diuresis.
Prostaglandins. Prostaglandins are produced by endothelial cells,
vascular smooth muscle cells, mesangial cells and tubule and interstitial cells
in the renal medulla. They are predominantly produced in response to angiotensin
II, sympathetic activation and ADH. Thereby, prostaglandins locally limit the
effects of these vasoconstrictors to maintain blood flow and GFR under stress
conditions. We will review these regulation systems in more detail
later, after considering glomerular filtration and reabsorption of sodium and
water.
The glomerular
filter consists of three components:
1.
The
endothelial layer: endothelial cells are fenestrated, providing no barrier for
solutes; still, a role of the negatively charged glycocalyx in permselectivity is being discussed.
2.
The
basement membrane with lamina rara
interna, lamina densa and lamina rara externa. The basement
membrane is a joint product of endothelial cells and podocytes. It is directly
continuous with the glycocalyx and consists of a dense network of collagen type
IV and proteoglycans. The proteoglycans carry lots of anionic charges, e. g.,
in the form of sialic acid. This decreases the filter's permeability for
negatively charged solutes, e. g., for albumin. The basement membrane prevents
any blood cell to pass the filter.
3.
The
foot processes of podocytes, which are connected by slit diaphragms. The slit
diaphragm
is a dense screen consisting of two major proteins, the long transmembrane
proteins nephrin and neph1, whose cytosolic ends are fixed by the protein
podocin.
Congenital
nephrotic syndrome of the Finnish type: this very rare syndrome is caused by mutations in the
gene encoding nephrin. Massive amounts of protein are filtrated and lost via
the urine. Results are kidney failure, malnutrition and infections, as antibodies
are lost, too. It thus seems likely that the slit diaphragm is the structure
primarily responsible for permselectivity.
Two factors
contribute to the permselectivity of the glomerular filter:
1.
Physical
pore size: with about 8 nanometers in diameter, pore size prevents larger
proteins from passing through. The defining property of a protein in this
respect is its Einstein-Stokes radius, which is the radius of a hard sphere
diffusing at the same velocity as the protein. Globular ("ball-like")
proteins with Einstein-Stokes radii of 4 nm, equaling diameters of 8 nm,
typically have masses around 70 kDa: larger proteins cannot pass the filter.
For example, antibodies (150 kDa and larger) are completely retained, while
isolated immunoglobulin light chains (22 kDa) or their dimers (44 kDa) are able
to pass through.
2.
Charge
selectivity: both basement membrane and slit diaphragm are characterized by
dense arrays of negative charges. As most plasma proteins are also negatively
charged at the physiological pH of 7.4, electrostatic repulsion prevents them
from slipping through pores that spatially would be wide enough to let them
pass. To think of an analogy, you might be able to squeeze through between the
bars of crowd barriers, but if they are freshly painted, you will probably
refrain from trying. Human serum albumin, for example, is a 66 kDa protein. It
is not globular, but rather triangular or heart-shaped, with an effective
diameter of 7 nm, which would allow it to pass through the 8 mm pores. Yet,
albumin is negatively charged, so that less than 1% of all albumin molecules
actually pass the glomerular filter.
Most of this filtrated albumin is reabsorbed by the
proximal tubule. In a healthy person, less than 30 mg of albumin per day is
excreted in the urine. Interestingly, some people, especially young adults,
when standing or sitting in an upright position, excrete somewhat higher
amounts. This phenomenon is termed orthostatic proteinuria and is not
considered pathologic. Apart from that, albumin excretion in the range between
30 and 300 mg per day is called microalbuminuria and constitutes a sensitive
parameter for glomerular damage, e.g., by hypertension or diabetes mellitus.
Glomerular
filtration rate (GFR)
In a 70 kg male young adult, the total filtrated
volume is approximately 180 liters per day, or 125 ml/min. Renal filtration
correlates better with body surface area than with weight. As the surface area
of the standard human being is assumed to be 1.73 m2, normal GFR in males is usually
reported as being 125 ml/min per 1.73 m2. In young females, the respective GFR is
110 ml/min per 1.73 m2.
GFR varies greatly with age: until the age of two,
babies have smaller GFRs due to incomplete development of kidneys. In a young
adult, each kidney contains about 1 million nephrons. A typical feature of
aging is the progressive loss of nephrons, which goes hand in hand with a
gradual decline in GFR.
How to measure GFR? The experimental gold standard is
via inulin clearance. Inulin is a polysaccharide extracted from plants.
Water-soluble and non-toxic, it freely passes the glomerular filter but is
afterwards neither absorbed nor secreted by tubulus cells. If the plasma level
of inulin is kept constant by continuous infusion, the amount of inulin filtered
over a given time equals the amount excreted in the urine over the same time:
inulin plasma concentration x GFR = inulin urine concentration x urine
volume/sampling time
from which GFR is easily calculated as the ratio of
the concentrations times the urine flow.
With normal renal function, the concentration of
inulin in the urine is much higher than in plasma, as all the filtered inulin
is excreted, but almost all of the filtered water is reabsorbed in the tubule
and collecting duct. In other words, virtually the entire filtered volume is
cleared from inulin. Thus, inulin clearance equals GFR.
More generally
speaking, the clearance of a
specific solute is the virtual volume of blood plasma that is cleared from the
solute per unit time via excretion into the urine. Clearance may vary between the
extremes of
·
zero,
e. g. for glucose, which is quantitatively reabsorbed by the tubule and
normally does not appear in urine, and
·
"all",
about 700 ml/min, which is the total volume of plasma that reaches the kidneys,
for substances that are efficiently secreted by the tubules so that all of the
solute brought to the kidney ends up in the urine after a single pass. As we
have seen before, this is true for infused para-aminohippurate, which thus can
be used to measure renal plasma flow.
Determining inulin clearance is an established,
precise method to measure GFR, but measuring inulin concentrations is too
cumbersome for use in clinical routine. Easier to measure is, for example,
Iohexol, a molecule originally used as an X-ray contrast agent, which has very
similar distribution and elimination characteristics.
Even with iohexol and other alternatives, the determination of GFR remains laborious. On the one hand, the substances must be administerd, on the other hand, measurements are cumbersome. For iohexol, what is usually determined is plasma clearance, i. e., it is not the concentration in the urine that is assayed, but rather, in a series of measurements, decreasing plasma concentration. It would be much more convenient to
use a solute that is produced by our organism itself in a steady state way,
that is filtered freely and that is neither reabsorbed nor secreted by the
tubule. In addition, it should be easy to measure. Well, this ideal substance doesn't exist, but creatinine comes close.
Creatinine
clearance as a proxy for GFR
Creatinine is a metabolite of creatine, a liver-synthesized product that
functions as an ultra-short-term energy store in skeletal muscle in the form of
phosphocreatine. Creatinine is continuously released from muscle. It is freely
filtrated, yet also slightly secreted by the tubule. On the other hand, the
usual colorimetric assay tends to overestimate plasma creatinine concentration,
so both effects tend to cancel each other out.
Usually, the patient is asked to collect urine for 24
h and to refrain from eating a lot of meat, which contains creatinine. Three
measurements are required: the volume of
urine, the concentration of creatinine in collected urine and the concentration
of creatinine in plasma. Creatinine clearance can then be calculated as
follows:
Creatinine clearance (ml/min) = urine creatinine /
plasma creatinine x urine volume (ml) / duration of collection (min): usually
1440 for 24 h.
The patient's height and weight may be measured so
that surface area can be calculated and creatinine clearance adjusted to the
standard body surface area of 1.73m2.
It is important to
keep in mind that several factors may impinge on the reliability of the
calculated creatinine clearance:
·
the
patient may have been sloppy in collecting urine, usually at day time (e. g., didn't
want to be seen to go to the bathroom with a giant dark bottle)
·
the
patient nonetheless may have succumbed to the temptation of a steak
·
the
patient may have a health problem affecting skeletal muscle
·
the
patient may have an impairment of liver function. Synthesis of new creatine
occurs mainly in the liver
Serum creatinine
as a measure of renal function
The next step down the ladder of reliability is to throw out urine collection altogether, and to use steady state serum creatinine to assess renal function. In theory, a halving of GFR should lead to a doubling of serum creatinine concentration. In everyday medicine, we use a level of 1 mg/dl as a cutoff: whoever doesn't exceed it is considered to have acceptable renal function. By doing this, however, we miss a considerable percentage of patients who have low actual GFR while still managing to keep serum creatinine levels below the threshold. Typically, this is observed in patients with lower muscle mass, e. g., in delicately built females or in people of advanced age.
To improve on these shortcomings, formulas have been
developed to calculate estimated GFR from serum creatinine measurements by using empirical correction factors, e. g.
for female sex, for age or for African ancestry implying increased muscle mass.
One of these was developed by the Modification of Diet in Renal Disease Study
Group and thus termed MDRD-formula:
eGFR (ml/min)= 186 x serum creatinine (mg/dl)-1.154 x age-0.203 x [0.742 if female] x [1.210 if black]
Parameters like eGFR and albumin-to-creatinine ratio
in urine (ACR) contribute to prediction of renal outcomes, as well as to prediction
of vascular outcomes beyond traditional risk factors.
4. REGULATION OF SODIUM AND EXTRACELLULAR VOLUME
In the extracellular fluid compartment of our body, which accounts for a little more than a third of its total fluid, we carry a lot of Na+ with us. Life on our planet originated in the sea, and in our extracellular volume, we carry a diluted form (from 3.5% to 0.9% salinity, or from about 480 mmol/l Na+ to 140 mmol/l and from about 550 mmol/l Cl− to 110 mmol/l) of "inner" sea with us, which we need to maintain carefully. About 65% of the total Na+ in our body is located in the extracellular fluid. An additional 5-10% is in the ICF compartment. Together, these 70-75% of the total sodium pool of our body constitute the "exchangeable" sodium stores. The word exchangeable stems from experiments with radioactive Na+, which rapidly equilibrates with this part of our total body Na+. The rest is non-exchangeable, meaning osmotically inactive, at least in the short term, and bound in some form, mostly in skin, muscle and bone. Non-exchangeable Na+ has been shown in the
skin, where it seems to be bound to glycosaminoglycans. There, it is not
completely osmotically inactive: it seems to be "locked away" in a
hyperosmotic compartment, the accessibility of which is modulated by the
density of lymphatic vessels, which in turn is regulated by macrophages via
VEGF-C (vascular endothelial growth factor-C). Prolongued intake of a high-salt
diet may over time increase these stores, which increase with age and are
higher in hypertensive than in normotensive persons. Our knowledge about the pathophysiological
significance of these stores is quite inadequate; experimental data suggest
that over a longer time frame, these stores may still act as an additional
sodium buffer. This way, they may contribute to salt-sensitive hypertension. In
tightly controlled space simulation experiments with fixed NaCl input over
months, periodic oscillations in sodium storage of 200-400 mmoles were
observed, with rhythms of weekly and monthly or longer periodicity. Thus, while
sodium balance is by and large maintained (input equals output), this is not strictly
true for specific 24-hour periods.
One gram of salt contains 17 mmol of NaCl. If we take
up 140 mmol (8.2 g) of NaCl, for osmotic reasons, we expand our inner sea, our
extracellular volume, by roughly 1 liter. Today, on our Western Diet, we take
up a lot of salt. But that has not always been the case.
Until a few thousand years ago, maintaining our inner
sea was a challenge, because Na+ was quite scarce in human
nutrition. Thus, our organism is optimized for salt conservation. For millions
of years, the human organism confronted the following problem: Our "inner
sea" consists mainly of water and salt; while water is a sine qua non and had to be readily
available to fill up, salt was not. The human body needed to excrete a lot of
water-soluble substances (like urea), yet retain the salt.
This problem is solved by our kidneys. Every single
day, we filtrate up to 180 liters of "inner sea" containing (180x140
mmol) 25 mol salt. This is about 1.5 kg of salt, while our daily intake
contains only a few grams! There is only one option: we need to retrieve 99.5%
of all filtrated salt. This is achieved in steps: 67% of filtrated sodium is
reabsorbed in the proximal tubule, mainly in exchange for H+.
Twenty-five percent is reabsorbed in the thick ascending limb of the loop of
Henle by cotransport of Na-K-2Cl. In the distal convoluted tube, a further 5%
are absorbed by Na-Cl-cotransport. Actual Na+-excretion is fine-tuned
by the renin-angiotensin-aldosterone system, which controls reabsorption of the
last 0-3% in the collecting duct via the epithelial Na+ channel (ENaC).
Let's take a look at the individual segments:
·
In
the proximal tubule, there is a huge gradient between the 140 mM Na+ in the primary filtrate and the cytoplasm of the tubule cell, which is
continuously evacuated from Na+ by the powerful basolateral
Na-K-ATPase. In addition, there is the cell-interior negative electrical
potential generated by more Na+ leaving the cell than K+ entering. This powerful combined electrochemical gradient produces an inward Na+ cascade that supplies the energy to cotransport solutes like glucose, amino
acids, phosphate, citrate etc. out of the filtrate into the cell. Reabsorption
of all these valuable solutes is enabled simply by inserting the respective cotransporters
("mills") into the apical plasma membrane. From a quantitative
viewpoint, however, a second mechanism is even more important: the Na+ cascade's energy is used to extrude H+ from the cell via the
Na-H-exchanger (NHE), with the ultimate goal of HCO3− recovery. The speed of this exchange depends on the rate of intracellular proton
generation. H+ and HCO3− are generated
by carbonic anhydrase II; HCO3− and Na+ leave the cell basolaterally to the blood. Numerous aquaporin units in the
membranes of proximal tubule cells allow water to freely follow sodium,
resulting in isotonic reabsorption. Apical inflow of Na+ into the
cell and basolateral outflow of K+ to the blood create a
lumen-negative transepithelial voltage (caution: don't confuse this with the
transmembrane voltage!), causing some of the transported Na+ to leak
back into the lumen by the paracellular pathway.
Pharmacology cross reference: blocking carboanhydrase (by acetazolamide) reduces
the amount of H+ available to exchange against Na+. While
at first glance that would seem to produce a huge diuretic effect, compensatory
Na+ uptake in later nephron segments reduces it to a rather mild end
result. As a diuretic, acetazolamide is obsolete, yet it continues to be used
for glaucoma and other indications.
·
In
the thick ascending limb, Na+ and Cl− gradients
drive uptake into the cell via the Na‑K‑2Cl cotransporter. In this
process, the limiting factor is K+, which is low in extracellular
fluid and has to be dragged in by its partners against its concentration
gradient. In order to make more of it available, K+ is recycled from
the cell via an unusually high number of apical K+-channels (ROMK, renal outer medullary K+-channel). The
resulting strong apical outflow of K+ leads to a lumen-positive transtubular
voltage, which is the driving force for additional, paracellular Na+-flow.
About 50% of Na+ reuptake in the thick ascending limb is due to this
passive transport. Of course, the same mechanism also drives paracellular
reuptake of other cations such as K+, Ca2+ and Mg2+.
An important feature of the thick ascending limb is a remarkably low water
permeability of its apical membrane, which is due to the selection of
transmembrane proteins expressed on it (e.g., no aquaporins); tight junctions
between cells are water-impermeable, too. These tight junctions contain
specialized claudins which manage to let pass Na+ (claudin 10b), Ca2+ and Mg2+ (claudins 16 and 19), but hardly any water. Na+ is transported out, water stays in: this leads to a diluted, hypoosmolar fluid
at the end of this "dilution segment", while interstitial osmolality
is built up. Interstitial osmolality may be raised up to 200 mOsm above the
level of the adjacent tubule content.
Pharmacology cross reference: Loop diuretics (furosemide) block the Na-K-2Cl cotransporter
by competing for the chloride site, producing a strong diuretic effect.
Therefore, a lot of Na+ and Cl− arrive at the macula densa at the end of this segment.
Normally, that would induce a massive reduction of GFR via tubuloglomerular
feedback. Yet, such reduction would be mediated by Cl− uptake into macula densa cells via precisely the
Na-K-2Cl cotransporter that is being blocked by the drug. Thus, there is no reduction of GFR, which contributes
to the diuretic effect. Of course, this also reduces the passive, paracellular
reabsorption of other cations and may cause considerable losses of Ca2+ and Mg2+. As we noted previously, hypocalcemia increases the open probability of Na+ channels in neurons and muscle cells and may lead to cramps. Via the same
mechanism, loop diuretics may induce secondary hyperparathyroidism. The other
way round: loop diuretics may be used in the treatment of hypercalcemia.
Obviously, downstream nephron segments try to compensate for the looming Na+ loss, causing increased excretion of K+. The mechanism of this
"exchange" is explained below, in the subsection on aldosterone.
·
In
the distal convoluted tubule, the diluting effect has taken Na+ concentration down to about 70 mM. That is still more than 5 times the
intracellular level. Na+ enters the cell via the Na‑Cl cotransporter
and is pumped out the back door via Na-K-ATPase.
Pharmacology cross reference: Thiazide diuretics, like hydrochlorothiazide, block
the Na‑Cl cotransporter. Here again, distal attempts to make up for Na+ losses lead to increased excretion of K+.
·
In
the collecting duct, Na+ enters tubule
cells via epithelial Na+ channels (ENaC), which are under control of
aldosterone and ANP. Aldosterone increases the number of open sodium channels;
ANP decreases it. Importantly, the inflow via ENaC is electrogenic, as Na+ enters without an anion partner. This is different from the electroneutral
uptake in the previous segments and leads to an indirect exchange with K+,
as we will see in a minute in the aldosterone subsection.
Pharmacology cross reference: Amiloride and triamterene block ENaC; spironolactone
and eplerenone block the mineralocorticoid (aldosterone) receptor. The effect of these
diuretics is mild, as they only affect the last 3% of Na+ left in
the lumen. These drugs do not lead to losses of K+; to the contrary,
they may cause K+ retention.
Atrial natriuretic peptide (ANP) is released from myocardial cells of the atria in response to volume expansion, which is sensed as atrial stretch. It is a 28-amino acid polypeptide binding to a receptor with guanylate cyclase activity so that its second messenger is cGMP. ANP has renal effects and systemic effects:
1.
In
the kidney, it increases excretion of Na and water:
-ANP closes apical Na+ channels (ENaC),
reducing Na+ reabsorption.
-It increases GFR and filtration fraction by dilating
the Vas afferens more than the Vas efferens. As a result, it also increases
blood flow to cortex and medulla, and tends to wash out the osmotic gradient in
the renal medulla. In addition, it reduces the sensitivity of the
glomerulotubular feedback.
2.
Only
at higher concentrations ANP acts as a direct vasodilator in the systemic
circulation, lowering arterial blood pressure.
This second effect limits the first, as the reduction
in blood pressure negatively affects GFR and with that, the excretion of Na+ and water.
The effect of ANP in volume regulation depends on
conditions:
The effect of the
Renin-Angiotensin-Aldosterone system on extracellular volume
A decrease in effective circulating volume increases renin
secretion in three ways:
1.
Deactivation
of stretch receptors in the afferent arteriole directly leads to renin
secretion.
2.
Deactivation
of stretch receptors in the carotid and the aortic arch result in enhanced
sympathetic signaling to the juxtaglomerular apparatus via adrenergic β1-receptors.
3.
The
mechanism mediating tubuloglomerular feedback affects renin secretion as
well. As we noted before, delivery of
more NaCl to the macula densa leads
to a reduction of GFR. The other way round, if less NaCl is arriving at the
macula densa, this is sensed via reduced uptake of Cl− by the
Na‑K‑2Cl cotransporter and somehow translated into an increase in
renin release. It is not yet clear how the message travels from macula densa to adjacent granular cells;
possibly via alterations in the rate of prostaglandin production.
However, we should keep in mind that renin release in
response to increased sympathetic activation may also occur in the absence of a
decrease in effective circulating volume. In this case, the result is an
increase in circulatory volume, which may play an important role in the
development of arterial hypertension.
Renin, a protease released into the vas afferens,
excises the decapeptide angiotensin I from the α2 plasma globulin
angiotensinogen, which is continuously produced by the liver. Angiotensin I is
converted to the octapeptide angiotensin II by angiotensin converting enzyme
(ACE) via removal of two amino acids. ACE is expressed by endothelial cells, at
high levels in lung and glomerular capillaries, and exposed on their luminal
surface. The kidney receives systemic
angiotensin II and produces additional angiotensin II on the endothelial
surface of the glomeruli. The result is that angiotensin II concentrations at
the vas efferens are higher than elsewhere, and that vasoconstriction affects
the vas efferens more than the vas afferens.
The majority of angiotensin II's effects are mediated
via the AT1 receptor:
Together, these mechanisms contribute to maintain and
fill up extracellular volume.
Aldosteron has two main functions: firstly, it is our
Na+ saving hormone, secondly, it is required to excrete K+ excess. Consequently, aldosterone is activated by angiotensin II as well as by
an increase in plasma K+ concentration.
Aldosterone has a
threefold effect on principal cells of the collecting tubule:
1.
it
induces Na-K-ATPase. An increase in activity over and above levels in other
cells is necessary, as Na+ entry via ENaC is passive and works only
as long as there is a concentration gradient. In cases where urine Na+ approaches its lowest possible value of about 3 mM, intracellular Na+ has to be lower.
2.
it
increases the surface area of the basolateral membrane to accommodate more
units of Na-K-ATPase
3.
it
induces the apical ENaC (epithelial Na+ channel)
Indirect exchange
of Na+ for K+:
Aldosterone makes more Na+ cross the apical
membrane. Inflow of these positive charges affects membrane potential. Membrane
potential is primarily a function of K+ trickling out. The outflow
of K+ limits itself, as accumulation of positive charges on the
outside and negative charges on the inside stop the flow electrostatically at a
certain level of polarization. Incoming Na+ reduces this voltage,
allowing more K+ out. The more Na+ is ushered in by
aldosterone, the higher net secretion of K+. If Na+ reabsorption in upstream tubulus segments is inhibited by diuretics such as
furosemide or thiazide, high amounts of K+ are excreted with the
urine. In contrast, if little Na+ is left in the fluid reaching the
collecting tubule, K+ secretion is low. Therefore, the ratio is also
flow-dependent: at low tubular flow, a larger proportion of Na+ is
taken up in upstream segments together with Cl−, and a lower
proportion is exchanged for K+.
Aldosterone also has a Na+-retaining effect in other organs: in the intestinal epithelium, especially in the colon, in the sweat glands and in the placenta. In cooperation with Angiotensin II, it also promotes salt hunger in the CNS. The corresponding aldosterone responsive neurons are located in the nucleus tractus solitarii in the medulla oblongata.
Physiologically, glucocorticoids increase the GFR and
decrease water permeability of the distal nephron. Thereby, they increase flow
and the amount of Na+ reaching the collecting duct, which enhances K+ secretion.
Cortisol and aldosterone are quite similar. While in
principle, each of them has its own receptor, both hormones bind to both
receptors. In cells that ought to respond to aldosterone, this causes a
problem: cortisol, with its much higher molar concentration, would bind and
activate the mineralocorticoid receptor, causing an unregulated all-out
response. To prevent that, these cells express the enzyme
11β-hydroxysteroid-dehydrogenase 2(11β-HSD2), which converts cortisol
to cortisone. Cortisone does not bind to the mineralocorticoid receptor (nor
does it bind to the glucocorticoid receptor).
Pharmacology cross
reference: When used as drugs, glucocorticoids reach far higher molar concentrations than
in the physiological situation. The sheer number of glucocorticoid molecules
overwhelms the capacity of 11β-HSD2 to protect the mineralocorticoid
receptor. Thus, the higher the dose of glucocorticoids, the higher the
proportion binding to the mineralocorticoid receptor and the more pronounced
the aldosterone-like effects, producing, e.g., arterial hypertension.
Glycyrrhizin, a constituent extracted from the liquorice root, inhibits 11β-HSD2,
increasing the mineralocorticoid effect of glucocorticoids. Depending on the
ingested dose, it may cause Na+ and water retention by its own.
ADH (anti-diuretic hormone = AVP, arginine
vasopressin), a nonapeptide, is synthesized in neurons of the Nucleus supraopticus and the Nucleus paraventricularis of the
hypothalamus. Neurons of these nuclei are stimulated by osmoreceptor neurons located
still outside the blood-brain barrier in two circumventricular organs: the Organum vasculosum laminae terminalis and the subfornical organ. The firing rate of these osmoreceptor
neurons is influenced by mechanosensitive cation channels that are able to
sense osmotic swelling or shrinking.
Separate, but parallel osmoreceptor-neurons in the
same organs also project to hypothalamic regions generating thirst.
On rises in plasma osmolality, ADH is released in incremental amounts from the posterior lobe of the pituitary. Normally, no ADH is secreted below 280 mOsm. However, there is an exception: in case of plasma volume depletion, ADH may be secreted even at low plasma osmolality. In this case, the stimulus for ADH release is originated by high-pressure baroreceptors in the carotid sinus and aortic arch and by low-pressure baroreceptors in the right and left atria. ADH release is much more sensitive to osmoregulation than to volume regulation. Increases of 1-2% in plasma osmolality will stimulate release of ADH, while a 10-15% reduction in blood volume or pressure is needed. However, once this threshold is reached, further response to volume depletion is exponential. Since ADH is a product of the hypothalamus, central
nervous stimuli also influence its release. In slightly dehydrated mice, the
sensory perception of available water within seconds reduces the release of
ADH, before any change in the increased osmolality occurs. In humans, pain,
nausea and vomiting, IL‑6 release, elevated body temperature as well as
hypoglycemia have been reported to increase ADH release. ADH therefore not only
reacts to osmolality, but more generally is part of the organism's acute stress
response ("In the event of danger, we better hold back our water
reserves").
The hormone has a short half-life of about 20 minutes.
Depending on initial levels, the maximum diuresis after a water load is
therefore delayed for one to two hours.
Regular effects via the V2 receptor:
·
At
the distal nephron, ADH activates AVPR2, also known as V2-receptor, a Gs-coupled
receptor signaling via cAMP. PKA-mediated protein phosphorylation causes fusion
of aquaporin-2 water channel-containing vesicles with the apical membrane, increasing
water permeability of the cortical collecting tubule as well as outer and inner
medullary collecting ducts. Thus, in the distal nephron, osmotic resorption of
water may be increased more than tenfold. Longer term stimulation by ADH
induces aquaporin‑2 gene expression, leading to higher numbers of protein
units per cell.
·
In
addition, ADH increases urea permeability only at the inner medullary
collecting duct by adding and phosphorylating units of UT-A (urea transporter-A, SLC14A2). This second
function is crucial for urea recycling required to build peak osmotic pressure
in the antidiuresis situation. Under conditions of maximal antidiuresis, urine
is isotonic with the inner medulla of the kidney. Thus, the degree of
hypertonicity of the medullary interstitium determines maximum urine
concentration.
·
ADH
increases NaCl reabsorption in the thick ascending limb by stimulating Na‑K‑2Cl
cotransport.
Stress-related effects via the V1 receptor:
·
Only
at maximal secretion in acute stress situations does the molecule actually act
as "vasopressin", i.e. lead to contraction of smooth muscle cells in
arteriolar walls via AVPR1a (V1a receptor).
·
Another
renal effect of ADH via the V1 receptor is to stimulate production of
prostaglandins in many cell types, including the glomerular mesangium and the
tubulus. The prostaglandins promote vasodilation and in all likelihood are
important to maintain GFR and renal perfusion in the face of elevated ADH
levels and antidiuresis.
Pharmacology cross reference: the widespread use of NSAIDS has the unwanted side
effect of suspending this safeguard mechanism.
·
V1a
receptors are also expressed by hepatocytes and platelets, promoting
gluconeogenesis and clotting readiness.
·
Via
AVPR1b (V1b) receptors on corticotropes in the pituitary, ADH/AVP promotes the
release of ACTH.
Pharmacology cross reference:
·
Stimulants
of ADH release may be nicotine and drugs like morphine or barbiturates.
·
Inhibitors
of ADH release are alcohol and opiate-antagonists.
Diabetes insipidus ("unquenchable flow-through") is the result
of failing ADH function, which may be caused either by lack of ADH itself
(central diabetes insipidus) or by a
failure of the kidney to respond (nephrogenic diabetes insipidus). As the latter parts of the nephron remain
impermeable to water, the patient produces large amounts of diluted urine
(polyuria). The loss of water tends to result in hypovolemia, making patients
thirsty and inducing them to drink large amounts (polydipsia). If patients do
not drink enough, they quickly develop hypotension, marked hypernatremia and
shock. Central diabetes insipidus may
be treated by nasal administration of the synthetic ADH analog desmopressin or DDAVP
(1 Desamino-8-D-Arginine-Vasopressin).
Pharmacology cross
reference: Nephrogenic diabetes insipidus may be
an unwanted response to certain drugs, e. g., colchicine and Li+.
Lithium therapy is used to treat bipolar disorder. Li+-ions decrease
cAMP, the second messenger of ADH, by inhibiting adenylyl cyclase.
The syndrome of inappropriate ADH secretion (SIADH) is
the opposite of diabetes insipidus, with inappropriately high levels of ADH in
relation to osmolality. Full-fledged water resorption leads to volume expansion
with ANP release. Water retention combined with excretion of concentrated urine
containing relatively high amounts of Na+ may result in critical hyponatremia.
Circumstances under which SIADH may occur:
·
Certain
malignant tumors may produce ADH or ADH-like peptides, e. g., small cell lung
carcinomas.
·
CNS
disorders, including traumatic brain injury, meningitis or encephalitis.
·
Pulmonary
diseases such as pneumonia, tuberculosis.
·
SIADH
is sometimes seen in the course of surgical procedures, probably via pain
afferents. Most likely, this is abetted by genetic factors, although we don't
yet know the responsible genes and alleles.
·
SIADH
is likely to be part of the problem in exercise-associated hyponatremia, which
is caused by overdrinking during endurance sports. Normally, overdrinking
should just result in production of more urine. Yet, most probably again on the
basis of genetic factors, under stress of competition some individuals retain the fluid, develop hyponatremia
and, in extreme cases, exercise-associated hyponatremic encephalopathy, a form
of cerebral edema. As in the case of surgical procedures, ADH secretion is due
to non-osmotic stimuli. Concomitant use of NSAIDs to alleviate aches and pain
may compound the problem by reducing GFR and renal perfusion. At the end of the
2002 Boston Marathon, 13% of surveyed participants were reported to be
hyponatremic.
·
Pharmacology cross reference: Likewise, a long list of drugs may increase
production of ADH (central SIADH) or sensitize renal tubules to ADH (renal
SIADH). SIADH may occur in patients receiving opioids, tricyclic
antidepressants or chemotherapy containing cyclophosphamide or vincristine.
The same pattern may develop in two endocrine deficiency syndromes: hypothyroidism and cortisol deficiency (primary or secondary, verifiable with an ACTH stimulation test); cortisol has a mild inhibitory effect on ADH secretion in the physiological concentration range. Since the excess of ADH in these two conditions is secondary and can be cured by hormone replacement, they are usually not categorized as "SIADH". Pregnancy: During pregnancy, ADH release and thirst are
stimulated already at lower osmolalities to satisfy the need for increased
blood volume. Thus, pregnancy is often associated with an 8 to 10 mosmol
decrease in plasma osmolality. A similar but smaller change may occur in the
progesterone-dominated late phase of the menstrual cycle, resulting in slightly
increased body weight.
Pharmacology cross
reference: The effect of ADH may be blocked by V2 receptor antagonists like Tolvaptan.
Causes. In theory, hyponatremia may result either from loss of
sodium or from retention of water. While we may lose considerable amounts of
sodium by vomiting, diarrhea or sweating, Na+ concentration in these fluids is almost always lower
than in plasma. Thus, we lose more water than Na+ which tends to increase plasma Na+ concentration. Consequently, hyponatremia is only
seen in cases of water retention leading to an excess of water in relation to
Na+. As long as everything works according to plan, we have an enormous
capacity to excrete water by completely shutting down ADH. In the absence of
ADH, urine osmolality can fall to 40 mosmol/kg, about one-seventh of the value
in plasma (290 mosmol/kg). Therefore, water retention resulting in
hyponatremia occurs only when there is an inability to suppress the secretion
of ADH. An exception to this rule occurs only in patients who, due to mental illness, consume so much
fluid that they exceed the water elimination capacity at
zero ADH.
Persistent ADH
release in the face of reduced osmolality is seen in:
1.
Effective
circulatory volume depletion
·
Hepatic
cirrhosis
·
Use
of thiazide diuretics, in combination with the next point:
·
electrolyte-containing
fluid (renal, gastrointestinal, skin) losses partially replaced by water
2.
SIADH,
as discussed above (paraneoplastic, CNS disease, tuberculosis, surgery,
overdrinking in endurance sports, as a side effect of medication)
3.
Adrenocortical
insufficiency (cortisol deficiency, salt loss due to mineralocorticoid
deficiency)
4.
Hypothyroidism
Some patients with asymptomatic mild hyponatremia have
a reset osmostat. In these patients, the threshold for ADH release is reduced
below the normal osmolality of 280 mosmol/kg, so that a lower level of
plasma Na+ is maintained. In these patients, hyponatremia cannot and
should not be treated, as these patients would only increase ADH release and
develop thirst on attempts to elevate their plasma sodium concentration.
Symptoms. Hyponatremia is critically more dangerous if it
develops acutely than if it develops chronically. With acute hyponatremia,
plasma osmolality falls, and water starts to move inside cells. Cells start to
swell by osmotically taking up water. For most cells, this is no big problem,
yet it is for brain cells, which are encased in a rigid skull. The brain can accommodate
a maximum increase of 7-8% in volume by shifting fluid from ventricles out of
the skull, but then starts to herniate through the Foramen magnum at the base of the skull. The patient may develop
headaches, nausea, vomiting, confusion and eventually, seizures and coma. In
the worst case, brain stem compression may lead to respiratory arrest and
death.
If hyponatremia develops slowly and chronically, on
the other hand, brain cells adapt by releasing osmotically active substances.
These comprise K+ and Na+, but also organic solutes, e.
g., myoinositol, choline compounds, glutamine and glutamate. Therefore, chronic
hyponatremia typically produces no symptoms for a long time.
When diagnosing
and correcting hyponatremia, it is essential to differentiate between the two
states:
·
Acute hyponatremia
in an overhydrated marathon runner may be safely and rapidly corrected by
administration of hypertonic saline. The SIADH component requires that Na+ concentration of the infusion be higher than the Na+ concentration
in the excreted urine.
· Were the same treatment administered to a patient with asymptomatic chronic hyponatremia, the rapid increase in extracellular osmolality would induce a massive shift of water out of the already adapted brain cells and potentially produce an osmotic demyelination syndrome with irreversible neurologic damage (central pontine myelinolysis). In chronic hyponatremia, correction needs to be performed with extreme caution: very slowly and only partially at first.
Causes. Any increase in osmolality beyond normal levels
induces a strong sensation of thirst. Hypernatremia is thus usually seen in
situations involving unreplaced water losses where thirst is either impaired or
cannot be quenched.
Water losses may
be due to:
1.
Sweating
and insensible losses via skin and respiration: people in the desert or on
boats at sea; patients with fever and infections
2.
Urinary
losses due to diabetes insipidus or osmotic diuresis due to hyperglycemia
3.
Gastrointestinal
losses
Symptoms. As with hyponatremia, symptoms are predominantly seen
when the condition develops rapidly and include lethargy, seizures and
ultimately, coma. In chronic hypernatremia, elevated levels of Na+ may be reached in the absence of symptoms other than thirst.
Hypernatremia is commonly seen in elderly patients in
nursing homes suffering from an infection, especially if their mobility or
mental status had already been reduced to begin with. Alternatively,
hypernatremia may develop in infants and toddlers yet too small to help
themselves or unable to express themselves. Think of children left in car seats
in the heat.
Hypo- and hypernatremia may also be seen in patients
suffering from diabetes mellitus. Hyperglycemia
causes osmolality to increase, leading to osmotic water movement out of cells.
In addition, increased osmolality leads to the sensation of thirst, causing the
patient to drink more water. Both effects contribute to lowering plasma Na+ by dilution. The longer this state of hyperglycemia is maintained, the more
water is lost due to osmotic diuresis. Yet, Na+ concentration in the
urine is well below plasma concentration: proportionally, more water is lost
than Na+. Treatment with insulin will correct hyponatremia by
causing water (with glucose and potassium!) to shift back into cells, unmasking
the previous relative accumulation of Na+. Plasma Na+ may
now climb through normal values and temporarily reach levels of hypernatremia.
The ability to
concentrate urine is inextricably linked with conditions that pose a danger to
the medulla
Like a penguin is able to regulably separate its
warmth (40°C!) from the coldness in its feet by a counter-current procedure,
our kidneys are able to regulably build a steep osmotic gradient by an
osmolality counter-current procedure. This gradient is the basis of our
kidneys' ability to concentrate urine. Like the system allows the penguin in
the cold to lose little warmth to the environment, it allows us to lose little
water to the environment if water is scarce.
The kidney's ability to concentrate urine depends on
its handling of NaCl and of urea. Of these two, urea is more variable. Urea is
produced by the liver when metabolizing amino acids. The higher the dietary
protein content, the greater the kidney's concentrating ability. In the deepest
portion of the medulla next to the papilla, interstitial urea contributes about
half of total osmolality. Additional solutes that sometimes have to be excreted
at high concentrations are integrated into the gradient.
The kidneys keep our organism's osmolality constant by
adjusting excretion of osmotically active solute particles. They are able to
produce a wide range of urine concentrations, from very concentrated to
extremely dilute. A given urine volume may be thought of consisting of two
parts: part 1 is the volume required to dissolve all the excreted solutes at
the same osmolality as in plasma (290 mosmol/l). Part two is the volume of pure
or solute-free water that has to be added or subtracted to arrive at the given
urine volume. In case urine osmolality is lower than that of plasma, this free
water volume is positive; if urine osmolality is higher, free water volume is
negative. Excreted volume of free water per unit time is called (solute-) free
water clearance.
With a normal diet, our daily production of excreted
solutes amounts to about 600 mosmol. Dissolved in an average urine volume of
1.5 l, they result in urine osmolality of 400 mosmol/kg, which is a little more
concentrated than the 290 mosmol/kg of plasma. Free water clearance that day
would be negative at about -0.5 l, because isoosmotic excretion of 600 mosmol
would require a little more than 2 liters.
A patient with diabetes
insipidus produces extremely diluted urine around 40 mosmol/kg. At that
dilution, the patient excretes 600 : 40=15 liters of urine. This is the amount
he also has to drink to prevent hypovolemia. Free water clearance of that day
would be +13 liters.
If we get lost in the desert, we need to concentrate
the 600 mosmol in the smallest volume possible. Our young kidney is able to
push up osmolality to 1300 mosmol/kg. That way, the 600 mosmol can be excreted in a little less than half a
liter of urine. Free water clearance per day is -1.6 liters.
Urine specific gravity is easily measured, allowing a
rough estimate about its concentration and osmolality:
1010 g/l: corresponds to the 290 mosmol/ kg of our
body
1015-1022 g/l: a frequent range, with slightly
concentrated urine
1001 g/l: extremely dilute, around 40 mosmol/kg
1040 g/l: extremely concentrated, about 1300 mosmol/kg
If extremely concentrated urine at osmolalities up to
1300 mosmol/kg is excreted, of course, all solutes contribute to this enormous
osmolality according to their individual concentrations: Na+, Cl−,
K+, urea, NH4+, phosphate, etc.
Why is it
impossible to prevent dying from thirst by drinking sea water?
In young healthy adults, the people with "optimal
kidneys", maximum measured concentration ability for Na+ in
urine is about 270 mmol/l, with individual maxima varying between 240 and 295
mmol/l. Na+ concentration of sea water is about 480 mmol/l. Thus,
drinking sea water would only worsen the situation. Here, we assess the problem using the more
familiar numbers for Na+, yet the result is even more valid for Cl−.
To provide a zone with extreme osmolality in the
kidney, it is necessary that this zone be perfused as sparingly as possible to
prevent the otherwise inevitable washout. Of course, this entails the danger of
hypoxia, and with prolonged hypoxia, the danger of necrosis. The papilla is
therefore the most endangered region of the kidney once additional
cell-stressing factors coincide. These additional stress factors may be
remembered by the mnemonic POSTCARDS:
Pyelonephritis
Obstruction of the urogenital tract
Sickle cell disease
Tuberculosis
Chronic liver disease
Analgesia or alcohol abuse
Renal transplant rejection
Diabetes mellitus
Systemic vasculitis
How to protect one's kidneys? Just by drinking
sufficiently! If it's not necessary to concentrate urine, perfusion of renal
medulla is higher and oxygen supply is better.
Starling's law states that net filtration is
proportional to the permeability of the capillary wall, the surface area
available for filtration, and the differential between hydraulic pressures in
capillary and interstitium minus the differential in oncotic pressures. The
differential in oncotic pressures is codetermined by the reflection coefficient
of proteins across the capillary wall, σ. How much of this filtrated
volume remains in the interstitium is determined by the rate of fluid removal
via the lymphatics.
Thus, edema may
result from:
1.
Increased
capillary pressure
2.
Decreased
oncotic pressure differential
3.
Increased
capillary permeability
4.
Lymphatic
obstruction
Each of these conditions causes a reduction in plasma
volume, which is corrected by the kidneys by retaining Na+ and water
until plasma volume is normalized. In other words, in case of edema, renal
retention of Na+ and water is an appropriate compensation to
normalize perfusion of all organs of the patient's body. We may feel that we
would like to remove the underlying edema by treating the patient with a
diuretic, yet we have to keep in mind that this may diminish tissue perfusion
in parts of her organism.
Edema does not become apparent until the interstitial
volume has increased by at least 2.5 l. The only way to recognize it earlier on
is by weighing the patient on a daily basis.
Pharmacology cross
reference: Classes
of drugs known to sometimes promote sodium and water retention include
glucocorticoids, non-steroidal anti-inflammatory drugs (NSAIDS), Ca2+-antagonists
and thiazolidinediones. For glucocorticoids, the effect is due to activation of
the mineralocorticoid receptor. Prostaglandins not only maintain perfusion and GFR under
stress, they also have a slight tubular effect promoting excretion of Na+ and water; NSAIDs counter this effect. Ca2+-antagonists increase capillary filtration pressure,
because they dilate arterioles more than venoles; in the kidney, they tend to promote
diuresis. For Thiazolidinediones (glitazones), which have been used to treat
patients with diabetes mellitus type
2, the exact mechanism is not known. It has been shown that steady state levels
of several proteins involved in renal sodium and water transport are increased.
Note that thiazolidinediones are PPARγ agonists. Activated by endogenous
lipids like fatty acids, PPARγ is a nuclear receptor regulating expression
of a large number of genes. Obviously, some of these gene products affect
handling of Na+ and water.
5. DISORDERS OF POTASSIUM HOMEOSTASIS
Disorders of potassium balance may occur by losses,
retention or excessive administration of potassium or, rarely, by a very low-potassium
diet over a long time. In addition, imbalances may occur by potassium shifts
between intra- and extracellular volume.
While exact regulation of Na+ is required
for its importance in determining extracellular volume, exact regulation of K+ is essential for its role in membrane excitability. Membrane potential is
foremost a K+-potential, because at equilibrium, membrane permeability
it highest for K+, which trickles out of the cell following its
concentration gradient. High intracellular K concentration is maintained by
Na-K-ATPase, which is not simply chugging along, but rather modified in its
activity by external factors. Insulin, epinephrine via β2 receptors
and aldosterone make it pump harder. Acidosis reduces the pump's activity.
Pondering this, we realize that we have to expect shifts in potassium once we
use drugs like insulin, beta-blockers, β2-adrenergic agonists or
aldosterone antagonists.
With a potassium-rich meal, we take up a high load of
K+, while K+ concentration in blood is low and needs to
be kept strictly within a narrow range. How do we manage to do that? Short-term,
the K+ taken up is shoveled into cells. This is strongly assisted by
insulin, which increases following a meal. Insulin is secreted into portal
blood, reaches the liver at high concentrations and helps to move a large part
of the K+ load into hepatocytes. Plasma K+ increases a
little, yet after an hour, most of it has vanished into cells. Longer term, the
kidney is able to smooth over the strong ups and downs of potassium intake with
food. This is done via the collecting tubule, which is able to secrete large
amounts of potassium via the ROMK channel of principal cells, or to reabsorb almost all K+ by α‑intercalated cells. A healthy kidney has an enormous K+-excreting
capacity.
In response to prolonged nutritional K+ deprivation, the apical membrane of intercalated cells, which contains H+‑
K+‑ATPase pumping in K+ from the lumen in exchange
for protons, gets amplified much like in parietal cells of the stomach.
Plasma K+ concentration may also be
perturbed by intense exercise. In a fast sprint, we use almost all of our
muscles. At depolarization, Na+ streams into muscle cells, at
repolarization, K+ pours out. Concurrent sympathetic activation
guards against an excessive increase in plasma K by prompting other cells,
e.g., hepatocytes or cells of inactive muscles, to take up more of it. Once the
sprint is over, epinephrine still remains active for a while. In summary, K+ tends to increase during strenuous exercise, but tends to decrease following
cessation.
Regarding the effects of changes in extracellular K+,
we may reflect as follows:
Extracellular hypokalemia should expedite the flow of
K+ out of cells, at least, if this change occurs within a short time
frame. This increases membrane polarization. With membrane potential farther
from the threshold, it is harder to trigger an action potential.
Extracellular hyperkalemia should impede K outflow and
reduce membrane polarization. That should facilitate triggering action
potentials in neurons and muscle cells. Unfortunately, it's not as simple as
that. As we have seen in cardiac pacemaker cells, at low polarization, Na+ channels
sometimes fail to regenerate after an action potential, reducing the
probability of further action potentials.
At this point in our considerations, we give up and
content ourselves with the general assessment that deviations of plasma K
concentration in both directions may cause arrhythmia and weakness in skeletal
muscle.
Acidosis leads to hyperkalemia. Low pH inhibits Na-K-ATPase,
leaving more K outside in extracellular space. Inside cells, increased protons
displace K ions from proteins so that more K+ exits from cells.
Likely, these effects are most prominent in the large volume of muscle tissue.
In addition, acidosis reduces renal excretion of K+ in spite of
increased plasma concentrations: acidosis reduces the open-probability of the
apical ROMK-channel of principal cells.
Alkalosis leads to hypokalemia. This is mainly due to
uptake of K+ into cells, but in addition, more of this intracellularly elevated
K+ is secreted into the tubulus by cells of the collecting tubule.
Causes: A single high K reading in routine labs is no reason
to panic. Frequently, this is caused by artificial hemolysis due to improper
blood drawing or sample handling, resulting in release of K+ from
red blood cells.
Yet, K+ released from lysing cells may cause actual hyperkalemia in vivo. Examples are:
·
tumor
lysis syndrome in the initial phase of chemotherapy
·
burns
·
crush
injuries
·
rhabdomyolysis
A classical cause for hyperkalemia due to a shift of K+ out of cells is ketoacidosis in a patient with diabetes mellitus type 1. In
this situation, both lack of insulin and acidosis contribute to the inhibition
of Na-K-ATPase.
Pharmacology cross reference: some drugs have a tendency to induce hyperkalemia:
·
Beta
blockers
·
Digitalis
cardiac glycosides, which directly inhibit Na-K-ATPase, leaving K+ outside
·
potassium-sparing
diuretics: aldosterone antagonists, triamterene or amiloride
·
Trimethoprim
blocks ENaC, as well
·
to
a lower extent, ACE-blockers and AT1-antagonists
Due to the kidneys' enormous capacity to excrete K+,
persistent hyperkalemia is only possible if this function is impaired.
Conversely, as our kidneys usually secrete K+, hyperkalemia is
typical in end stage renal disease.
Symptoms of hyperkalemia are fairly unspecific, including
malaise, muscle weakness, arrhythmia. Sudden cardiac arrest or ventricular
fibrillation is possible. ECG changes start with a "tented", tall and
peaked T wave, then the p wave flattens or disappears and the QRS complex
widens to a pattern reminiscent of a bundle branch block.
Therapeutic
options include fast
and longer-term measures. The fastest way to stabilize membrane potential is
careful injection of Ca2+. Ca2+ ions impede Na+ channels, making it harder to trigger action potentials. Administration of
insulin, together with glucose to prevent hypoglycemia, moves extracellular K+ into cells. In case hyperkalemia is combined with metabolic acidosis, it may be
corrected by bicarbonate: Infusion of HCO3− helps
to move K+ into cells in exchange for H+. Furosemide, perhaps
concomitantly with an infusion of NaCl, will increase excretion of K+ provided the kidney is still in a state to do that. Nonabsorbable cation
exchange resins, introduced orally or by enema into the gastrointestinal tract,
may bind K+ and remove it from the body. In end stage renal disease,
dialysis is the only way to keep K+ values down.
Causes: Hypokalemia may result from losses of potassium from
the body, or from moving extracellular K+ into the cells.
Losses may occur
in the following situations:
·
diarrhea
or repeated vomiting, as gastrointestinal secretions contain a considerable
amount of K+
·
hyperaldosteronism,
in the majority of events secondary to loop diuretics or thiazides or by high-dose
glucocorticoid therapy with overwhelming of 11β-HSD
Intense exercise on a warm, humid day may cause
hypokalemia by a combination of K+-losses by sweating and uptake of
K+ into cells by adrenergic stimulation.
Shifting of K+ into cells may be caused by
administration of insulin or β2-sympathomimetic drugs. For
example, using solely insulin to treat a diabetic patient with high blood
glucose may result in hypokalemia. Instead, insulin should be accompanied by a
K+-containing infusion, which has the additional benefit of filling
up volume.
Strong sympathetic activation by itself may lead to
hypokalemia as well, e.g. by the combination of a drop in blood pressure, pain
and fear in myocardial infarction.
Symptoms: muscle weakness and cardiac arrhythmia, e. g., torsades de points. In ECG, a flattened
T wave merges in a prominent U wave.
For therapy,
it is important to figure out what caused hypokalemia in this particular
patient. In case hypokalemia has been the result of a shift of potassium into
cells, it has to be expected to get out again at some point in time. Filling up
K+ too forcefully risks hyperkalemia later on. This is of little
concern in hypokalemia by chronic losses, where intracellular K levels are on
the low side, either.
Therapy at first
sight looks easy: can't we just fill up the K+ deficit? Two things
have to be kept in mind when trying to do this by infusion:
·
K+ has to be properly diluted and the infusion has to run slowly. Blood from the veins reaches the heart very
quickly, and locally increased K+ concentration there would wreak
havoc with cardiac excitability.
·
In
addition, extracellular K+ is one of the main pain-generating agents.
Consequently, an infusion running too fast may cause pain or discomfort along
the patient's vein.
Therefore, if critical hypokalemia needs to be treated
quickly, it is advisable to do this by central venous access in combination
with heart monitoring. The infused solution should not contain components like
glucose or HCO3− that might shift additional K+ into cells. Rapid administration should proceed only until plasma K+ approaches the lower end of the normal range (about 3 mmol/l); then there is
enough time to complete the task slowly.
The safer route is to fill up by oral administration.
This is best done by having the patient swallow a solution containing KCl,
although this treatment is not popular, as the solution tastes quite awful.
Glucose is freely filtered at the glomerulus. At
physiological concentrations of around 100 mg/dl (5.5 mM), that is a lot of
valuable energy-containing fuel: 1 g for every single one of the 180 liters of
filtrate we produce in a day. Still, under normal circumstances, we excrete no
glucose at all. Virtually the entire load is reabsorbed in the proximal
tubulus. In the early part of the
proximal tubulus, this is an easy process. Low affinity/ high capacity Na+-Glucose
transporter 2 (SGLT2, SLC5A2) shovels most of it into the cells, from where it passes
the basolateral membrane via GLUT2-mediated facilitated diffusion. What glucose
remains in the lumen is taken up in the downstream part of the proximal tubule
by the high-affinity/ low capacity SGLT1 transporter, which uses the energy of
two Na+-Ions to push in the glucose unit. Here, the basolateral back
door is GLUT1.
Being a transporter-mediated process, glucose uptake
is saturable. Once plasma glucose concentration exceeds 250 mg/dl (14 mM) in
diabetics, the two cotransporters work at capacity, and incremental glucose is
excreted, causing osmotic diuresis. This has given the disease its name: diabetes mellitus is Greek for
"sweet flow-through".
Pharmacology cross reference: In patients suffering from diabetes mellitus type 2, one treatment option is to inhibit SGLT2 by specific blockers such as Dapagliflozin or Empagliflozin. Although backup transporter SGLT1 works like crazy, it is not able to fully compensate for the loss of SGLT2 activity. Lots of glucose is excreted via the urine, taking water with it. This lowers glucose levels by decreasing the threshold for excretion to values well below the normal 250 mg/dl. Compared to other treatments, the advantage is that excreted glucose cannot any more be metabolized to fat. In many cases, weight loss is an added benefit. The disadvantages: loss of glucose may lead to tiredness, excessive weight loss, dehydration and ketoacidosis. In addition, bacteria in the urinary tract are well fed by all this glucose, increasing the probability of urinary tract infections. Since blocking the reabsorption of glucose also slows
down the associated proximal co-transport of Na+, a higher Na+ concentration remains available for other co-transport processes. This can lead
to an increase in phosphate reabsorption. Higher extracellular phosphate levels
lower the Ca2+ concentration, increase parathyroid hormone release,
increase FGF23, and lower the active form of vitamin D. This may increase the
risk of bone fractures.
Don't have patients trouble taking up glucose form the
intestines? Glucose uptake in the small intestine is not affected by
SGLT2-blockers, as it is mediated by SGLT1. Although SGLT1 and -2 are closely
related, it would take a thousandfold concentration of Dapagliflozin to inhibit
SGLT1.
Filtrated proteins and peptides are 100% reabsorbed in
the proximal tubule. We need to realize that "reabsorption" means
something different for proteins than for electrolytes or glucose. Once "outside"
by filtration, proteins cannot reenter as such, they have to be broken down and
digested to amino acids. Still, amino acids are valuable enough to be
retrieved.
Most proteins and peptides are bound and endocytosed
by a receptor complex combining the three transmembrane proteins megalin-cubilin-amnionless.
For proteins and peptides small enough to be filtered, renal elimination is a
determining factor for their plasma levels. Via this mechanism, end-stage renal
disease can lead to elevated levels of glucagon, gastrin and ANP and may further
increase secondary hyperparathyroidism.
The pH of our blood, one of the most closely regulated
parameters in our body, is about 7.4. In most other fluid compartments, it is
very near that value. It is essential that pH remain near 7.4, because the
conformations of our proteins and thus, the activities of many enzymes, are
very sensitive to changes in pH.
In diluted solutions like our fluid compartments, pH
approximates the negative of the logarithm to base 10 of the concentration of
hydrogen ions in mol/l. In other words, the hydrogen ion concentration in our blood
plasma is 10-7.4 mol/l, that is 0.000 000 04 mol/l or 40 nmol/l.
This 40 nM proton concentration is a dynamic equilibrium, resulting from
an ever ongoing flourishing trade in protons by thousands of different donors
and acceptors. At every point in time, a tiny fraction of protons belongs to
nobody and is hurry-scurrying about: only this fraction determines pH. The
situation is complicated by the fact that this trade is conducted in
compartments separated by biological membranes. Some of the traders, like CO2,
can pass these membranes effortlessly, while others, like phosphate, cannot.
Furthermore, proton trade across membranes can't go on for long unless it
satisfies the requirement for electroneutrality. Unfortunately, our mind is
very ill-equipped to handle this complex trade involving thousands of
interdependent dynamic equilibria in an open system. Even our mathematical
models are inadequate. Our "knowledge" stems from experiments where
we try to vary the concentration of the biggest traders while keeping
everything else as constant as possible.
The biggest trading house in our body's proton market
is CO2 & HCO3−.
Luckily for us, the acidic one of its two partners has the habit of vanishing
into thin air, and if he didn't, the trade would go sour very soon. Each day,
we eliminate 15,000 mmol of CO2, a potential acid, via the lungs.
On the other hand, the buffering partner HCO3−, our main defense
against acidification, is in danger of vanishing via the kidney back door. Each
liter of primary glomerular filtrate contains 24 mmol HCO3−. At 180 liters per
day, that is 180 x 24 mM= 4320 mmol, or in the range of 4-5 moles. Loss of any appreciable
part of this would result in catastrophic metabolic acidosis: complete
reabsorption is therefore essential. This is accomplished by secreting H+ into the tubule, converting HCO3− to CO2, which is easily reabsorbed. This process is aided by
carbonic anhydrase IV, which is GPI-anchored to the outside of the apical
membrane, and carbonic anhydrase II inside the cell, which catalyzes the
opposite reaction. At the basolateral membrane, HCO3− is exchanged for Cl−or, following its
gradient, cotransported out together with Na+. This is accomplished
via the electrogenic Na-HCO3− cotransporter NBCe1 (N for Na+, B for bicarbonate, C for
co-transporter, e for electrogenic, SLC4A4). The carrier is able to transport
in both directions. The amount of HCO3− generated in the proximal tubule cell is such
that three HCO3− push a reluctant Na+ out of the cell. Eighty to ninety percent of
HCO3− is
recovered in the proximal tubule, the rest by α‑intercalated cells
in the distal tubule and collecting ducts.
H+ is extruded into the lumen via three
mechanisms. The first two of them are active in both the proximal and the distal
tubule, while the third one is restricted to distal tubule and collecting ducts:
1.
The
Na-H exchanger (NHE) is driven by the steep lumen-to-cell Na+ gradient in the proximal parts of the nephron. It mediates the majority of H+ transport but cannot build a strong proton gradient, i.e., a lower pH in the
lumen than in the cell.
2.
An
electrogenic H+ pump uses ATP to build a lumen-positive proton
gradient. This pump is expressed in all tubule segments, but highest expression
levels are found in α-intercalated cells of the cortical collecting tubule
and the medullary collecting duct.
3.
An
electroneutral H+-K+-pump in the distal tubule uses ATP
to exchange luminal K+ for H+. This system may contribute
to the generation of hyperkalemia in metabolic acidosis.
Thus, the majority of H+ is secreted in the
proximal tubule, but if a proton gradient is required, this is built up in the
distal tubule by a-intercalated cells. With all these mechanisms working at
capacity, urine pH may reach a value as low as 4.4; that is a thousand times
the H+ concentration in blood (40µmol/l compared to 40 nanomol/l in
blood). That sounds and is impressive, yet quantitatively, these 40µmol/l sink
into insignificance compared to the 4-5 moles of recovered HCO3− and, as we will
appreciate in a minute, compared to the up to 300 mmol of acid equivalents
eliminated in the form of NH4 per day. The kidney is thus able to
excrete acid equivalents over and above the recovery of HCO3−. If there is a
problem with acid excretion in an otherwise functioning kidney, we speak of renal tubular acidosis, which may
result from many genetic as well as acquired causes.
However, the kidney is also able to cope with an
alkaline challenge, e. g., loss of acid by vomiting. In this case, it simply
reduces H+ secretion so that a corrective amount of HCO3− is excreted with the
urine.
Pharmacology cross
reference: Acetazolamide blocks carbonic anhydrase and strongly inhibits HCO3− reabsorption,
leading to the excretion of alkaline urine.
On a typical Western diet, our metabolism produces a
surplus of about 70 mmol H+ as nonvolatile acids per day, which we
have to eliminate to prevent acidosis. This is mainly the result of a diet rich
in proteins. Protein from animal sources contains considerable amounts of
sulfur-containing amino acids cysteine and methionine. On metabolization, they
lead to the formation of sulfuric acid, which is a strong acid. Furthermore, the more we rely on proteins to satisfy our energy requirements,
the more nitrogen we have to dispose of and the more acidity we produce. In the
liver, gluconeogenesis produces waste from clipped-off amino groups, which can
be disposed of in two ways.
1.
Usually,
and depending on acid-base status, more than 90% are detoxified still in the
liver by integration into urea. While far more complicated in detail, the
process boils down to fusing two proton donors in the form of NH4+ and one proton acceptor in the form of HCO3−into one electroneutral molecule of urea,
leaving one acid equivalent on the table. So, this process consumes HCO3−.
2.
The
liver's second option to dispose of nitrogen is by packaging toxic NH4+ in non-toxic glutamine for transport. The kidney retrieves the NH4+.
The proximal tubule cell cleaves two ammonium ions from glutamine, leaving
α-ketoglutarate. The NH4+ is secreted. Two units of
α-ketoglutarate and 4 H+ from 4 CO2+4 H2O
are combined to glucose, leaving 4 HCO3−. Therefore, for each NH4+ secreted into the tubule lumen, one HCO3− leaves the cell at the basolateral membrane.
Gluconeogenesis with integrated acid secretion, in fact energy extraction
coupled with waste disposal: renally ingenious! Whether HCO3− is actually gained
in the kidney, or rather previously in the liver, by obviating the need to
consume it in urea synthesis is a matter of perspective. In summary, secretion
of acid equivalents in form of NH4+saves HCO3− that would otherwise
be consumed in urea synthesis.
Therefore, it is rather the liver than the kidney which regulates excretion of acid equivalents in the form of NH4+. As we will see later when discussing the pathophysiology of the liver, a tendency to acidosis causes the liver to dispose of more nitrogen in the form of glutamine and less in the form of urea. This will produce more new HCO3− and consume less existing HCO3−, thereby correcting incipient acidosis. Because the pKa of NH4+/
NH3 is 9.2, almost all of it is in the ionized form at pH 7.4
(At pH 7.2, 1% would be NH3). While NH3 readily
passes most cell membranes, NH4+ does not. As the luminal
fluid usually becomes more acidic in the later sections of the tubule, it
becomes progressively more difficult for NH4+ to slip back
through the membrane during the ultra-short time frames it adopts the form of
NH3; NH4+ thus becomes trapped and can be
excreted. The lower the urine pH, the higher is the ammonium excretion.
The kidney builds a considerable ammonium gradient in
the medulla, which helps to excrete ammonium and, thereby, acidity. The plasma membrane
of the thick ascending limb is relatively impermeable for NH3. Here,
NH4+ is reabsorbed by taking the slot of K+ in
the Na-K-2Cl-cotransporter and in K+-channels. At the higher
intracellular pH, it is easier for ammonium to leave the cell at the basolateral
side in the form of NH3, leading to accumulation of NH4+ in the interstitium of the renal medulla: the deeper into the medulla,
the higher the concentration of NH4+. This gradient leads
to net secretion into the descending parts of the nephron and a bypass of the
cortical portions of the distal nephron.
This way, it is possible to excrete higher percentages
of the relatively toxic ammonium. In addition, with every molecule of NH4+,
an acid equivalent leaves the body.
Some (about 30 mmol per day) of the protons extruded into the lumen are
buffered by acceptors like HPO42− (pKa of H2PO4−: 7.2), urate (pKa
5.75) and creatinine (pKa 5.0). They are called titratable acids as the amount
of buffered protons in acidic urine may be determined by measuring the amount
of sodium hydroxide required to bring pH back up to 7. NH4+ is not considered a titratable acid, because with a pKa of 9.2, it does not
release its proton in this maneuver.
If increased amounts of protons need to be excreted,
titrated acids are of little help. Phosphate buffering, the most important, is
limited by the amount of phosphate present in plasma. Only in case of
ketoacidosis, ketoanions like β‑hydroxybutyrate may contribute to a
considerable increase in titratable acids.
Thus, in acidosis, the predominant way to dispose of
additional acid equivalents is by increasing excretion of ammonium. This can increase from the usual 40 to 300 mmol per day.
"Acid ash"
diet does not lead to osteoporosis.
There is a common and often-repeated misperception
that, via generation of acid equivalents, a Western diet contributes to osteoporosis. There is no scientific basis for this assertion. While
it is true that osteoclasts break down bone substance with the help of
acidification, as we will discuss when dealing with bone metabolism, osteoclasts are careful to limit this acidification
to their very small operating range. And while there are good reasons to be
critical of our Western diet, the production of acid equivalents is not one of
them. Healthy kidneys are perfectly able to dispose of the 70 mmol per day of
non-volatile acidity resulting from a Western diet; in fact, acid elimination
could be ramped up to a multiple of that. Of course, blood and bone marrow pH remain
the same, irrespective whether people are stuffing themselves with sausages or prefer
a vegan diet. Sausage stuffers have higher blood pressure, more atherosclerosis
and more myocardial infarctions, but not more osteoporosis.
The situation is very different once there is a
serious problem with renal acid elimination. Renal tubular acidosis, which may
be due to genetic causes or may be secondary to, e.g., autoimmune diseases or
sickle cell anemia, leads to osteomalacia or to rickets in children. In this
case, acid equivalents are in fact buffered by phosphate from hydroxyapatite, which is thereby dissolved.
pH-dependent excretion
of pharmaceuticals: weak acids and bases
Excretion of weak acids and bases depends on pH, as
their non-ionic forms pass the membranes of tubulus cells a lot more easily
than their charged forms. Consider acetylsalicylic acid as an example. At low
urine pH, most of it is protonated and diffuses back into the blood. At high
urine pH, it is ionized as acetylsalicylate, stays in the tubulus lumen and is
excreted. The opposite is true for a weak base like quinine. As many
pharmaceuticals are weak acids or bases, urine pH has a profound influence on
their excretion. For example, an overdose of acetylsalicylic acid may be
treated by alkalinizing the urine by an infusion of HCO3−.
Elimination of organic anions and
cations
The late proximal tubule secretes several organic
anions and cations. Among the anions are oxalate, α-ketoglutarate, bile
salts and liver-manufactured glucuronate and sulfate conjugates. The yellowish
color of normal urine is mainly due to bilirubin-derived metabolites such as
urobilin. Among secreted cations are dopamine, epinephrine and norepinephrine.
Therefore, the rate of production of catecholamines may be determined from
their concentration in 24-hour urine.
Correction of
acidosis and alkalosis
An increase in arterial PCO2 is compensated by increased renal H+ secretion, which translates to production of new HCO3− via excretion of NH4+.
In chronic respiratory acidosis, the capacity of this mechanism is ramped up by
induction of the apical Na-H-exchanger and the basolateral Na-HCO3− cotransporter.
At high altitudes, air pressure and, concordantly, oxygen
partial pressure are diminished. We compensate for this by intensified
breathing, but in doing so we also lower the PCO2 and thus develop
respiratory alkalosis. That pulls more protons off albumin; the additional
negative charges bind more Ca2+ to albumin, which increases the risk
of cramps by relative hypocalcaemia. With PCO2 lowered, the kidney reduces its H+ secretion (or the liver its glutamine synthesis), so that a compensating amount
of HCO3− is excreted in the urine. This can be felt as so-called bicarbonate diuresis,
so we have to be careful to drink enough when in the mountains.
If, for some reason (e.g., lack of oxygen), tissues are incable of metabolizing glucose or fatty acids to CO2 and water with the help of mitochondria, they have to resort to anaerobic glycolysis; the result is lactic acidosis. Prolongued absence of insulin action makes the body switch into hunger mode: the result is ketoacidosis. Whatever the cause, increased acid equivalents are first buffered by HCO3−, generating CO2 and H2O. The first response is increased ventilation to reduce PCO2. In many cases, metabolic acidosis is the result of impaired renal function; of course, this precludes renal compensation. Yet, in chronic diarrhea, lactic acidosis or diabetic ketoacidosis, metabolic acidosis is caused by extrarenal factors. In these cases, the kidneys react via the same lines as in respiratory acidosis: H+ secretion is increased via induction of Na-H-exchanger and electrogenic H+ pump, combined with induction of the basolateral Na-HCO3− cotransporter. In addition, NH4+ production is increased by activation and induction of the enzymes involved in liver and kidney, enabling elimination of more acid equivalents which translates to generation of more HCO3−. Base excess allows estimating the magnitude of
metabolic deviations from acid-base equilibrium. Base excess is defined as the
amount of acid that must be added to each liter of blood to return the pH to
7.40 at standard respiratory conditions (100% oxygenation and PCO2 40 mm Hg at 37°C). In the absence of any metabolic deviation, there is nothing
to titrate; reference range is 0 ±2 mEq/l. Somewhat incongruous, the
"excess" may be positive or negative. Positive values > 2 mEq/l
indicate metabolic alcalosis, negative values < −2 mEq/l metabolic
acidosis.
For all practical purposes, base excess is not
assessed by separate titration. Rather, it is calculated in an arterial blood
gas test, which is performed anyway in case of acid-base problems.
[Just to illustrate that: base excess is calculated
with pH and PCO2 as input, from a combination of Siggaard-Anderson
and Hendersen-Hasselbalch equations:
base excess = 0.02786 x PCO2 x 10 (pH - 6.1) + 13.77 x pH − 124.58]
The best treatment of acidosis is correction of the underlying cause. Sometimes, however, metabolic acidosis needs urgent correction. In this case, base excess helps to assess the required amount of buffer. What is the fluid volume we need to take into consideration? Of course, it is not possible to correct only "the blood". A better approximation would be extracellular fluid volume: in our 70 kg female, that would amount to 14 liters and in the respective 70 kg male, to 17 liters. Yet, part of our buffer, sodium bicarbonate, equilibrates with intracellular volume which is also affected by acidosis. Therefore, 30% of body weight is assumed as target volume, or 21 liters in our 70 kg standard persons. As blood is only part of that volume, the calculation of base excess is slightly modified. Hemoglobin acts as a buffer itself. Standard base excess refers to a hemoglobin concentration of 5 g/dl, down from 12-18 g/dl in blood, to better reflect the properties of the target volume. If metabolic acidosis with a standard base excess of −10 mEq/l needs to be corrected in our 70 kg person, the amount of NaHCO3 required is approximately 70 x 0,3 x 10 = 210 mmoles. Caution! This type of correction is a critical procedure reserved for specialists, as it entails considerable risks (rapid increase in PCO2, risk of overcorrection, K+-shifts, osmotic effects like hypernatremia. Typically, half the dose is administered very slowly, followed by reassessment of the situation). The size of the metabolic deviation is important, yet it
is of little help in determining the cause of metabolic acidosis in the
individual patient. The next useful step in this regard is determining the anion gap. Plasma electroneutrality
implies that the number of anions equals the number of cations. The majority of
these ions are "visible" in routine labs: Na+,
K+ Cl−, and HCO3−. A
small remainder is invisible. In the patient with acidosis, could an unusual
amount of acids hide in this invisible corner? Lactate, for example, or maybe
ketone bodies acetoacetate of β-hydroxybutyrate? Well, it is possible to
estimate the amount of these anions by comparing anion and cation
concentrations. In doing this, we are lazy, ignoring K+: its
concentration is small anyway, and we are not going to fuss about a constant
error of 3%. There we go:
Anion gap = [Na+]
– ([Cl−]+[HCO3−] )
Normally, this value equals 12±2, reflecting...Um, what again? Anions, of course, for
example phosphate, but that's small,...Oops, almost forgot the proteins!
Proteins bear positive and negative charges, yet in total, more negative ones.
Think of albumin's 15 negative net charges. In fact, albumin constitutes no
less than 60% of plasma protein. Accordingly, the normal anion gap of 12 is
mainly albumin. But, albumin may be lost if there is a problem with the
glomeruli, albumin synthesis may decline in liver disease: we will have to take
it into account if albumin values are in any way unusual.
Acidosis with increased anion gap: if albumin is
normal, yet the anion gap is increased, the plasma obviously contains an
unusual amount of acids. We have to think about lactic acidosis, ketoacidosis,
uremia and a number of alternative causes.
Acidosis with normal anion gap: in this case, HCO3− has been reduced without concomitant addition of
acids. Possible causes include HCO3− losses by chronic diarrhea and renal tubular acidosis.
Among typical causes are persistent vomiting (as in bulimia) or
treatment with diuretics. Furosemide or
thiazides activate aldosterone, which exchanges not only K+, but
also H+ for Na+ in the collecting duct. Interstitial
HCO3−at
the basolateral side of proximal tubule cells is increased: more HCO3− leaks back into the
tubule lumen via the paracellular route. In addition, the increase depresses
the transport rate of the Na-HCO3− cotransporter, causing HCO3− to back up in the
cytosol. So, intracellular carboanhydrase, too, has to push against the backlog
of its product, generating lower amounts of H+ for secretion into
the tubule lumen. Apart from this effect in the proximal tubule, an additional
adaptation is found in the cortical collecting tubule: a shift in the ratio of
α to β intercalated cells, increasing the proportion of β cells.
While α cells pump H+ into the lumen and release HCO3− into the blood,
β intercalated cells have the opposite orientation, secreting HCO3−. Thus, in metabolic alkalosis, the cortical
collecting duct may switch from acid secretion to HCO3− secretion.
Volume contraction
stimulates H+ secretion
Volume contraction activates the RAAS and sympathetic
systems. Both Angiotensin II and norepinephrine stimulate Na-H exchange in the
proximal tubule, and aldosterone enhances H+ secretion in the
collecting duct. By these mechanisms, volume depletion not only lowers urine
volume; it lowers urine pH, too, which may contribute to stone formation.
Calcium oxalate stones and uric acid stones form especially at low urine pH (in contrast,
calcium phosphate stones form at increased pH).
Hypokalemia
stimulates H+ secretion
In the proximal tubule, K+ depletion
activates Na-H exchange and electrogenic Na-HCO3− cotransport. In addition, K+ depletion activates K-H exchange by α‑intercalated cells in the
collecting tubule. Hypokalemia may thus lead to metabolic alkalosis.
9. DISEASE MECHANISMS AFFECTING GLOMERULAR FUNCTION
In a young adult,
every kidney contains about one million glomeruli. Age is the invincible enemy
of our glomeruli: the older we become,
the more glomeruli we lose. These microfilters can only function if they
·
constantly
receive an adequate flow of blood and
·
remain
structurally intact
Problems with blood flow may be due to atherosclerotic
lesions, to hypovolemia, to hypertension or to other causes, which we deal with
elsewhere.
Structural alterations have many causes as well. For
example, arterial hypertension or diabetic metabolic state over time results in
structural damage and loss of glomeruli. Many of the mechanisms leading to
damage of the glomerular structure are immune in nature. As we have seen when
dealing with our defense system against microbial invaders, our immune system
commands a range of sharp weapons. The primary route of delivery of complement
components, neutrophils, monocytes, antibodies and T cells is via the blood.
Accordingly, plenty of these tools reach the glomeruli and in addition, there
is ample opportunity for them to get stuck in the filter. Depending on where
exactly the primary damage occurs, clinical symptoms may vary widely.
Damage to
podocytes: mainly nephrotic glomerular diseases
Membranous
glomerulopathy. Recall
that the slit diaphragm is the part of the filter with the tightest pores.
Antibodies may thus reach the capillary-facing side of the podocytes, even when
slit diaphragms are intact. If any of these antibodies cross-react with
structures on the podocyte surface, complement may be activated and podocytes
may react to the ensuing damage by retracting their foot processes.
Consequently, the slit diaphragm disappears, while the basement membrane part
of the filter remains largely unaffected. Proteins are filtrated in great
quantities, but blood cells are unable to pass. Now, antibodies reach the
lumen-facing side of podocytes as well.
Normally, with complement activation, small cleavage
products like C3a, C4a, and C5a diffuse into the surrounding tissue and attract
leukocytes. Here, in the midst of a rushing plasma cascade, any chemoattractant
is washed away. Accordingly, no inflammatory infiltration is seen in light
microscopy.
Yet, antibody and complement deposits sit on the
basement membrane-facing side of the podocytes, and podocytes react by
producing more basement membrane material. Basement membranes consist of
proteins like collagen type IV and laminin, as well as polysaccharides. The
polysaccharide component can be visualized with periodic acid Schiff
(PAS)-staining. Basement membranes appear markedly thickened, and analogous
changes may be seen in electron microscopy. In fluorescence microscopy for IgG
or complement, a grainy structure of staining follows the glomerular basement
membrane.
Emphasized membranes in the absence of leukocyte
infiltration have led to the morphological designation membranous
glomerulopathy. The term membranous glomerulonephritis is used frequently, too,
but is somewhat misleading as there is no inflammatory infiltration.
Which podocyte structures are targeted by antibodies? In
majority of cases, it is either a phospholipase-A2 receptor or
neutral endopeptidase (aka CD10 or CALLA). In children, antibodies against
nutritional bovine serum albumin from cow milk or beef have also been shown to
play a role. Having passed the intestinal barrier, maybe via M cells, the
protein is modified to a cationic state. In the cationic state, it becomes
implanted in the capillary wall near the surface of podocytes, which is studded
with numerous negative charges. There, it is attacked by antibodies against the
foreign protein.
Nephrotic
syndrome: With
membranous glomerulopathy, the patient usually loses large amounts of protein
of all sizes (thus including IgG) in the urine, but no erythrocytes or other
cells. Note that nephrotic syndrome also may be caused by marked, isolated loss
of albumin. Nephrotic syndrome is characterized by loss of plasma protein
combined with Na+ retention and edema. What is cause and what is
effect? This is debated and may vary from case to case:
Underfill
hypothesis: Loss of protein reduces intravascular oncotic pressure, so that
fluid is lost to the interstitium. Vascular underfill activates the
renin-angiotensin-aldosterone system causing compensatory Na+-retention.
Overflow
hypothesis: Increased glomerular filtration of proteins causes damage to the
nephron resulting in primary Na+ and fluid retention, potentially
via ANP resistance. Increased intravascular volume leads to overflow and
secondary edema formation. Several clinical and experimental observations seem
to support this view. In many cases with edema formation, oncotic pressures of
plasma and interstitium go down in parallel, leaving the differential constant.
If patients are treated with glucocorticoids, frequently edema retreats before
plasma albumin increases.
In nephrotic syndrome, GFR usually remains normal, but may be reduced in some cases.
Damage to
podocytes resulting in nephrotic syndrome may come in two additional variants,
"light" and "heavy":
·
light
functional damage: minimal change disease
·
massive
destruction: primary focal segmental glomerulosclerosis (may also develop
without causing nephrotic syndrome if glomerular destruction stops filtration
of proteins)
In
both cases, antibodies are usually not detected; therefore, the causative
damage must be of a different nature. Severity, rather than quality of the damaging
mechanism may determine which of the two forms develops.
Minimal change
disease: By light microscopy, the glomeruli look normal, neither antibodies nor
complement are detected by fluorescence microscopy. It is speculated that
podocytes react to soluble cytokines by retracting foot processes, a change
seen only in electron microscopy. Why cytokines? Usually, the patients enter
remission with glucocorticoid therapy and as we have learned, glucocorticoids
inhibit expression of many cytokines. Frequently, glucocorticoid withdrawal is
followed by relapse.
Primary focal
segmental glomerulosclerosis: This disease is characterized by necrosis and loss of podocytes, which
cannot be replaced. The result is massive flow through the basement membrane.
Proteins too big to pass through the pores of the basement membrane get stuck
and accumulate until the filter is completely clogged. "Glomerulosclerosis"
indicates collapse or obsolescence of glomerular capillaries.
"Segmental" indicates this involves part of the tuft of an individual
glomerulus; "focal" means less than 50% of glomeruli are affected as
judged by light microscopy.
Damage to the entire
capillary wall: mainly nephritic symptoms
(Diffuse) proliferative
Glomerulonephritis. Any
filter is prone to collect particles; if soluble immune complexes circulate in
the blood, lots of them are bound to end up in the glomerular basement
membrane. As there is no basement membrane separating capillary and mesangial
space, immune complexes also enter the mesangium. Alternatively, immune
complexes may assemble locally in the basement membrane if antibodies bind any
of its components.
Typical examples for the entrapment of circulating
immune complexes are poststreptococcal glomerulonephritis and lupus nephritis.
Complement is activated and C3a and C5a are generated in the immediate vicinity
of endothelial cells and leukocytes. Endothelial activation causes neutrophils
and monocytes to attach and to wriggle their way between endothelial cells to phagocytize
the complexes. Some of these leukocytes accumulate in the mesangium. In light
microscopy, increased numbers of cells or nuclei are visible, which enticed
pathologists of yore to describe the process as "proliferative". Although
this has proved incorrect, as it is infiltration rather than proliferation, the
name has stuck. Activated macrophages secrete IL‑1β, TNFα, IL‑6
and release their protease-containing granules. Locally, the basement membrane
is damaged or broken down, causing erythrocytes to pass the filter.
Nephritic
symptoms: The
damage to the basement membrane causes hematuria; erythrocytes and casts are
found in the urine sediment. Proteinuria is present but in many cases remains less
pronounced than in nephrotic syndrome, because not all glomeruli are damaged:
edema may or may not form. Depending on the extent of damage to
the glomeruli, GFR may range from normal to markedly reduced. RAAS activation may
cause hypertension.
This diffuse
proliferative glomerulonephritis ("diffuse" means most glomeruli
are affected, as opposed to "focal") is typically seen in
postinfectious glomerulonephritis and frequently resolves within a few weeks,
after the invading pathogen has been eliminated. However, if formation of
immune complexes does not stop, another type of morphology develops:
Membranoproliferative
glomerulonephritis. If infections become chronic, as in viral hepatitis, or in case of chronic
autoimmune processes like systemic lupus erythematosus, the load of immune
complex deposits increases over time despite the effort of immigrating
macrophages to clear the stuff out. Endothelial cells attempt to compensate by
synthesizing a new basement membrane over the piles of immune complex debris. In
the microscope, the thickened walls of capillaries seem like bands of amorphous
material. In PAS stains, typical "tram track" double contours may be
seen: two PAS-positive basement membranes are separated by a PAS-negative layer
of immune complex debris. More visible membranes and more cells: membranoproliferative glomerulonephritis.
Clinically, hematuria is likely to persist, GFR to come down; prognosis is
poor.
A similar picture develops in individuals who, due to
genetic factors, suffer from an especially trigger-happy complement system. As
we have seen previously, complement is constantly being activated via the
alternative pathway, yet reigned in on our cell membranes by inhibiting factors
like H, I or MCP (CD46). Homozygous defects in any of these regulatory factors
result in variant forms of membranoproliferative glomerulonephritis called
either dense deposit disease or glomerulonephritis C3. Erythrocyte
lysis may complicate the situation, completing the picture of atypical hemolytic-uremic syndrome (aHUS).
Pharmacology cross
reference: Eculizumab inhibits complement activation at C5 and may be used to treat aHUS.
IgA nephritis and
Henoch-Schönlein purpura. Immune complexes containing IgA1 are the common denominator of these two
related diseases. Usually, within only one or two days after the mucosal immune
system has been reactivated by a respiratory or gastrointestinal infection with
an agent "seen" previously by the immune system, IgA1-containing
protein complexes cause vasculitis with multiple disseminated small bleeding
episodes. If the process is limited to the kidney, hematuria may be the only
symptom. If more widely disseminated, the classical Henoch-Schönlein-trias develops
with purpura visible on legs and buttocks, arthritis and abdominal pain, with
or without hematuria.
The IgA1 deposited in immune complexes is
unusual in that it lacks galactose units normally present near its hinge
region. Why glycosylation is deficient in affected individuals is poorly
understood. It has been proposed that these galactose-deficient IgA1-units
act as an autoantigen and are bound by naturally occurring anti-glycan IgA1 or
IgG antibodies. These antibodies thus act like rheumatoid factors, resulting in
immune complexes consisting of two antibodies. The complexes seem to activate
complement indirectly via the lectin or alternative pathway and are recognized by
receptors expressed on mesangial cells. Mesangial cells start to proliferate,
secrete cytokines and overproduce extracellular matrix components. Prognosis is
good in children; in adults, the process tends to become chronic and
complications are more frequent.
Goodpasture
syndrome: Antibodies directed against the basement membrane. While the diseases mentioned above are type III hypersensitivity reactions caused by immune complexes, glomerular disease may
also be caused by a type II direct antibody attack. The antibodies are directed
against an epitope of the α3-chain of type IV collagen, which is the main
component of basement membranes. Unsurprisingly, many patients develop not only
renal but also respiratory symptoms, with shortness of breath, cough, coughing
up blood and chest pain. To Yours Truly, it remains unclear why basement
membranes of other organs are unaffected.
Glomerular damage is usually severe, with focal
necrosis and crescent formation and characteristic linear, not granular
deposition of IgG along basement membranes. Accordingly, kidney failure may
develop rapidly. For therapy, it is necessary to remove as much antibody as
possible by plasmapheresis and to suppress formation of new antibody by
aggressive immunosuppression. Regardless, in many patients kidneys are
destroyed within months.
Secondary focal
segmental glomerulosclerosis with hyalinosis ("renal aging"):
By light microscopy, the disease is often
indistinguishable from primary focal segmental glomerulosclerosis due to direct
podocyte damage. It is a sign of slowly progressing kidney damage, after
nephron loss by any cause has exceeded a certain limit.
The secondary variant reflects the growing strain on
remaining glomeruli, which hypertrophy and increase their single nephron
glomerular filtration rate. Higher single nephron filtration is reached by
opening of the vas afferens, combined with increase in systemic pressure, while
the efferent arteriole is constricted. With higher pressure hammering relentlessly
at the glomerular capillaries, the capillaries are distended and elongated,
increasing the volume of the glomerular tuft and the surface available for
filtration. Podocytes are terminally differentiated cells unable to divide;
they only increase in size. Once they are stretched to the maximum, they no
longer manage to keep intact all interdigitating foot processes and slit
diaphragms, leaving "tears" in the sheet below the basement membrane.
At these tears, the larger pores of the remaining basement membrane allow far
more protein to rush through. Because of reduced hydraulic resistance, also more
plasma volume rushes through. The tubule is overwhelmed by the amount of
filtrated protein. The patient develops microalbuminuria, later full-fledged
proteinuria. Big proteins like IgM or fibrin are too large to pass even through
the bigger pores of the basement membrane; they get stuck, over time clogging
the capillary wall with a hyaline deposit.
Increased shear stress and tension occasionally
ruptures the weaker part of the glomerular capillary wall towards the
mesangium, exposing mesangial matrix components to platelets. Thrombosis,
inflammation and organizing reaction follow, leaving behind collapsed
capillaries representing an area of segmental glomerulosclerosis in the
microscope.
Even before the entire glomerulus collapses, tubules
suffer, too. The fall in GFR causes phosphate retention (see FGF23 below). The
tears in slit membranes cause filtration of transferrin, which increases iron
load on proximal tubule cells; iron promotes generation of reactive oxygen
species.
The end is marked by the complete collapse of the
glomerulus. Once a glomerulus obliterates, the lack of blood flowing from its
efferent arteriole will damage not only that nephron's tubule, which would not
matter anymore, but also tubule segments of nephrons in close proximity.
A vicious cycle develops: the more nephrons die, the
higher the strain on the remaining ones. Clinically, the process manifests with
gradual elevation in creatinine concentration, slowly increasing proteinuria
and hypertension. Over time, it leads to end stage renal disease.
10. TUBULOINTERSTITIAL DISEASES
All of the tubules' blood supply stems from blood exiting
glomeruli via vasa efferentia. If one glomerulus is lost due to one of the disease
mechanisms mentioned above, the loss of the respective vessel loops negatively
affects the tubules of several neighboring nephrons.
Apart from problems with blood supply, tubules may be damaged by infections, autoimmune phenomena, toxic effects or inherited conditions.
Infections of renal
parenchyma are usually caused by bacteria that cause pyelonephritis via
ascending infection from the bladder. In children and adults, this occurs much
more frequently in females, due to
·
the
short female urethra, which allows passage of bacteria, especially during
sexual intercourse
·
the
presence of the vaginal microbiota
·
the
lack of antibacterial prostatic fluid
Ascending
infections require a number of bacterial and host factors to occur, including
properties allowing bacteria to adhere to glycolipids of the urothelium,
vesicoureteral and intrarenal reflux. Intrarenal reflux is promoted in compound
("twin –peak") papillae, where collecting duct openings are less
likely to be closed by the hydrostatic pressure in the pyelon. Pyelonephritic
scars are therefore typically found at the renal poles, where compound papillae
occur.
Pyelonephritis is critical because infection and the
inevitable damage from infiltrating neutrophils and macrophages affects a
tissue that receives little blood supply and operates under harsh conditions in
the best of times. Increases in interstitial pressure by increased vascular
permeability further reduce oxygen supply. With severe infection or delayed
antibiotic therapy, entire papillae vanish as lost tubules cannot be replaced.
What remains is a thin layer of renal cortex filled with useless glomeruli over
an empty calyx.
Drug-induced renal
impairment and injury
Drugs may interfere with kidney function by vascular
effects and by effects on tubule cells. Due to their concentrating effects,
renal tubules are especially likely to suffer unwanted injury from drugs. Recall
once more that cells in the loop of Henle and in the medullar collecting ducts,
while metabolically highly active, reside in a hyperosmotic, hypoxic
microenvironment. In addition, many drugs are metabolized in the proximal
tubule by cytochrome p450 and other systems, exposing downstream tubule
segments to unusual concentrations of metabolites.
Conversely, the kidneys' central role in drug
elimination implies that in pharmacotherapy, renal function has to be taken
into account. In many cases, loading and maintenance doses need to be
calculated based on eGFR, and concomitant volume expansion may be required to
limit concentrating effects. Let's have a look at a few examples.
Drugs with
primarily vascular side effects
Non-steroidal
anti-inflammatory drugs (NSAIDs) probably are the most frequently used drugs worldwide. Inhibition of
cyclooxygenase interferes with prostaglandins' role in keeping the vas afferens
open in in a broad range of physiologic and pathologic situations. Of course,
this effect is magnified in the presence of hypovolemia. Use of NSAIDs in
endurance sports enhances the risk of hyponatremia in case of overdrinking.
Angiotensin-converting
enzyme (ACE) inhibitors and AT1 receptor
blockers inhibit the effect of angiotensin II, which is important to
maintain GFR in the face of decreased systemic pressure. Accordingly, use of
these drugs may compromise renal function, more so when they are combined with
NSAIDs, cyclosporine or tacrolimus, especially in patients with congestive
heart failure or in situations of hypovolemia.
Calcineurin
inhibitors,
cyclosporine and tacrolimus, interfere with the autocrine IL‑2 feedback
loop necessary for T cell proliferation and are used for immunosuppression, e.
g., following renal transplantation. They have a very narrow therapeutic index
and may damage the kidney via several mechanisms, of which the most important
one is a constriction of the afferent arteriole.
Iodinated
radiocontrast agents are one of the most common causes of acute kidney injury in the hospital.
Intravenous administration is followed by an intense constriction of the vas
afferens, enhancing hypoxia in the medulla. In addition, the radiocontrast
agents damage tubule cells by an osmotic effect. Typically, a rise in serum
creatinine is seen within 24-48 hours. Previous administration of volume reduces
the frequency of complications.
Aminoglycosides have a cationic structure and are freely filtrated
and then reabsorbed, leading to accumulation in proximal tubule cells. This may
interfere with transport of cations like K+, Mg2+ and Ca2+,
leading to hypokalemia, hypomagnesemia and hypocalcemia. More pronounced
accumulation may result in cell death. Even at normal therapeutic
concentrations, nephrotoxic effects have been reported in 10-25%.
Sulfamethoxazole-trimethoprim may cause hyperkalemia, as trimethoprim inhibits ENaC
in the distal tubule, acting like the K+-sparing diuretic amiloride.
Amphotericin B is used to treat systemic fungal infections. Its name
originates from its amphoteric properties: it places itself in lipid membranes,
next to ergosterol, a sterol lipid present in fungal, but not in human
membranes. Its presence interferes with normal membrane function, e. g., it makes
K+ leak out. It also has some, albeit lower, affinity to
cholesterol, the sterol present in our cell membranes, which explains its
toxicity. Renal toxicity starts with tubular acidosis, concentration defects
and electrolyte imbalances and may end in acute tubular necrosis. As
therapeutic use is limited by nephrotoxicity, a liposomal formulation has been
developed that has an improved renal safety profile.
Acyclovir, administered via venous infusion to treat serious
infections by herpes viridae, may induce tubular injury by precipitation. Slow
infusion and volume expansion to increase diuresis are essential.
Lithium therapy is used to treat bipolar disorder. Li+-ions
inhibit adenylyl cyclase, thus decreasing cAMP, the second messenger of ADH.
This reduces the number of aquaporins brought to the membrane of the collecting
duct and may cause renal diabetes insipidus.
HMG CoA reductase
inhibitors or statins block the rate-limiting step in cholesterol synthesis and are among
the most widely used drugs. A common side effect of this class of
pharmaceuticals is myopathy, which may range from very frequent myalgia to very
rare rhabdomyolysis. In rhabdomyolysis, the contents of muscle cells leak out
into the bloodstream. Myoglobin, a 17 kDa protein with a heme group to
bind oxygen, is released and filtrated in large amounts. Along with other muscle
proteins, myoglobin is taken up via megalin/cubilin-mediated endocytosis and
thus concentrated in proximal tubule cells. Recall that heme groups with their
central iron are dangerous compounds that enable redox reactions in carefully
regulated systems like cytochrome P450 enzymes or in the mitochondrial
respiratory chain. Here, uncontrolled reactions produce radicals of reactive
oxygen species that over time may result in cell death. So, rhabdomyolysis in
response to statins is at least as much a kidney problem as a muscle problem.
Cisplatin is, of course, an extremely toxic molecule to begin
with, intended to kill tumor cells by crosslinking DNA. This toxicity is
enhanced by accumulation in proximal tubule cells.
Methotrexate blocks dihydrofolate reductase and is intended to
deprive tumor cells of purines and thymidine, the building blocks for DNA
synthesis. Nephrotoxicity is strongly enhanced by crystallization in the
tubulus, which is favored at the acidic pH that is the typical result of our
Western diet high in protein.
Polycystic kidney disease is an umbrella term for
genetic diseases leading to tubule dysfunction and cyst formation. There are
indications that compromised function of the primary cilium may be a common
denominator of various forms of the disease. Almost all cell types have a
single immotile structure protruding from the cell. This primary cilium carries
many receptors and might be seen as a kind of antenna that concentrates signal
reception.
Autosomal dominant polycystic kidney disease is the
most common form with an incidence of 1:500. Affected genes PKD1 and PKD2 encode
transmembrane proteins forming a complex on the primary cilium, which probably
serves as a mechanosensor for the flow rate in the tubule: PKD2 encodes a Ca2+ channel. Already in utero, cysts start to form in a
small percentage of nephrons. Over time, these cysts grow and compress
surrounding parenchyma. An early sign may be a reduction in the kidney's
ability to concentrate urine. The condition may eventually lead to end-stage
renal disease in late adulthood.
Autosomal recessive forms are less common, but tend to be more severe. In polycystic kidney disease, it is especially important
to treat arterial hypertension. As blood pressure contributes to
"inflating" the cysts, it is essential to bring it back to a normal
range. In addition, affected tubule cells are usually sensitive to ADH/AVP,
which may cause enhanced proliferation. Therefore, the -very expensive- V2
antagonist tolvaptan may slow progression in part of the patients.
11. RENAL HANDLING OF CALCIUM AND PHOSPHATE
We will come back to the regulation of calcium and
phosphate when we study bone metabolism. For now, we will concentrate on the renal aspects.
Calcium and phosphate stores of our body are regulated according to different
principles:
·
For
calcium, the main variable regulated to maintain long term stores is uptake
from the intestines. This is accomplished by "switching on" vitamin D
by hydroxylation. This mechanism is too slow to regulate plasma Ca2+,
which needs to be kept in a very narrow range. This is achieved via a
short-term mechanism: if necessary, parathyroid hormone releases Ca2+ from bone. Even this "fast" mechanism requires a few hours to react;
in the meantime, fluctuations in Ca2+ levels are buffered physically
by the bone revervoir. Increased requirement for parathyroid hormone switches on vitamin D
to fill up bone stores.
·
Phosphate
is treated as a precious resource: we take up all we can get and excrete only
the surplus via the kidneys. The main tool to do this is fibroblast growth
factor 23, which goes up in response to an intestinal phosphate load and
stimulates phosphate excretion.
Let's have a look at the system in more detail:
Parathyroid hormone (PTH) is released by the four
parathyroid glands behind the thyroid. An increase in concentration of free Ca2+ activates the calcium-sensing receptor (CaSR)
located at the membrane of their chief cells, throttling PTH production.
Pharmacology
cross-reference: Cinacalcet is a small molecule binding to another site of the calcium-sensing
receptor, allosterically sensitizing the receptor to free Ca2+. Its
main use is in treating secondary hyperparathyroidism in patients with chronic
renal failure
PTH increases Ca2+ concentration via two
main mechanisms: by mobilizing it from bone and by adjusting renal function. In bone, osteoclasts release Ca2+ and phosphate by
local acidification. Not much would be gained by just taking Ca2+ and phosphate from bone: due to their low solubility product, they would quickly
reprecipitate somewhere else in the body.
Therefore, PTH has three effects in the kidney:
1.
It
simultaneously lowers plasma phosphate levels by inhibiting renal reabsorption.
By activating its receptor, PTH stimulates two G
proteins. Gαs stimulates adenylyl cyclase and, via cAMP,
protein kinase A (PKA). Gαq activates phospholipase C and, via
diacylglycerol and Ca2+ release, protein kinase C (PKC). Phosphorylations
by PKA and PKC promote removal of the Na‑Pi cotransporter from the apical
membrane into a vesicle compartment below the membrane, thus strongly reducing
phosphate reabsorption in the proximal tubule.
2.
It
minimizes Ca2+ losses.
At the same time, in the thick ascending limb and the
distal collecting tubule, PTH increases the open probability of the apical Ca2+ channel and induces a Ca2+-buffering protein in the cytosol, thereby increasing Ca2+ reabsorption.
3.
It
switches on vitamin D.
In
proximal tubule cells, PTH induces CYP27B1, the enzyme hydroxylating 25‑hydroxy-vitamin
D at position 1 to produce the biologically active 1,25‑dihydroxy-vitamin
D. Active vitamin D then proceeds to fill up calcium pools from outside.
Lipid-soluble vitamin D3 may be taken up from food,
especially fatty fish, and may be produced in our own skin from
7-dehydrocholesterol with the help of UV-B from sunlight.
Vitamin D, which already contains one hydroxyl group, is
activated by two successive hydroxylations resulting in 1,25-dihydroxy-vitamin
D or calcitriol. The first hydroxylation is performed at position 25, the end
of the side chain, in the liver. The second, at position 1, is performed in the
proximal tubule and is carefully regulated. PTH stimulates hydroxylation, while
the end product calcitriol as well as increased levels of phosphate, via FGF23,
act inhibitory. Calcitriol activates the vitamin D receptor (VDR), a member of
the nuclear receptor superfamily, which is expressed by most cells in our body.
As a ligand-dependent transcription factor, one of its functions is the
induction of genes that are necessary to maintain Ca2+ reserves.
Its main target of filling up calcium pools is
achieved by enhancing uptake of Ca2+ from food in the duodenum. In
the kidney, the action of vitamin D parallels that of PTH by increasing
reabsorption of Ca2+ in the distal tubule, although its effect is
much weaker. Contrary to PTH, vitamin D also enhances reabsorption of phosphate.
Fibroblast growth
factor-23 (FGF23)
FGF23, a member of the large fibroblast growth factor
family of extracellular signaling molecules, primarily regulates serum phosphate
levels. It is produced by osteocytes and osteoblasts in response to dietary phosphate
loading and to 1,25-dihydroxyvitamin D. How bone cells get the message of
intestinal phosphate loading is not yet clear. FGF23 increases renal phosphate
excretion by reducing the number of Na‑Pi cotransporters in the apical
membrane of the proximal tubule. In this function, it acts similar to PTH. Yet,
it counters PTH by inhibiting 1α-hydroxylation of vitamin D; FGF23
downregulates CYP27B1, the enzyme performing 1-hydroxylation, but upregulates an
enzyme which degrades vitamin D. FGF23 is eliminated if it is fully able to do
its job, although we are not yet sure about the precise mechanism. If it is not
entirely successful, FGF23 levels rise, possibly by reduced renal breakdown.
What happens in
chronic kidney disease?
With progressive loss of nephrons, fewer nephrons have
to work harder to get the job done. As eGFR decreases, FGF23 increases to keep
serum phosphate in the normal range. Nephrons are admirable workers: they are
able to redouble their efforts without a hitch. But of course, there is a
limit: once only about a quarter of nephrons are left, they start having
trouble to secrete the necessary amounts of H+, K+ and
phosphate. While dietary phosphate loading continues, FGF23 increases further, but to
no avail: the remaining nephrons excrete all the phosphate filtrated, but that
is not enough. Phosphate slowly creeps up. The low solubility product forces
precipitation of calcium phosphate somewhere in the body, for example in the
media of large arteries, leading to Mönckeberg’s calcific sclerosis.
This reduces aortal windkessel function and leads to left ventricular
hypertrophy and to deterioration of coronary blood flow.
So, phosphate is on the high side, FGF23 is high,
keeping active vitamin D down. Too little Ca2+ is taken up from
outside. How do we keep up Ca2+? The only way to maintain Ca2+ near required levels is to crank up PTH. This is what is being done, leading to
strong secondary hyperparathyroidism. This is not really a good solution: continuous
nibbling on bone leads to renal osteodystrophy or CKD-MBD (chronic kidney
disease- mineral and bone disorder).
How would you go about designing a system to make sure
there is always just the right number of erythrocytes? Maybe we would try to
count and replace the ones we break down or lose. Even if we succeeded in doing
that, it would not do the trick. Once we try to climb a mountain somewhere up
high, say, in the Himalayas, gaspingly, we would soon find out that we need
more erythrocytes than at home. So, the system would have to be able to respond
to changes in ambient pressure, better still, it should as well respond to
longer-term changes in oxygen consumption. To implement such a system, there
would be no way around measuring oxygen concentration somewhere in the body.
Where in the human body would you place an oxygen sensor? Not in the skin, of
course, because the skin receives lots of oxygen from outside; it would tell
you nothing about oxygen transport capacity. Not in the gastrointestinal
system, because blood supply there varies greatly: think of a fright-or flight
reaction. We would have to look for a place deep inside the body with a high,
reliable, constant blood supply. Notwithstanding that, at the same time it
should be a place where oxygen concentration may approach its critical lower
boundary.
Nature placed our oxygen sensor at the renal lcortical-medullar boundary. In
an elegant feedback mechanism, a reduction in partial oxygen pressure
(PO2) in this region stimulates production of new erythrocytes in the bone marrow.
Like most cells, peritubular cells in the renal medulla constantly produce a
transcription factor, hypoxia inducible factor (HIF-1), which at normal PO2 is broken down fast and efficiently. Breakdown via the ubiquitin-proteasome
pathway is initiated by an oxygen-dependent enzyme, a prolyl-hydroxylase. This
enzyme just barely works as long as there is enough oxygen in the cell. As soon
as PO2 sinks below a certain threshold, the enzyme ceases to
function. HIF-1 stops to be hydroxylated, is not broken down any more yet
remains to be constantly produced: so it starts to accumulate. In the nucleus,
accumulating transcription factor HIF-1 activates its target genes, including
the gene encoding erythropoietin (EPO). In summary, this sensing mechanism
translates a fall in renal medullar PO2 into a secretion of EPO.
EPO increases the rate of proliferation in erythroid
progenitors in the bone marrow (colony forming units, CFUE and burst
forming units, BFUE). The net effect is an increase in erythrocyte
output, eventually increasing hematocrit or, in other words, the blood's
oxygen-transport capacity. In time, this leads to normalization of PO2 in the kidney, reactivating the oxygen-dependent hydroxylase. As soon as HIF-1
is degraded again, the negative feedback loop is closed.
Tubulo-interstitial disease interferes with the renal
medulla's ability to produce EPO. Thus, chronic kidney disease is typically
associated with anemia. Previously, dialysis patients had to receive blood
transfusions at regular intervals, over time resulting in iron overload,
infections and transfusion complications.
A big improvement was made in 1989, when the FDA
approved recombinant EPO, developed by the then-small company Amgen with
late-stage help by Johnson&Johnson (J&J). EPO is a 30 kDa glycoprotein
of 165 amino acids, with three N-linked and 1 O-linked carbohydrate chains of
variable structure providing about 40% of its mass. For patients on dialysis,
it was usually administered concomitantly with dialysis two times per week.
To bring recombinant EPO to market, Amgen had entered
into a contractual obligation with J&J that reserved the dialysis market in
the US for Amgen, while all other areas and uses remained the realm of J&J.
EPO became a huge worldwide commercial success, as not only the numbers of
patients on dialysis increased greatly, EPO was also used in cancer patients on
chemotherapy to minimize the duration of anemia.
Soon, recombinant EPO was used for doping in
professional endurance sports like cycling and cross-country skiing, as it was
an undetectable way to increase oxygen transport capacity. Eventually, it was
found that recombinant EPO differed slightly from endogenous EPO in the number
of sialic acid (=N-acetylneuraminic acid/ NANA) residues at the end of the
carbohydrate chains. That way, doping could be identified by isoelectric
focusing of EPO variants from athletes' urine. Post-hoc analysis of stored
urine samples revealed numerous athletes had used the good stuff.
If differences between recombinant and endogenous EPO
exist, don't they lead to antibody formation? There was indeed a scare around
the year 2002. In Europe at that time, there was a drive to replace human serum
albumin, which until then had been used as stabilizer of the injectable EPO
solution in prefilled syringes. Human serum albumin is not produced
recombinantly (too big, too expensive), yet purified from pooled blood plasma from
donors. The recent cases of variant Creutzfeldt-Jakob disease had made
authorities aware of the potential that prions transferred from any pooled
donor material might cause problems later on. With the best of intentions, in
Europe, the stabilizer was therefore reformulated to avoid pooled donor
material: albumin was replaced by the detergent polysorbate 80 plus glycine.
Shortly thereafter, very low, but increasing numbers of patients developed pure
red cell aplasia (PRCA) because of neutralizing antibodies, which inactivated
not only recombinant, but also remaining endogenous erythropoietin. It was
proposed that organic compounds leached from uncoated rubber stoppers in
prefilled syringes containing polysorbate 80 might have an adjuvant effect,
breaking B cell tolerance. Alternatively, it was proposed that increased
formation of EPO aggregates might play a role. As a hypothetical model, EPO
aggregates or EPO-including micelles
formed by polysorbate 80 might cross-bridge several B cell receptors, while
that may not have happened with the old formulation. To address the problem quickly,
several changes were made simultaneously: subcutaneous administration was
stopped, rubber stoppers were Teflon-coated and the cooling chain from
production to patient was improved. With these measures, the problem soon
abated; however, it was not possible to discriminate cause and effect of a
single action. An undisputable lesson seems to be: there is a fine line between
tolerability and antigenicity, and that line is quite unpredictable.
Over time, Amgen developed a molecule they termed
novel erythropoiesis stimulating protein (NESP), which differed from EPO in
that it contained two additional N-linked glycosylation sites. To reach that
goal, five amino acids had to be replaced. Surprisingly, this considerable
change does not lead to more frequent antibody formation. The additional
glycosylation increased serum half-life, allowing less-frequent dosing. A
dispute arose between J&J and Amgen whether NESP was also covered by the
original contract. By arbitration, the dispute was settled in favor of Amgen,
which from then on was able to sell its product in Europe under the generic
name darbepoetin alfa (trade name: Aranesp).
At the peak in the year 2006, worldwide sales of
EPO-related drugs exceeded $10 billion. Then, several studies indicated that there
is no added benefit, even a disadvantage resulting from thrombosis and
hypertension, in trying to raise hemoglobin concentrations to levels seen in
healthy controls. A target range of hemoglobin of 10 g/dl to 12g/dl was
stipulated by FDA and EMEA in 2007 and 2008, which brought down sales in the
following years. In the meantime, the upper limit has been further reduced to
11 g/dl. As patents have expired and the market remains attractive nonetheless,
a range of biosimilars has meanwhile become available in Europe.
After having studied all these renal functions, let's
assemble a picture of a person whose kidneys do not function adequately over a
prolonged period; e.g. in end-stage renal disease (ESRD). We expect to find
symptoms in the following areas:
·
Uremia is the word we use for the combined toxic effect of all those
substances that are normally eliminated via the kidney. One of those substances
is ammonium, NH4+. Apart from ammonium, we measure blood urea
nitrogen and creatinine as surrogate markers, but these two are not toxic at
the concentrations reached. Importantly, we know very little about which other
toxins are responsible for the symptoms encountered. As a complex form of
intoxication, uremia affects many systems. The central nervous system is one of
the first to be affected, leading to symptoms including weakness, fatigue,
problems with memory and concentration, confusion, anorexia, somnolence during
daytime, restlessness at nighttime. Without intervention, the condition will
progress to stupor, coma and death. The peripheral nervous system is affected
with polyneuropathy, the gastrointestinal system with nausea and vomiting, and
the heart with pericarditis.
·
Retention of sodium and water with edema and hypertension, as long as the person
adheres to the usual Western diet. Fluid retention may be alleviated by cutting
back on Na+ intake. In principle, however, at normal water and very
low Na+ intake, hyponatremia may occur, too.
·
Acidosis, again, at the relatively high protein intake typical of our usual diet
·
Hyperkalemia, contributing to weakness, arrhythmia and congestive
heart failure
·
Hyperphosphatemia leading to itching and contributing to bone damage
·
Hypocalcemia, contributing to muscle weakness, cramps and
secondary hyperparathyroidism with chronic kidney disease- mineral and bone
disorder
·
Anemia, resulting in massive weakness and increased strain on the heart
***
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TEXTBOOK SOURCES AND FURTHER READING: in English: Kumar V. et al. (eds.): Robbins and Cotran Pathologic Basis of Disease, 9th Edition, Saunders, Philadelphia, 2015 Boron W. F. and Boulpaep E. L. (eds.): Medical Physiology, 3rd Edition, Elsevier, Philadelphia, 2016 H. G. Rennke & B. M. Denker: Renal Pathophysiology, 4th Edition, Lippincott, Philadelphia, 2014 K. S. Kamel & M. L. Halperin: Fluid, Electrolyte and Acid-Base Physiology, 5th Ed., Saunders, Philadelphia, 2016 in German: Schwarz et al. (Eds.): Pathophysiologie, Maudrich, Wien, 2007 Siegenthaler W. und Blum H. E. (Eds.): Böcker W. et al. (Eds.): Pathologie, 4. Auflage, Urban und Fischer, 2008 |