You are on page 1of 47

Biochemistry

of Kidney
Kusumo hariyadi
(KSH)
A normal eGFR is 60 or more. A low
eGFR number may suggest kidney
disease. Use this scale to see what your
eGFR may mean.
A normal eGFR is 60 or more.
A low GFR number may suggest kidney
disease.
Primary urine formed in result of filtration (about
200 L per day).
Reabsorbtion. Lenght of renal tubules is about
100 km. So, all substances important for our
organism are reabsorbed during passing these
tubules.
Epithelium of renal tubules reabsorb per day 179
L of water, 1 kg of NaCl, 500 g of NaHCO3, 250
g of glucose, 100 g of free amino acids.
Reabsorbtion.
All substances can be divided into 3 groups
1. Actively reabsorbed substances
To the first group belong Na+, Cl-, Mg2+,
Ca2+, H2O, glucose and other
monosaccharides, amino acids,
inorganic phosphates, hydrocarbonates,
low-weight proteins, etc.
Na+ reabsorbed by active transport to the
epithelium cell, then into the extracellular
matrix.
Cl- and HCO3- following Na+ according to the
electroneutrality principle,
water according to the osmotic gradient.
Substances go from extracellular matrix to the
blood vessels.
Mg2+ and Ca2+ are reabsorbed with help of
special transport ATPases.
Glucose and amino acids use the energy of Na+
gradient and special carriers.
Proteins are reabsorbed by endocytosis.
GLUCOSE REABSORPTION
LUMEN
Na+ Blood
Na+ Na-K ATP ase
K+ active

glucose
glucose
GLUT 1

electro GLUT 2
no energy req
chemical gradien
Na-K ATPase pump
2. Substances, which are reabsorbed in a
little amount.
Urea and uric acid are little reabsorbable
substances.
3. Non-reabsorbed substances.
Creatinin, mannitol, inulin and some other
substances are non-reabsorbable.
Some substances (K+, ammonia and other) are
secreted into urine in the distal part of tubules.
K+ is changed to Na+ by the activity of Na+-
K+ATPase.
Kidney have a very high level of metabolic
processes.
They use about 10 % of all O2, which used in
organism.
During 24 hours through kidney pass 700-900 L
of blood.
Carbohydrates are the main fuel for kidney.
Glycolysis, ketolysis, aerobic oxidation and
phosphorylation are very intensive in kidney.
A lot of ATP formed in result
Metabolism of proteins also presents in
kidney in high level. Especially, glutamine
deaminase is very active and a lot of free
ammonia formed.
In kidney take place the first reaction of
creatin synthesis.
Kidney have plenty of different enzymes:
LDG (1, 2, 3, 5), AsAT, AlAT. Specific for
kidney is alanine amino peptidase, 3rd
isoform.
Utilization of glucose
Utilization of glucose in cortex and
medulla is differs one from another.
Dominative type of glycolysis in cortex is
aerobic way and CO2 formed in result.
In medulla dominative type is anaerobic
and glucose converted to lactate.
Two sources contribute to the renal ammonia: blood
contribute to the renal ammonia
ammonia (is about one-third of excreted ammonia), and
ammonia formed in the kidney.
The predominant source for ammonia production within
the kidney is glutamine, the most abundant amino acid in
plasma, but a small amount may originate from the
metabolism of other amino acids such as asparagine,
alanine, and histidine.
Ammonia is secreted into the tubular lumen throughout
the entire length of the nephron. Secretion occurs both
during normal acid-base balance and in chronic acidosis.
Metabolic acidosis is accompanied by an adaptive
increase in renal ammonia production with a
corresponding increase in urinary ammonium excretion.
Kidney cortex like liver appear to be unique in that it
possess the enzymatic potential for both glucose synthesis
from noncarbohydrate precursors (gluconeogenesis) and
glucose degradation via the glycolytic pathway
regulate gluconeogenesis in the two organs
1) The liver utilizes predominately pyruvate, lactate and
alanine. The kidney cortex utilizes pyruvate, lactate,
citrate, -ketoglutarate, glycine and glutamine.
2) Hydrogen ion activity has little effect upon hepatic
gluconeogenesis, but it has marked effects upon renal
gluconeogenesis.
Thus, when intracellular fluid pH is reduced (metabolic
acidosis, respiratory acidosis or potassium depletion), the
rates of gluconeogenesis in slices of renal cortex are
markedly increased.
The ability of the kidney to convert certain organic acids
to glucose, a neutral substance, is an example of a
nonexcretory mechanism in the kidney for pH regulation.
Role of kidney in regulation of blood pressure.
Regulation of urine formation.

In response of blood pressure decreasing


juxtaglomerular apparatus of kidney secrete
renin.
This hormone activates angiotensinogen, which
transformed to angiotensin I in liver.
In lung angiotensin I transformed to angiotensin
II strong pressure agent, main actions of which
are vessels contraction and stimulating of
aldosteron secretion by cortex of adrenal glands.
Aldosteron increase Na-reabsorbtion and
decrease quantity of urine.
Na-uretic hormone (produced in heart) decrease
reabsorbtion of Na+, and quantity of urine
increased.

Decreasing of blood pressure is due to activity of


kinines (blood plasma polypeptides, which can
widen vessels and increase permeability of
capillar walls).
Source of kinines is kininogen which undergone
to activity of different proteolytic enzymes.
In result formed kalidin and bradikinin main
substances in this system.
Role of kidney in acid-base balance
regulation.
Kidney have some mechanisms for
maintaining acid-base balance.
Na+ reabsorbtion and H+ secretion play
very important role.
1. Primary urine has a lot of Na2HPO4 (in
dissociated form). When Na+ reabsorbed,
H+ secreted into urine and NaH2PO4
formed.
2. Formation of hydrocarbonates. Inside
renal cells carboanhydrase forms from
CO2 and H2O H2CO3, which dissociated
to H+ and HCO3-. H+ excreted from cell
into urine (antiport with Na+) and leaded
with urine. Na+ connect with HCO3-,
NaHCO3 formed and go to the blood,
thereupon acidity decreased.
BICARBONATE REABSOPTION
LUMEN TUBULAR CELL BLOOD

FILTRAT

Na+
+ + O
- Na Na
HCO3
+
H+ + - -
O H HCO3 HCO
3
H CO H CO3
2 3 2
CA
H O + CO CO + H2O
2 2 2
tight jucntion

Brush Border
3. Formation of free ammonia. NH3 used
for formation of NH4+ (H+ ion associated),
and different acid metabolites excreted as
ammonia salts.
Role of kidney in water balance regulation

Excessive entrance of water leads to dilution of


extracellular fluid. Decreasing of osmolality inhibits
secretion of antidiuretic hormone. Walls of collective
tubules stay non-penetrated to water and dilutive urine
formed.

If volume of blood circulation increases, circulation in


kidney increases also and hyperosmotic medium of
kidney medulla removed. Some substances in these
conditions return into blood. So, excess of water carried
with urine and a lot of soluble substances are
reabsorbed into blood. After water loading stopped,
hyperosmolality in kidney medulla returns for previous
stage during some days.
Physical and chemical characteristics of urine

Urine amount in healthy people is 1000-2000 ml per


day.
Daytime diures is in 3-4 times more than nighttime.
Decreasing of urine amount called oliguria (due to fever,
diarrhea, vomiting, acute nephritis, cardiac insufficiency).
Sometimes anuria occurred (amount of urine per day
less than 50 ml).
The main causes of anuria are leads or arsenics
intoxication, nephrolytiasis, strong stress.
Increasing of urine amount called polyuria (due to
diabetes mellitus, another types of diabetes, usage of
diuretic drugs, some another diseases).
colour of urine
Normal colour of urine is yellow (like hay or amber), what
is due to presence of urochrom (derivate of urobilin or
urobilinogen), some another colour substances are
uroerythrin (derivate of melanin), uroporphyrines,
rybophlavine and etc.
Colour depends from urine concentration.
Colour of urine can be changed due to different diseases
and usage of some drugs. For example, red or pink-red
colour can be due to massive haemolysis (hematuria
and hemoglobinuria occurred) or usage of amidopyrin,
santonin.
High concentration of urobilin and bilirubin gives to urine
brown or red-brown colour.
Green or blue colour can be due to decay of proteins in
the intestine.
Urine characteristic depends from amount of different
salts (oxalates, urates, phosphates), amount of present
epithelium cells and leucocytes.
For differentiation of turbidity origin the following simple
test is used. Add in urine acid product and heat it.
Disappearing of turbidity means that main cause of this
state were urates and phosphates of calcium and
magnesium. If turbidity stays after heating we must think
about inflammatory process in urinary tracts (turbidity in
this case is due to presence of leucocytes, epithelium
cells and another substances).
Density of urine
Density of urine depends from concentration of soluble substances.
Borders of variation are from 1002 to 1035 g/l.
Near 60-65 g of hard substances are excreted with urine per day.
Increasing of density is due to intensive excretion with urine a lot of
organic and inorganic compounds.
Patients with diabetes mellitus have polyuria with increased density
of urine due to presence in urine a lot of solved substances like
glucose, ketone bodies etc.
Patients with diabetes insipidus have polyuria with decreased
density of urine due to insufficiency of antidiuretic hormone and
disorders of water reabsorption in renal tubules.
In case of severe kidney insufficiency urine has density near 1010
g/L (like primary urine). This state is called isostenuria.
pH
In normal conditions urine has acid or weak acid reaction
(pH=5,3-6,8). This depends from presence of NaH2PO4
and KH2PO4. A lot of meat and proteins in diet gives to
urine acid reaction, plants give alkaline reaction. Alkaline
reaction can be due to cystitis, pyelitis, after vomiting.
Expressed acid reaction can be due to diabetes mellitus,
fevers and starvation.

Fresh urine has a specific smell, which is due to


presence of flying acids. But a lot of microorganisms,
which are present in urine, split urea and free ammonia
formed.
Organic compounds of urine.
Proteins. Healthy people excrete 30 mg of proteins per day. As a
rule these are low weight proteins.

Urea. This is main part of organic compounds in urine. Urea


nitrogen is about 80-90 % of all urine nitrogen. 20-35 g of urea is
excreted per day in normal conditions.

Uric acid. Approximately 0,6-1,0 g of uric acid is excreted per day in


form of different salts (urates), mainly in form of sodium salt. Its
amount depends from food.

Creatinin and creatin. Near 1-2 g of creatinin is excreted per day,


what depended from weight of muscles. This is the constant for
each person. Men excrete 18-32 mg of creatinin per 1 kg of body
weight per day, women 10-25 mg. Creatinin is non-reabsorbable
substance, so this test used for evaluating of renal filtration.
Amino acids. Per day healthy person excretes 2-3 g of amino acids
(free amino acids and different low weight molecule peptides). Also
products of amino acids metabolism can be found in the urine.

Couple substances. Hypuric acid (benzoyl glycine) is excreted in


amount 0,6-1,5 g per day. This index increases after eating a lot of
berries and fruits, and in case of proteins decay in the intestines.

Indican (potassium salt of indoxylsulfuric acid). Per day excretion of


indican is about 10-25 g. Increasing of indicans level in urine is due
to intensification of decay proteins in the intestines and chronic
diseases, which are accompanied by intensive decomposition of
proteins (tuberculosis, for example)
Organic acids. Formic, acetic, butyric, -oxybutyric, acetoacetic and
some other organic acids are present in urine in a little amount.

Vitamins. Almost all vitamins can be excreted via kidney, especially,


water-soluble. Approximately 20-30 mg of vit C, 0.1-0.3 mg of vit B1,
0.5-0.8 mg of vit B2 and some products of vitamins metabolism.
These data can be used for evaluating of supplying our organism by
vitamins.

Hormones. Hormones and their derivates are always present in


urine. Their amount depends from functional state of endocrinal
glands and liver. There is a very wide used test determination of
17-ketosteroids in urine. For healthy man this index is 15-25 g per
day.

Urobilin. Present in a little amount, gives to urine yellow colour.

Bilirubin. In normal conditions present in so little amount that cannot


be found by routine methods of investigations.
Galactose. Present in the newborns urine, when digestion of milk or
transformation of glalactose into glucose in the liver are violated.

Fructose. It is present in urine very seldom, after eating a lot of


fruits, berries and honey. In all other cases it indicates about livers
disorders, diabetes mellitus.
Glucose. In normal conditions present in so little amount that cannot
be found by routine methods of investigations.

Pentoses. Pentoses are excreted after eating a lot of fruits, fruit


juices, in case of diabetes mellitus and steroid diabetes, some
intoxication.

Ketone bodies. In normal conditions urine contains 20-50 mg of


ketone bodies and this amount cannot be found by routine methods
of clinical investigations.

Porphyrines. Urine of healthy people contains a few I type


porphyrines (up to 300 mkg per day).
Inorganic compounds of urine
Urine of healthy people contains 15-25 g of inorganic
compounds.

NaCl. Per day near 8-16 g of NaCl excreted with urine.


This amount depends from amount of NaCl in food.

Potassium. Twenty-four hours urine contains 2-5 g of K,


which depends of amount of plants in the food.

Different drugs can change excretion of Na and K. For


example, salicylates and cortikosteroids keep Na and
amplify excretion of K.
Calcium. Twenty-four hours urine contains 0.1-0.3 g, which depends
from content of calcium in the blood.

Magnesium. Content of magnesium in urine is 0.03-0.18 g. So little


amount of calcium and magnesium in urine can be explained by bad
water solubility of their salts.

Iron. Amount of iron in urine is about 1 mg per day.

Phosphorus. In urine are present one-substituted phosphates of


potassium and sodium. Their amount depends from blood pH. In
case of acidosis two-substituted phosphates (Na2HPO4) react with
H+ and one-substituted phosphates (NaH2PO4) formed. In case of
alkalosis one-substituted phosphates react with bases and two-
substituted phosphates formed. So, in both cases amount of
phosphates in urine increases.

Sulfur. Amount of sulfur in twenty-four hours urine is 2-3 g per day (in
form of SO42-).

Ammonia. Ammonia is excreted in ammonium sulfates and couple


substances. Ammonium salts make up 3-6 % of all nitrogen in urine.
Their amount depends from character of food and blood pH.
Pathological components of urine, which are
occur dueOccurs
Glucosuria. to different
in two casesmetabolic
when level of disorders
glucose in bloodin
more than kidney threshold (8-10 mmol/l), so called extrarenal
organism.
glucosuria (diabetes mellitus), and when the kidney cannot reabsorb
even normal quantity of glucose, so called renal glucosuria (kidney
insufficiency).

Ketonuria. Due to some diseases and pathological conditions


(diabetes mellitus, starvation, severe heart weakness, when amount
of fat in the food more than amount of carbohydrates) level of
ketone bodies increases in a big amount (up to 20-50 g per day).
This is the index of deep metabolic disorders, especially in
carbohydrates metabolism.
Bilirubinuria. It occurs in case of hepatic parenhimatous
inflammatory processes or in case of obstruction of gall bladder
ductus. Urine has a colour as dark bear. After some times it stays
yellow-green (bilirubin oxidized to biliverdin).

Urobilinuria. Increasing of amount of urobilin is due to haemolytic or


parenhimatous hepatitis, when decomposition of mesobilinogen in
liver is depressed.

Creatinuria. Amount of creatin in urine increases due to different


pathological processes in muscles like myopathy and myodistrophy,
starvation, hypovitaminosis E, radiation sickness, hyperthyreosis.
Also this is present in small children and in women after delivery.
Indicanuria. Increasing of indicans level in urine is due to
intensification of decay proteins in the intestines, weaken of
intestine peristaltic (atony, constipation), and chronic diseases,
which are accompanied by intensive decomposition of proteins
(tuberculosis, for example).

Phenylketonuria. Innate deficiency of phenylalanine hydroxilase in


liver makes transformation of phenylalalnine in tyrosine impossible.
Amount of phenylalanine in our organism increases more than 10
times and this amino acid utilized by another pathway with
phenylbutyric and phenylacetate formation. These substances
cannot be utilized and heaped up in blood and tissue. Growth of
brain is stopped. With urine a lot of phenylbutyric and phenylacetate
excreted. Fresh urine with FeCl3 gives olive-green colour. This test
is wide spread in maternity homes, because special diet can prevent
problems with child health.
Causes of changes of normal components
content in urine.of urea in urine is due to deficiency of
Urea. Decreasing
protein in diet, disorders of liver functions (especially,
liver cirrhosis, phosphorus intoxication), acidosis
(ammonia used for neutralization of acids), inflammatory
or destructive processes in kidney (nephritis, when urea
is not excreted and uraemia appeared). Increasing of
urea in urine is due to excess of proteins in diet, and
different diseases, which are accompanied by intensive
proteins decomposition (diabetes mellitus, malignant
tumors, infectious diseases with fever).
Creatinin. Amount of creatinin in urine is decrease in case of
disorders of glomerular filtration (amount in blood increases in the
same time). Increasing of creatinin in urine is due to intensive
muscle work, intensive proteins decomposition, excess of creatinin
in diet (meat).

Amino acids. Increasing of amino acids level take place in case of


intensive decomposition of tissues proteins (trauma, burns,
radiation sickness etc.). Also it indicates about liver functions
disorders, especially about depressing of proteins and urea
formation.

Uric acids. Decreasing of uric acid in urine is present, when diet has
mainly carbohydrates without purines. Meat, caviar, gland tissues,
where a lot of nucleoproteins are present, can be cause of
increasing level of uric acid in urine. Leucoses, gout, burns,
radiation sickness, usage of aspirin and corticosteroids also can be
causes of hyperuricuria.
Enzymes. Different enzymes can be present in urine according to
disorders of organs functions and these dates can be used for
precise topic diagnostic.

Inorganic compounds. Concentration of inorganic compounds


depends from their amount in diet (particularly for sodium,
potassium), from character of diet (plants contains a lot of
potassium, and level of this ion in urine increases when diet consists
of plants mainly; meat and other proteins can be cause of increasing
of ammonia salts in urine), from blood pH (in case of alkalosis or
acidosis level of phosphates in urine increases), from some
diseases and physiological stages (during pregnancy and in case of
parathyroid glands hypofunction level of calcium in urine
decreases).

General nitrogen. General nitrogen its a sum of all compounds


which contains nitrogen. Near 80-90 % of general nitrogen belongs
to urea. Another parts uric acid, creatinin and amino acids. So,
level of general nitrogen depends from level of these substances.
Indeces of renal functions
disorders.
Proteinuria. Proteinuria can be kidney and extralidney
origin. Kidney proteinuria is due to damages of
nephrons, when blood plasma proteins can pass through
glomerular membranes. In this case albumins and
globulins are present in urine. Extra kidney proteinuria is
due to damages of urinary tracts and prostate.

Hematuria. This pathological component is due to


damages of kidney or urinary tracts. In most cases
nephrolytiasis is accompanied by hematuria. Some times
hematuria is indicator of traumatical kidney damage.
Glucosuria. In most cases glucosuria is a symptom of diabetes
mellitus, when level of glucose in blood more than kidney threshold
(8-10 mmol/L). But sometimes glucose can be present in urine even
its level in blood is normal. This is so called renal glucosuria which
is due to disorders of glucose reabsorbtion in tubules.

Pyuria. In normal urine leucocytes are present in a very little


amount. Due to different inflammatory processes of urinary tracts,
urine bladder, prostate, also due to nephrolytiasis amount of
leucocytes in urine increases and this situation is called pyuria.

Creatin. Decreasing of creatin in urine is index of kidney


insufficiency.
determination in urine. Clinical significance.

Proteins can be detected in urine after denaturation


(during heating, because proteins under high
temperature denaturated and stay visible, and after
interactions with mineral acids sedimentation with
sulfosalicylic or nitric acid). Quantitative determination of
proteins in the urine can be perfomed by Roberts-
Stolnikov method. The method is based on the known
fact that in adding of urine to nitric acid the white ring is
formed on the border of two solution. This ring is formed
within 2-3 min if the protein contents in urine is 0,033 g/l.
Using variety of dilutions find that one forming the white
ring within 2-3 min.
Glucose can be detected qualitatively with help
of some reductive reactions Felling (the
reaction is based on the reduction of Cu2+ to
Cu+. In this reaction the glucose is oxidized.
Cuprous oxide (Cu+) has red color) or Tromer
reactions, special indicator paper Glucotest,
and quantitatively with help of Althauzens
method (in boiling of mixture of urine containing
glucose with alkaline the different tints of brown
color (from yellow to dark-brown) are formed.
Tint depends from the glucose concentration in
urine).
Ketone bodies can be detected in urine qualitatively by
Legal or Herhardt (in adding of FeCl3 solution to urine
containing ketone bodies the sediment of phosphates is
formed) tests and quantitatively in reaction with acetic
acid, ammonia or sodium nitroprussicum (if urine
contains acetone the violet ring is formed in layer of
ammonia on the urine containing natrium nitroprussicum
and concentrated acetic acid. The velocity of ring
appearance depends on the concentration of acetone in
urine. The appearance of ring between 3 and 4 min
means that concentration of acetone is 0,0085 g/l).
Blood in the urine can be detected qualitatively by benzidin test
(blood peroxidase oxidizes benzidin in the presence of H2O2.
Oxidized benzidin has the dark blue colour).

Fushe (after sedimentation by barium salts bilirubin is oxidized by


FeCl3, which is a part of Fushe reactive, and give blue-green or
blue colour), Gmelin (biliverdin and bilirubin are easy oxidized
substances, which formed after oxydation different coloured
products yellow, red, violet, blue and green), Rosin (bilirubin
oxidized into biliverdin under influence of iodine and gives a green
colour), Florance (urobilin with hydrochloric acid formed a red-
coloured substance) and Rosenbah (modification of Gmelin test)
tests are used for qualitative detection of bile pigments in urine.