Download Biochemical Tests of Renal Function

Kidney Function Testing
DR. SAID AL GHORA
An Introduction to the Urinary System
Produces urine
Transports urine
towards bladder
Temporarily store
urine
Conducts urine
to exterior
The Function of Urinary System
A)  Excretion & Elimination:
removal of organic wastes products
from body fluids (urea, creatinine,
uric acid)
B)  Homeostatic regulation:
Water -Salt Balance
Acid - base Balance
C)  Enocrine function:
Hormones
Kidney – basic data
 Urine excreted daily in adults: cca 1.5L
 Kidney only ca 1% of total body weight, despite it
 The renal blood flow= 20% of cardiac output
 Plasma renal flow= PRF ca 600 mL/Min./1.73 M2
 Reflects two processes
 Ultrafiltration (GFR): 180 L/day
 Reabsorption: >99% of the amount filtered
Renal threshold
 Renal threshold of a substance is the concentration
in blood beyond which it is excreted in urine
 Renal threshold for glucose is 180mg/dL
 Tubular maximum (Tm): maximum capacity of the
kidneys to absorb a particular substance
 Tm for glucose is 350 mg/min
Kidney Function
 A plumbers view
Input
Arterial
Filter
Processor
Output
Venous
Output
Urine
How do you know it’s broken?
 Decreased urine
Input
Arterial
production
 Clinical symptoms
Filter
 Tests
Processor
Output
Venous
Output
Urine
Where can it break?
 Pre-renal
Input
Arterial
 Renal
Filter
(intrarenal)
Processor
 Post-renal
Output
Venous
Output
Urine
(obstruction)
Causes of kidney functional disorders
 Pre-renal e.g.
decreased
intravascular volum
 Renal e.g. acute
tubular necrosis
 Postrenal e.g. ureteral
obstruction
Signs and Symptoms of Renal Failure
 Symptoms of Uraemia (nausea, vomiting,





lethargy)
Disorders of Micturation (frequency, nocturia,
dysuria)
Disorders of Urine volume (polyuria, oliguria,
anuria)
Alterations in urine composition (haematuria,
proteinuria, bacteriuria, leukocytouria, calculi)
Pain
Oedema (hypoalbuminaemia, salt and water
retention)
Lab findings
 Rising creatinine and urea
 Rising potassium
 Decreasing Hb
 Acidosis
 Hyponatraemia
 Hypocalcaemia
Why Test Renal Function?
 To identify renal dysfunction.
 To diagnose renal disease.
 To monitor disease progress.
 To monitor response to treatment.
 To assess changes in function that may impact on
therapy (e.g. Digoxin, chemotherapy).
When should you assess renal function?












Older age
Family history of Chronic Kidney disease (CKD)
Decreased renal mass
Low birth weight
Diabetes Mellitus (DM)
Hypertension (HTN)
Autoimmune disease
Systemic infections
Urinary tract infections (UTI)
Nephrolithiasis
Obstruction to the lower urinary tract
Drug toxicity
Biochemical Tests of Renal Function
 Measurement of GFR
Clearance tests
 Plasma creatinine
 Urea, uric acid and β2-microglobulin
 Renal tubular function tests





Osmolality measurements
Specific proteinurea
Glycouria
Aminoaciduria
 Urinalysis







Appearance
Specific gravity and osmolality
pH
osmolality
Glucose
Protein
Urinary sediments
Biochemical Tests of Renal Function
 Measurement of GFR
 Clearance
tests
 Plasma creatinine
 Urea, uric acid and β2-microglobulin
Biochemical Tests of renal function
In acute and chronic renal failure, there is effectively a loss of
function of whole nephrons
 Filtration is essential to the formation of urine  tests of
glomerular function are almost always required in the
investigation and management of any patient with renal disease.
The most frequently used tests are those that assess either the
GFR or the integrity of the glomerular filtration barrier.
Measurement of glomerular filtration rate
GFR can be estimated by measuring the urinary excretion of a substance that is completely filtered
from the blood by the glomeruli and it is not secreted, reabsorbed or metabolized by the renal
tubules.
 Clearance is defined as the (hypothetical) quantity of blood or plasma completely cleared of a
substance per unit of time.
(Uinulin  V)
V is not urine volume, it is urine flow rate
Pinulin
 Clearance of substances that are filtered exclusively or predominantly by the glomeruli but
neither reabsorbed nor secreted by other regions of the nephron can be used to measure GFR.
GFR =
 Inulin
The Volume of blood from which inulin is cleared or completely removed in one minute is
known as the inulin clearance and is equal to the GFR.
Measurement of inulin clearance requires the infusion of inulin into the blood and is not
suitable for routine clinical use
Biochemical Tests of Renal Function
 Measurement of GFR
 Clearance
tests
 Plasma creatinine
 Urea, uric acid and β2-microglobulin
Creatinine
1 to 2% of muscle creatine spontaneously converts to creatinine
daily and released into body fluids at a constant rate.
Endogenous creatinine produced is proportional to muscle
mass, it is a function of total muscle mass the production
varies with age and sex
 Dietary fluctuations of creatinine intake cause only minor
variation in daily creatinine excretion of the same person.
 Creatinine released into body fluids at a constant rate and its
plasma levels maintained within narrow limits  Creatinine
clearance may be measured as an indicator of GFR.
Clinical Significance
Elevated Creatinine is found in
 Impaired renal function
 Very high protein diet
 Vary large muscle mass: body builders, giants, acromegaly
patients
 Rhabdomyolysis/crush injury
 Drugs:
 Trimethoprim
 Amiloride
Clinical Significance
 For renal transplant patients, an increase in serum
creatinine of 2 mg/L has been used as a criterion of
establishing rejection.
 In other persons a change in creatinine of 2 mg/L would
represent a 20% loss in renal function.
Specimen
 One can analyze serum, plasma, or diluted urine.
 The common anticoagulants (fluoride and heparin) do
not cause interference, though heparin, which can be
formulated as the ammonium salt, must be avoided in
enzymatic methods that measure ammonia production.
 Storage


7 days at 4-25oC
At least 3 months at -20oC
Specimen
 Urine should be diluted 1:100
 Bacterial contamination has been found to falsely lower
creatinine values measured using the Jaffé reaction.
 The mechanism of this interference appears to be
bacterial production of a substance that retards the rate
of the Jaffé reaction.
Enzymatic Method
Creatinine aminohydrolase
 Creatinine + H2O
 Creatine +ATP
Creatine
Creatine Kinase
Creatine-P + ADP
Pyruvate Kinase
 ADP + Phosphoenolpyruvate
ATP + Pyruvate
 Pyruvate + NADHLactate dehydrogenaseLactate + NAD+
 The difference in absorbance at fixed times during conversion is
proportional to the concentration of creatinine in the sample
Creatinine Clearance
 Creatinine clearance is used to estimate the
glomerular filtration rate (GFR).
 One method of determining GFR from creatinine is to
collect urine (usually for 24-hours) to determine the
amount of creatinine that was removed from the blood
over a given time interval.
 Clearance is defined as the (hypothetical) quantity of
blood or plasma completely cleared of a substance per
unit of time.
 The most frequently used clearance test is based on
the measurement of creatinine.
 Creatinine is chosen because it is freely filtered at the
glomerulus and is not reabsorbed by the tubules.
 However, a small amount of the creatinine (about 5%)
in the final urine of healthy persons is derived from
tubular secretion.
 To do the test, one needs a precisely timed urine
collection and a blood sample taken during the
collection period.
 Best results are obtained from a 24-h urine collection.
 The test is initiated by having patients empty their
bladder at the beginning of the timed period.
 Urine is collected throughout the period, the bladder is
again emptied at the end of the time period.
 The 'clearance' of creatinine from plasma is directly related to
the GFR if:
The urine volume is collected accurately
 There are no ketones or heavy proteinuria present to interfere with the
creatinine determination.

 It should be noted that the GFR decline with age (to a greater
extent in males than in females) and this must be taken into
account when interpreting results.
Creatinine Clearance
 Creatinine determinations are performed on both
samples. The creatinine clearance is calculated from
the following formula:
 A person has a plasma creatinine concentration of
0.01 mg/ml and in 1 hour produces 60ml of urine with
a creatinine concentration of 1.25 mg/mL.
 Creatinine clearance (mL/min)= (UV)/P X 1.73/S
 where U is urinary creatinine (mg/L), V is volume of
urine (mL/min), P is plasma creatinine (mg/L), S is the
calculated surface area of the patient, and 1.73 is the
surface area (m2) of a standard 70 kg person.
 The range of creatinine clearance in healthy persons
corrected to a surface area of 1.73 m2 is 90 to 120
mL/min.
 At low filtration rates, the creatinine clearance does not
parallel true glomerular filtration rate because a
relatively large portion of the urine creatinine is
secreted rather than filtered.
Effect of Muscle Mass on Serum Creatinine
Normal
Muscle
Mass
Normal
Muscle
Mass
Normal
Kidneys
Diseased
Kidneys
Increased
Muscle
Mass
Reduced
Muscle
Mass
Creatinine
Input
Plasma
Pool
Content
Kidney
Output
Normal
Kidneys
Diseased
Kidneys
Biochemical Tests of Renal Function
 Measurement of GFR
 Clearance
tests
 Plasma creatinine
 Urea, uric acid and β2-microglobulin
Measurement of nonprotein nitrogencontaining compounds
Catabolism of proteins and nucleic acids results in formation of
so called nonprotein nitrogenous compounds.
Protein
 Proteolysis, principally enzymatic
Amino acids
 Transamination and oxidative deamination
Ammonia
 Enzymatic synthesis in the “urea cycle”
Urea
Plasma Urea
Urea is the major nitrogen-containing metabolic product of protein
catabolism in humans,
 Its elimination in the urine represents the major route for nitrogen
excretion.
 More than 90% of urea is excreted through the kidneys, with losses
through the GIT and skin
 Urea is filtered freely by the glomeruli
 Plasma urea concentration is often used as an index of renal glomerular
function
 Urea production is increased by a high protein intake and it is decreased
in patients with a low protein intake or in patients with liver disease.
Plasma Urea
Many renal diseases with various glomerular, tubular, interstitial or vascular damage can
cause an increase in plasma urea concentration.
The reference interval for serum urea of healthy adults is 5-39 mg/dl. Plasma
concentrations also tend to be slightly higher in males than females. High protein diet causes
significant increases in plasma urea concentrations and urinary excretion.
Measurement of plasma creatinine provides a more accurate assessment than urea
because there are many factors that affect urea level.
Nonrenal factors can affect the urea level (normal adults is level 5-39 mg/dl) like:
Mild dehydration,
high protein diet,
increased protein catabolism, muscle wasting as in starvation,
reabsorption of blood proteins after a GIT haemorrhage,
treatment with cortisol or its synthetic analogous
Clinical Significance
 States associated with elevated levels of urea in blood
are referred to as uremia or azotemia.
 Causes of urea plasma elevations:



Prerenal: renal hypoperfusion
Renal: acute tubular necrosis
Postrenal: obstruction of urinary flow





Increased protein catabolism:
Increased dietary protein
Severe tress: fever, etc
Rhabdomyolysis
Upper GI bleeding

Causes of urea plasma decrease


Decreased dietary protein
Increased protein synthesis ( Pregnant women ,
children )
severe liver disease
Overhydration (IV fluids)


Specimen
 Serum and heparinized plasma can be used for the
urease/GLDH methods.
 Fluoride will inhibit the urease reaction; therefore
methods employing urease cannot use serum preserved
with fluoride.
 Ammonium heparin also cannot be used as an
anticoagulant for urease methods.
 Stability in serum or plasma:
 7 days at 4–8°C
 1 year at -20°C
 Because of urea’s susceptibility to bacterial degradation,
serum and urine samples should be kept at 4° to 8° C until
analysis.
Urease/GLDH Method
 The method is optimized for 2-point kinetic
measurement.
 Decrease in absorbance at 340 nm is proportional to
concentration of urea
 BUN / Creatinine Ratio
39








Normal BUN / Creatinine ratio is 10 – 20 to 1
Creatinine is another NPN
Pre-renal increased BUN / Creat ratio
BUN is more susceptible to non-renal factors
Post-renal increased ratio BUN / Creat ratio
Both BUN and Creat are elevated
Renal decreased BUN / Creat ratio
Low dietary protein or severe liver disease
Increased BUN
Normal Creat
Increased BUN
Increased Creat
Decreased BUN
Normal Creat
Uric acid
 In human, uric acid is the major product of the catabolism of the purine
nucleosides, adenosine and guanosine.
 Purines are derived from catabolism of dietary nucleic acid (nucleated cells,
like meat) and from degradation of endogenous nucleic acids.
 Overproduction of uric acid may result from increased synthesis of purine
precursors.
 In humans, approximately 75% of uric acid excreted is lost in the urine;
most of the reminder is secreted into the GIT
Uric acid
Renal handling of uric acid is complex and involves four sequential steps:
Glomerular filtration of virtually all the uric acid in capillary plasma
entering the glomerulus.
Reabsorption in the proximal convoluted tubule of about 98 to 100%
of filtered uric acid.
Subsequent secretion of uric acid into the lumen of the distal portion
of the proximal tubule.
Further reabsorption in the distal tubule.
 Hyperuricemia is defined by serum or plasma uric acid concentrations higher
than 7.0 mg/dl (0.42mmol/L) in men or greater than 6.0 mg/dl (0.36mmol/L)
in women
Greater-than-normal levels of uric acid
(hyperuricemia) may be due to:
 Alcoholism
 Diabetes
 Gout
 Hypoparathyroidism
 Lead poisoning
 Leukemia
 Nephrolithiasis
 Renal failure
 Toxemia of pregnancy
 Purine-rich diet
 Excessive exercise
 Chemotherapy-related side effects
Lower-than-normal levels of uric acid may be due to:
 Fanconi syndrome
 Wilson's disease
 Syndrome of inappropriate antidiuretic hormone
(SIADH) secretion
 Multiple Sclerosis
 Low purine diet
Gout
 Gout is a kind of arthritis that occurs when uric acid
builds up in the joints.
 In Gout increased serum levels of uric acid lead to
formation of monosodium urate crystals around the
joints.
 Acute gout is a painful condition that typically affects
one joint.
 Chronic gout is repeated episodes of pain and
inflammation, which may involve more than one joint.
Specimen
 Serum or plasma may be used; slight but insignificant
positive bias (0.2 mg/dL) has been noted in plasma
specimens as compared with serum.
 Stability in serum / plasma:



6 months at -20°C
7 days at 4-8°C
3 days at 20-25°C
Enzymatic Colorimetric
 Uric acid + H2O + O2
Uricase
Allantion + CO2 + H2O2
 TBHBA + 4- Aminoantipyrine + 2H2O2
POD
Quinoneimine + 3 H2O
 Uric acid is oxidized to allantoin by uricase.
 The generated hydrogen peroxide reacts with 4-
aminophenazone/ESPT to quinoneimine.
Acute Renal Failure
Metabolic features:
 Retention of:




Urea & creatinine
Na & water
potassium with hyperkalaemia
Acid with metabolic
acidosis
Classification of Causes:
 Pre-renal

reduced perfusion
 Renal



inflammation
infiltration
toxicity
 Post-renal

obstruction
Pre-renal versus intrinsic ARF
Test
Result
Pre-renal
Renal
Urea & Creatinine Disproportionate
rise in Urea
Tend to rise
together
Protein in urine
Present on
dipstick
testing
Uncommon
Reference Ranges
49
 BUN
10 - 20
mg / dl
 Creatinine
0.5 - 1.5 mg /dl
 Uric Acid
3.0 - 7.0 mg / dl
 Creatinine Clearance
90 - 130 ml / min
 Ammonia
20 - 60 ug / dl
 BUN / Creat Ratio
10 - 20 to 1
Case Study—Renal Failure
Case Study #1
 HSL is a 63 year-old male with HTN, CAD, and
hyperlipidemia; routine physical examination reveals
asymptomatic hematuria.
 What do you do?
Case Study 2
 Ms. Garcia, a 54 yr old Hispanic female, dx with IDDM for 10
years. Admitted to the hospital with CHF, ESRD, altered lab
values (K+=6.2; BUN 45, Creatinine 3.5; Hgb 6.2; Hct 18.6).
 States that her “breathing keeps getting worse and worse, …can’t
get around, bones break…decreased appetite but keep gaining
weight…funny taste in mouth…blood sugar real high…legs jump
at night”. States that a doctor told her she had “bad kidneys”.
 1. What lab work is typically done?
 2 If ESRD, what lab results would be anticipated?
 3. What S&S of ESRD does Ms. Garcia display and WHY?
 4. What conservative measures might have delayed ESRD?
Discuss dietary, fluid, medications, etc…
Case Study
 1. What lab work is typically done? Chem 12; H&H ; 24 hr creatinine
clearance most helpful;
know normals!
 2 If ESRD, what lab results would be anticipated? ^ BUN, serum
creatinine; low H&H; ^ K+, metabolic acidosis from kidneys inability
to excrete acid load (especially NH3) and from defective reabsorption
of bicarbonate.
Case Study
 3. What S&S of ESRD does Ms. Garcia display and WHY?
Metabolic Acidosis due to decreased ability to excrete acid
metabolites therefore Kussmauls breathing in effort to blow
off excess CO2. musculoskeletal system affected with
renal osteodystrophy as GFR dec., kidney cannot eliminate
phosphate; high phosphate binds with Ca which is drawn
from bone; in CRF, kidneys do not metabolize vitamin D to
its active form which is required for reabsorption of Ca
from intestinal tract; weight gain from Na and water
retention; uremic damage causing peripheral
neuropathy..plus other symptoms including anemia from
decreased production of erythropoietin; and HTN..
 4. What conservative measures might have delayed
ESRD? Discuss dietary, fluid, medications,
etc…Control HTN usually by Na and fluid restriction
and antihypertensives, esp by ace inhibitors; restrict
phosphate intake and use phosphate binders and give
with meals; inc. Ca levels by adm. of active Vit D;
monitor K levels; adm erythropoietin; avoid use of
nephrotoxic drugs…such as aminoglycosides; protein
restriction in diet.
Labs
 Urinalysis
 2+ protein
 3+ occult blood
 >60 RBC per HPF
 Biochemical test
 K+ 4.0
 BUN 19
 sCr 1.5
 Glucose 116
Repeat Labs
 sCr 1.7 (MDRD 45)
 Glucose 88; HBA1c 5.9
 Hct 40.4
 LDL 92
 CrCl 46 ml/min
 24 hr Uprot 741.6 mg
 Renal U/S: normal
 Modification of Diet in
Renal Disease
 One month ago…

sCr 1.6 (49)
 6 months ago…

sCr 1.3 (59)
 One year ago…

sCr 1.5 (54)
To summarize:
1. Use the Creatinine Clearance as the best estimate of
GFR
2. Use the Serum Creatinine to follow renal function over
time
3. Use the Creatinine Index to check the adequacy of a
urine collection
4. Use the BUN to help assess GFR, volume status, and
protein intake