Lab Medicine
Conference :
Renal & Liver Function
Tests
Jim Holliman, M.D., F.A.C.E.P.
Professor of Surgery and Emergency Medicine
Director, Center for International Emergency Medicine
M. S. Hershey Medicial Center
Penn State University
Hershey, Pennsylvania, U.S.A.
Lab Medicine Conference :
Renal and Liver Function Tests
Lecture Objectives
–Review renal & liver physiology as it relates
to clinical testing
–Review methodology for RFT's & LFT's
–Discuss indications for obtaining RFT's &
LFT's
–Determine cost-effectiveness of RFT's &
LFT's
Physiology of Creatinine
Is breakdown product of creatine (the
storage source for high-energy
phosphate in muscle cells)
CPK acts to add high energy phosphate
to creatine from ATP
Creatine-phosphate transfers the
phosphate to re-make ATP when energy
is needed for metabolism
Physiology of Creatinine
(cont.)
Synthesis of creatine
–First step (guanidoacetate) occurs in kidney,
small bowel, pancreas, liver
–Second step (methylation of guanidoacetate)
occurs in liver
Distributed throughout body, mainly to
muscle
Total body content relatively constant &
proportional to muscle mass
Metabolic Breakdown of
Creatine
Creatine phosphate undergoes
spontaneous & irreversible breakdown
to creatinine
Converted at constant rate : 2 % of total
body stores per 24 hours
Muscle mass is main determinant of
amount produced
Renal Handling of Creatinine
Found in all body secretions, including CSF
Has no metabolic "usefulness"
Excreted almost entirely via kidneys
Freely filtered at glomerulus
No passive or active reabsorption along
nephron
So, major determinant of serum level is degree
of renal function
Rate of urine flow has no effect on serum level
Renal
handling of
creatinine
Urea Physiology
Major end product of metabolism of
nitrogen-containing substances (mainly
protein)
Generated mainly in liver
–Small amount made in brain
Freely diffusable across cell
membranes except that of urinary
bladder
Renal Handling of Urea
Excreted mainly renally
–Small amounts lost in sweat or metabolized by
gut bacteria
Freely filtered at glomerulus
1/2 of filtered urea reabsorbed in proximal
tubule
Water reabsorption in distal tubule (via
ADH) & collecting ducts increases tubular
luminal concentration of urea
While urea concentration in urine is high,
only 40 to 80 % of filtered urea is excreted
Renal handling of urea
Urine Flow Effects on Urea
Levels
Major determinant of urea reabsorption is
rate of urine flow
–Depends on glomerular integrity & state of
hydration
At high urine flow rates (> 2 ml/min.) : 40 %
of filtered urea is reabsorbed
At lower flow rates, amount of reabsorbed
urea proportionately increases
Urea load filtered varies with degree of
dietary protein intake & tissue breakdown
General Measurement
Methodology for Creatinine &
BUN
Done on serum
Red top tube
Should be run in 2 to 3 hours
No problems related to sample
collection
Free hemoglobin may interfere with
assays
Measurement of Creatinine
Jaffe reaction is standard method
–Red solution results from reaction of
creatinine & picric acid in alkaline medium
–Color change is proportional to amount of
creatinine, & follows Beer's Law
–Reaction is sensitive to temp. & pH
–Pre-Rx with aluminum silicate (Lloyd's
Reagent) improves specificity
Sakaguchi color reaction is alternate
method
Urea Analysis Quantification
Enzyme urease added to specimen
–Catalyzes hydrolysis of urea to carbonic acid
& ammonia
–Amount of ammonia produced is directly
proportional to amount of urea
Ammonia is then quantified
–Automated analyzers used
ƒ React ammonia with alphaketoglutaric acid
ƒ Or, an ammonia - sensing electrode is used
Urea Analysis : Alternate
Method
Diacetyl reaction with urea
–Forms a measureable chromogen
–Simple to perform
–Disadvantages :
ƒ Less specific
ƒ The reagents stink
ƒ Non-linear photometric curve
Normal Reference Ranges
for BUN & Creatinine
BUN :
–8 to 26 mg/dl
–2.9 to 9.3 mmol/liter (International Units)
Creatinine
–0.7 to 1.5 mg/dl
–0.062 to 0.113 mmol/liter
Normal BUN : Creat. ratio :
–8 to 15 : 1
Azotemia
Represents abnormal condition in
which the non-protein nitrogenous
(NPN) compounds of urea & creatinine
are elevated
Classed as :
–Prerenal
–Renal
–Postrenal
General Causes of
Hyperuremia
(BUN > 26)
Prerenal azotmia
Postrenal azotemia
Renal dysfunction
Increased protein load to liver
–Endogenous
–Exogenous
Prerenal Azotemia
Functional integrity of nephrons
maintained
Due to :
–Inadequate renal perfusion
ƒ Dehydration
ƒ Shock
ƒ Blood loss
ƒ Congestive heart failure
ƒ Renal artery stenosis
–Or, increased NPN production
Prerenal Azotemia :
Causes of Increased NPN
Production
Endogenous
–GI hemorrhage
–Catabolic states
–Antianabolic medications (steroids,
tetracycline)
–Cancer chemoRx
Exogenous
–Increased protein intake
Causes of Postrenal Azotemia
Generally due to urinary tract obstruction
& stasis of urine flow
–Renal vein thrombosis
–Bilateral ureteral stricture, calculi, or
compression
–Prostatic hypertrophy or tumor
–Bladder obstruction
ƒ Tumor
ƒ Trauma
ƒ Stone or foreign body
ƒ Autonomic dysfunction (spinal cord
dysfunction)
Causes of Renal Azotemia
Due to renal insufficiency or failure due
to intrinsic renal disease
Not reversed by correcting pre- or postrenal problems
Etiologies :
–Acute tubular necrosis
–Acute interstitial nephritis
–Nephrotic syndrome
–Collagen vascular diseases
–Malignancy
–Metabolic diseases (esp. diabetes)
Mechanisms Causing Increased
NPN
Compounds with Renal Disease
Renal vasoconstriction / decreased
renal blood flow
Urine stasis from tubular obstruction by
debris
Back leakage of filtrate into blood
Decreased glomerular permeability &
GFR
Shunting or redistribution of renal
blood flow resulting in decreased GFR
Causes of Hypouremia
(BUN < 6 mg/dl)
Physiologic
–Newborn
–Pregnancy (increased GFR & urine flow)
–Overhydration
–Decreased protein intake
Pathologic
–Acute or chronic liver disease
General Factors Affecting
the BUN Level
BUN is dependent on :
–Protein intake
–Functional integrity of kidneys
–Functional integrity of liver
–Urine flow rate
Causes of Elevated Creatinine
Levels (> 1.5 mg %)
Intrinsic renal disease
Mild elevations from pre- or post- renal
azotemia
(Transiently) from ingestion of large
amounts of meat
Extensive muscle trauma
Muscle wasting diseases (MD, ALS,
myasthenia gravis)
Factitious (lab assay interference)
Causes of Factitious
Elevations
of Creatinine Levels
Ketone bodies
Hyperglycemia
Other proteins
Barbiturates
Penicillins
Cephalosporins
Methanol
Causes of Low Creatinine
Levels
(< 0.7 mg %)
Basically due to decreased muscle mass :
–Children
–Females
–Pathologic : later stages of muscle-wasting
diseases
Indications to Check
BUN & Creatinine
Assess dehydration not obvious by physical
exam
Differentiate renal vs. pre- or post- renal
azotemia as cause for decreased urine output
Indicate presence of "occult" blood in upper GI
tract
Verify renal function O.K. prior to dye studies,
surgery, or nephrotoxic Rx
Evaluate for transplant rejection
Monitor for ongoing nephrotoxic drug effect
Situations NOT Requiring
Checking BUN & Creatinine
Dehydration in healthy adults from
gastroenteritis
Preop in healthy adults for simple
abdominal or orthopedic surgery
Uncomplicated UTI's
Uncomplicated respiratory tract and
head & neck infections
Mild to moderate back trauma without
hematuria
Clinical Situations Requiring
Periodic
BUN & Creatinine Monitoring
Aminoglycosides
Amphotericin
ACE inhibitors
Moderate to severe hypertension
Diabetes
Structural renal disease (polycystic, etc.)
Renal transplant
Rhabdomyolysis
Lab Charges at Hershey Med
Center for BUN / Creatinine
Both together ("renal profile") : $11.00
Either separate : $ 11.00
"SMA-7" : $ 12.00
Stat fee (for E.D. or inpatients) : 11/2
Times the above $
Laboratory Evaluation of Liver
Disease :
Topics Covered
Enzymes
–Alkaline phosphatase
–Aminotransferases (transaminases)
–Lactate dehydrogenase (LDH)
Bilirubin
–Direct
–Indirect
Serologies for viral hepatitis
Alkaline Phosphatase (ALP)
Physiology
Is heterogeneous group of enzymes catalyzing
same reaction using different substrates
Hydrolyze phosphomonoesters to alcohol &
inorganic phosphate at alkaline pH
 Play role in transport of sugars & phosphates :
–Intestinal mucosa
–Renal tubules
–Bone
–Placenta
Isoenzymes exist but difficult to separate
Causes of Increased ALP
Activity
in Serum
Physiologic
–Rapid growth periods in children (ages 5 to 14
years)
ƒ Value is 2 to 3 X normal
–Pregnancy
–Aging
–Post fatty meal
Pathologic Causes of Increased
ALP Activity in Serum
Hepatic lesions
–Acute hepatitis, mononucleosis, cirrhosis,
cholestasis
Osteoblastic lesions
–Hyperparathyroidism, Rickets, Paget's, fractures,
tumors
Tumors
–Ectopic production
Gastrointestinal lesions
–Stomach, duodenal, or colon ulcerations
Infarcts
–Cardiac, pulmonary, renal, spleen
Physiology of Aminotransferases
(Transaminases)
Catalyze reversible transfer of amino group from
an alpha amino acid to an alpha keto acid
Results in formation of oxaloacetic & pyruvic
acids
2 main ones in serum :
–Aspartate aminotransferase (AST)
ƒ Formerly glutamate oxaloacetic transaminase
(GOT)
–Alanine aminotransferase (ALT)
ƒ Formerly glutamate pyruvate transaminase
(GPT)
Aspartate Aminotransferase
(AST)
Found in heart, liver, skeletal muscle, brain,
kidney
Catalyzes transfer of amino group from
aspartate to alpha ketoglutarate
Present in both mitochondria & cytosol
Present in serum as both apoenzyme &
holoenzyme (i.e., with & without cofactor
pyridoxal-5-phosphate; same as for ALT)
Currently measurement of AST isoenzymes
not clinically useful
Causes of Elevated AST
Levels
Myocardial infarction
Acute hepatic necrosis
Pulmonary infarction (mild elevations in
30 %)
Congestive heart failure (passive liver
congestion)
Pericarditis
Rheumatic fever
Skeletal muscle injury
Alanine Aminotransferase
(ALT)
Localized primarily in liver
Catalyzes transfer of amino group from
alanine to alphaketoglutarate
Is specific marker for hepatic disease or
injury
Only in cytosol (not in mitochondria)
Lactate Dehydrogenase (LDH)
Physiology
Catalyzes the reversible reaction :
–lactate + NAD
pyruvate + NADH
Maintains balance between anabolism &
catabolism of carbohydrates
In liver is involved in gluconeogenesis &
glycogen synthesis from lactate
In heart enables lactate to enter citric acid
cycle & be used as fuel to generate ATP &
NAD
Most tissues have high quantities of LDH
Isoenzymes of LDH
Are tetramers made of 4 subunits
containing one of 2 tissue types : H
(heart) or M (skeletal muscle)
There are 5 isozymes of LDH which
consist of combos of the monomers
Normal LDH activity in serum is mainly
of erythrocyte origin
Isoenzymes of LDH
TYPE
MONOMERS
ORGAN
LOCATION
LDH 1
HHHH
Myocardium,
erythrocytes
LDH 2
HHHM
Myocardium,
erythrocytes
LDH 3
HHMM
Brain, kidney, less
in liver & muscle
LDH 4
HMMM
Liver, brain, kidney,
muscle
LDH 5
MMMM
Liver, muscle,
less in kidney
Conditions with Increased
LDH Levels
Cardiac
–Myocardial infarction
–CHF
–Pulmonary infarction
Hematologic
–Megaloblastic anemia
–Sickle cell disease
–Hemolytic anemia
–Leukemias
–Lymphoma
–Infectious mononucleosis
Conditions with Increased
LDH Levels (cont.)
Hepatic
–Hepatitis
–Obstructive jaundice
–Cirrhosis
–Metastatic tumors
Skeletal
–Muscular dystrophy
–Delerium tremens
LDH Isoenzyme Patterns
in Different Conditions
DISEASE
Acute MI
Meg.anemia
Hem.anemia
Mus.dystro.
Leukemia
Pancreatitis
Ca. mets
Pulm infarct
C.H.F.
Hepatitis
Cirrhosis
LDH-1
LDH-2
+
+
+
+
+
+
+
+
LDH-3
+
+
+
LDH-4
LDH-5
+
+
+
+
+
+
+
Measurement Methodology
for Liver Enzymes
ALP
–Rate of conversion of p-nitrophenylphosphate (p-NPP)
to p-nitrophenol (p-NP) in presence of buffer AMP
–Change in absorbance at 405 nm due to formation of
p-NP is proportional to ALP activity
AST, ALT
–Change in absorbance at 340 nm due to
disappearance of NADH is proportional to AST & ALT
activity
LDH
–Change in absorbance at 340 nm due to appearance of
NADH is proportional to total LDH activity
–LDH isozymes separated electrophoretically & stained
General Diagnostic
Interpretation of AST & ALT
Levels
ALT is most specific measure of hepatocellular
damage (necrosis)
Highest AST & ALT levels occur with :
–Acute viral hepatitis
–Toxin - induced hepatic necrosis
–Circulatory shock
Damage to as little as 1% of liver cells raises ALT
AST & ALT rise 7 to 14 days before jaundice
AST elevation can be screen for Reye's Syndrome
General Interpretation of
AST & ALT Levels (cont.)
Degree of elevation not necessarily related to
severity of disease process
Levels < 500 U/liter usually mean mild illness
Ratio of AST : ALT > 2 highly suggestive of
alcoholic hepatitis (unless ALT > 300, then
this does not apply)
LDH usually normal or only slightly elevated
with hepatitis or obstructive jaundice
Patterns of Enzyme Elevations in
Liver and Biliary Diseases
DISEASE
ALP
AST
ALT
LDH
Acute Liver Injury
4 - 10 X
>20 X
>20 X
+/-
Alcoholic hepatitis
2-4X
4 - 10 X
2-4X
+/-
Infectious
Mononucleosis
Cholestatic
jaundice
2 - 10 X
10 - 20 X
10 - 20 X
10 - 20 X
4 - 10 X
4 - 10 X
Primary or Secon.
cancer
10 - 20 X
4 - 10 X
4 - 10 X
4 - 20 X
Primary biliary
cirrhosis
10 - >20 X
4 - 10 X
4 - 10 X
2-4X
Alcoholic fatty
liver
2-4X
2-4X
+/-
Cirrhosis
2-4X
2-4X
2-4X
2-4X
Chronic active
hepatitis
2-4X
10 - 20 X
4 - 10 X
2-4X
2 - 10 X
+/-
+/-
Time pattern of serum transaminases
Bilirubin Metabolism
Originates from breakdown of heme (from
hemoglobin, myoglobin, & cytochromes) into
biliverdin which is reduced to form bilirubin
Can be produced by most cells
Free (unconjugated) bilirubin enters plasma from
sites of production
–Is tightly bound to albumin
–Not filtered at glomerulus (not excreted in urine)
–Taken up by hepatocytes
–Conjugated in microsomes by enzyme bilirubin
glucuronyl transferase
–Bilirubin diglucuronide (water soluble) then
excreted via bile
Disposition of Conjugated
Bilirubin
Enters intestine via bile
Further reduced by colonic bacteria to
stercobilinogen / urobilinogen, which is
spontaneously oxidized to brown bilin
pigment (accounts for normal stool color)
Some of this pigment undergoes
enterohepatic cycling
Trace amounts excreted in urine as
urobilinogen, which autooxidizes to urobilin
"Direct" versus "Indirect"
Bilirubin
"Indirect" = unconjugated (non- liver
metabolized)
–Is nonmiscible with aqueous diazonium salts
–So solvent such as methyl alcohol is needed
to render it water soluble, permitting a color
reaction
"Direct" = conjugated (acted upon by liver cells)
–Reacts directly with diazo reagents to make a
measureable color change
Normal serum total bilirubin is 0.5 to 1.2 mg/dl (<
20 % unconjugated)
Bilirubin
metabolism
Bilirubin
metabolism in
hepatic disease
Bilirubin
metabolism in
extra- hepatic
obstruction
Causes of Jaundice from
Unconjugated
Hyperbilirubinemia
Pigment loading
–Hemolytic anemia
–Extravascular blood (surgery or trauma)
–Liver disease (unable to conjugate)
Gilbert's Syndrome
–Usually benign
–Bilirubin levels elevate with fasting
Crigler-Najjar Syndrome
–If homozygous is severe & needs liver
transplantation
Jaundice from Conjugated
Hyperbilirubiinemia
Usually reflects cholestasis
–Retention of bilirubin & bile salts
–Can be intra- or extra- hepatic cause
–Urine is dark brown (from conjugated
bilirubin)
–Urine froths if shaken (from detergent action
of bile acids)
–Patients often have pruritis from bile acids
Intrahepatic Causes of
Conjugated
Hyperbilirubinemia
Hepatocellular injury
Biliary atresia
Primary biliary cirrhosis
Steroids (especially estrogens)
Space - occupying hepatic lesions
Dubin-Johnson Syndrome
Extrahepatic Causes of
Conjugated Hyperbilirubinemia
Choledocholithiasis
Caroli's Disease
Postoperative biliary tract strictures
Sclerosing cholangitis
Cholangiocarcinoma
Pancreatitis
Ampullary or pancreatic cancer
Compression from adjacent cysts
Parasites
Other Tests to Consider for
Evaluation of Liver Disease
Protime
–Measures presence of liver-synthesized vitamin K dependent
factors II, VII, X
–Factor VII has half life < 12 hours
–Indicates significant liver dysfunction if prolonged > 2
seconds
Serum albumin
–Normal level 3.5 to 5 g/dl
–Synthesized exclusively in liverMax. synthesis is 25 g/day
(half life is up to 20 days)
Serum globulin
–Levels often > 2 g/dl (normal < 1.1 g/dl) with chronic liver
disease
Gamma glutamyl transpeptidase
Gamma Glutamyl Transpeptidase
(GGTP)
Found in liver, kidney, pancreas, heart,
brain
Elevates in cholestatic disorders
Inducible by many drugs
Half life 26 days
If GGTP level is normal, it suggests a
concurrent ALP elevation is from bone or
placenta
Can be elevated in non-liver disorders
(other LFT's are then normal)
Additional Tests to Consider To
Rule Out Specific Liver
Disorders
Alpha-1-antitrypsin level
–Rule out alpha-1-antitrypsin deficiency
–These patients can have CAH & COPD
–Is autosomal recessive ; relatives should be
screened
Serum ceruloplasmin
–Rule out Wilson's Disease
–Should confirm with liver biopsy
–Is treatable
Serum iron / TIBC
–Rule out hemochromatosis ; also treatable
Elevated AST
Algorithm for evaluation of elevated alkaline phosphatase
Hepatitis A Serologies
Hepatitis A IgM antibody (IgM anti-HAV)
–If positive, represents current or recent
acute hepatitis A
–Persists typically 4 to 6 months (but up to
12) post infection
Hepatitis A total antibody (total antiHAV)
–Tests IgM & IgA early, and mainly IgG later
Time course of hepatitis A serologies
Hepatitis B Serologies
Antibody to hepatitis B surface antigen
(anti-HBs)
–If present indicates :
ƒ Prior hepatitis B, now immune
ƒ Or prior hepatitis B vaccination
ƒ Or recent hepatitis B immune globulin
prophylaxis
–If HBsAg also present, indicates chronic
hepatitis B (carrier)
Hepatitis B surface antigen (HBsAg)
–If present, indicates acute or chronic hepatitis
B and patient is infectious
Hepatitis B Serologies (cont.)
Hepatitis B e antigen (HBeAg)
–If present, indicates acute or chronic
hepatitis B with active viral replication
Antibody to hepatitis B e antigen (antiHBe)
–Indicates suppression of hepatitis B viral
replication
Hepatitis B Serologies (cont.)
Hepatitis B core IgM antibody (IgM antiHBc)
–Indicates current or recent hepatitis B (in past 4
to 6 months), or chronic hepatitis B with active
viral replication (less common)
Hepatitis B core total antibody (total antiHBc)
–Just indicates prior hepatitis B infection, but
does not indicate infectivity or chronicity
Time course of acute hepatitis B serologies
Time course of chronic hepatitis B serologies
Serologies with hepatitis D superinfection
Serologies with acute hepatitis D coinfection
Hepatitis C Serologies
Hepatitis C antibody by enzyme immunoassay (antiHCV by EIA)
–Indicates chronic hepatitis C ( rarely detectable for
acute hepatitis C)
Hepatitis C antibody by recombinant immunoblot
assay (anti-HCV by RIBA)
–Indicates chronic hepatitis C (useful for evaluating
suspected false positive anti-HCV by EIA)
Hepatitis C RNA by polymerase chain reaction (HCV
RNA by PCR)
–Indicates acute or chronic hepatitis C
These antibodies do not confer protection against
infection
Lab Charges for Liver
Function Tests at Hershey
Med Center
"LFT panel" (ALP, AST, total bili) : $18
ALT alone : $10
AST alone : $10
Hepatitis serologies :
–HCV antibody : $42
–HAV antibody : $37
–HBc antibody : $30
–HBsAg :
$30
–anti-HBs :
$43