LIVER FUNCTION TESTS. G M Kellerman. Hunter Area Pathology Service
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1 LIVER FUNCTION TESTS G M Kellerman Hunter Area Pathology Service
2 FUNCTIONS OF LIVER Carbohydrate metabolism storage (glycogen), release, synthesis (gluconeogenesis), interconversion (galactose, fructose), recycling (lactate, glycerol, aminoacids) Lipid metabolism lipogenesis (from glucose), scavenging (LDL, IDL, FFA), synthesis (cholesterol, VLDL, apoproteins, choline) Protein metabolism aminoacid interconversions and disposal, urea synthesis, synthesis of many plasma proteins, inflammatory markers (CRP), coagulation factors, transport proteins (Fe, vitamins, hormones) Storage vitamins (B 12, D,A) Detoxication including most drugs (Cytochrome P 450 ) Excretion Bilirubin, drugs, toxins Reticuloendothelial system clearance (from gut absorption)
3 TESTING LIVER FUNCTION A very limited selection of these functions is currently testable in the biochemical laboratory Protein synthesis: albumin as index of nutrition or synthetic capacity, but it can be lost acutely into tissues or chronically into urine or faeces Prealbumin (transthyretin) as index of nutrition Clotting factors, thrombin time, INR Inflammatory markers CRP Iron metabolism ferritin, transferrin Transport proteins Vitamin A, hormones (sex, CORT, TBP) Alpha-1 antitrypsin and other protease inhibitors Alpha-fetoprotein in hepatocellular carcinoma Blockage of bile ducts GGT, ALP, bile acids Integrity of hepatocytes ALT, AST, LD Excretory function bilirubin (conjugated and unconjugated) Storage Vitamins A, E, D, B 12
4 OTHER TYPES OF TEST Chemical tests should be integrated with other tests: Ultrasound can demonstrate dilated bile ducts behind a block Ultrasound can demonstrate primary or metastatic cancers Ultrasound can demonstrate a fatty liver Computerised tomography can help with the above also Signs of portal hypertension splenomegaly, spider naevi, palmar erythema, abdominal veins, oesophageal varices, haemorrhoids Detection of ascites and analysis of the fluid for cells, protein (and fluid/plasma ratios), cell breakdown products (cholesterol, LD), leucocytes and bacteria
5 HEPATOCELLULAR - 1 Disease primarily affecting the integrity of liver cells is characterised by elevations in the transaminases and LD with no or minor increase in GGT and ALP. In cytoplasm ALT>AST so less severe cell damage gives a greater rise in ALT, which is often used as marker of episodes in HepC In the mitochondria AST>ALT, so that when cells die and all contents leak out, AST>ALT However, the half life of AST is normally about hours, and of ALT 36 hours, so that the AST falls much more rapidly and the ratio is of value only very soon after the onset of the problem Transaminases rise in acute illnesses such as hepatitis, infections such as cholangitis, ischaemic damage from cardiac failure or arrest, liver trauma and nonspecifically in many severe illnesses
6 HEPATOCELLULAR - 2 Transaminases are elevated in chronic conditions where liver cells undergo episodic or continuous damage. In most cases ALT>AST and the elevations are relatively mild. Such conditions include steatohepatitis (fatty liver with damage) which is becoming a leading cause of liver pathology in the current epidemic of adiposity Other causes are cirrhosis of any type, chronic hepatitis (B or C are commonest), autoimmune hepatitis, haemochromatosis etc Alcoholic hepatitis is relatively unique as it frequently shows AST>ALT, suggested to be due to patchy necrosis of cells rather than more generalised malfunction. NOTE that in the terminal stages of liver disease there may be insufficient cell mass left to give increased transaminases
7 CHOLESTATIC - 1 When bile ducts are blocked, there is a reaction in the cells behind the blockage that leads to induction of GGT and ALP, levels of which then rise in the plasma. This increase is presumably related to the amount of duct that is blocked, so that complete blockage either of common bile duct or all of the small ducts should lead to the greatest increases Blockage of only part of the biliary tree will therefore lead to a rise in GGT and ALP, but the liver lobules whose ducts are still patent can excrete bilirubin so that there will not be any jaundice Complete blockage can result from stone or carcinoma blocking the common bile duct. Commonest is carcinoma of pancreas or ampulla, or cholangiocarcinoma involving the common bile duct or both of its branches, or metastatic nodes in the porta hepatis
8 CHOLESTATIC - 2 Primary biliary cirrhosis can obstruct all ducts When complete blockage occurs, excretion of bile salts is not possible and they accumulate and itch occasionally measured Partial blockage, recognised by the combination of high GGT and ALP with normal bilirubin, is a feature of metastatic disease in the liver, commonly from GI cancers Both GGT and ALP are also induced by a number of drugs, including alcohol, and moderate increases in their plasma levels up to a few hundred units can occur. This induction does not itself cause liver damage, but it affects drug metabolism and the enzyme changes can be misinterpreted unless the cause is recognised ALP is also secreted by other organs, especially bone and placenta, and isoenzyme studies can be useful
9 MIXED PICTURES It is very common to find a mixed picture with both cholestatic and transaminase enzymes elevated Mild elevations often occur nonspecifically in severe disease of other organs or sepsis Prolonged duct blockage often leads to some cell damage Cholangitis usually damages cells and at least partially blocks some ducts so that all enzymes are elevated Architectural changes consequent on hepatitis and cell necrosis frequently lead to some cholestasis during the recovery period Chronic diseases such as steatohepatitis and developing cirrhosis from any cause often present with a mixed picture
10 BILIRUBIN - 1 Biliverdin is formed by oxidative opening of the porphyrin ring of haem proteins (predominantly haemoglobin) in the RES cells and is oxidised to bilirubin. (C 1 -> CO) Bilirubin is very insoluble and circulates bound to albumin through its 2 anionic side chains. This form is not excreted in urine and is called indirect or unconjugated because it does not react in our analytical methods unless extra reagents are added In the liver there are carrier proteins that release the bilirubin from albumin and transport it into the hepatocyte In the hepatocyte the 2 anionic side chains have glucuronic acid added to form bilirubin di-glucuronide ( conjugated or direct bilirubin, which reacts immediately with our reagents) This reaction is defective in Gilbert s syndrome The bilirubin glucuronide is then excreted across the other side of the cell into the biliary canaliculus and thus into the intestine
11 BILIRUBIN - 2 Normally a trace of the bilirubin glucuronide escapes from the hepatocyte back into the blood, so that normal plasma contains small amounts of both conjugated and unconjugated bilirubin When there is a blockage in the excretory process rarely in the hepatocyte/bile canaliculus membrane, almost always somewhere in the biliary tract then all the conjugated bilirubin diffuses back into the blood and the plasma has a high concentration of conjugated bilirubin which is soluble enough to be excreted in urine Thus haemolytic jaundice with unconjugated bilirubin only, with normal liver function, does not cause bilirubin in urine Obstructive jaundice with conjugated bilirubin has bilirubinuria Hepatic pathology usually has both forms of bilirubin in plasma (proportions vary widely) and therefore bilirubinuria occurs
12 ALBUMIN Decrease in albumin can result from many causes, but they have to be present for some time as its half life is of the order of 3 weeks Decreased protein intake and malnutrition?prealbumin better Decreased synthetic capacity of liver cirrhosis etc Negative acute phase reactant Loss into extracellular space due to increased capillary permeability N.B. This can occur rapidly Large volume saline resuscitation (e.g. in ICU) Ascites Renal loss as in nephrotic syndrome grams per day Intestinal loss protein losing enteropathy e.g. blocked lymphatics Increase in albumin can be a sign of intravascular volume depletion
13 COAGULATION FACTORS All synthesised in liver, 6 of them (Factors II, VII, IX, X, Proteins C and S) are vitamin K dependent and warfarin sensitive Failure to absorb vitamin K can result from steatorrhoea due to any cause. Lack of bile salts due to bile duct blockage results in failure to absorb vitamin K. (Vitamin K synthesis by gut bacteria) Assessment is primarily made by prothrombin time compared to the normal for that set of reagents, normally seconds. Usually calculated as INR for patients on warfarin Levels of clotting factors need to drop to about 10% of normal before coagulation defects appear Diminished levels are a sign of serious hepatocyte loss if adequate vitamin K is available or administered Fibrinogen is also an acute phase reactant
14 OTHER PROTEINS With the exception of alpha fetoprotein, useful as a tumour marker especially for hepatocellular carcinoma, the other proteins synthesised by liver are measured as markers of problems in other systems Ferritin and transferrin in iron metabolism Carbohydrate deficient transferrin as index of ethanol intake Alpha-1 antitrypsin in emphysema (and in faeces for protein loss) Binding proteins in endocrine disorders and vitamin A testing Inflammatory markers in infections Other proteases in prostate cancer, nephrotic syndrome
15 PLASMA ENZYMES - 1 FROM DISEASE IN OTHER ORGANS Almost all cells are capable of anaerobic metabolism, and contain lactate dehydrogenase (LD). Thus this enzyme is non specific. It has 4 subunits, either of the H (heart) or M (muscle) type. H subunits have a long half life about 3 days; M about 15 hours Heart has mostly H 4 or H 3 M, most other tissues are M 4, and red cells have mostly H 2 M 2. These isoenzyme studies are largely obsolete LD is liberated whenever tissue dies, and IN CONJUNCTION with the patient s history can give useful information Myocardial infarction (discussion later), post chemotherapy, most neoplastic diseases before and after treatment, haemolytic disease (and also artefact of bad sample collection), platelet damage (DIC), liver cell damage, muscle damage.
16 OTHER TISSUES - 2 Alkaline phosphatase (ALP) is found in bone, liver, intestine and placenta. The placental enzyme is a separate gene, the others are different in particular by their glycosylation patterns, and antibodies have been made specific for the bone enzyme but are not yet used in general assay methods The placental enzyme is very heat resistant and can be measured when all others have been denatured by 60º for 30 min. It is occasionally useful also as a tumour marker in gut cancer Bone ALP is elevated in childhood and adolescence (growth spurt of puberty), with healing fractures, and in Paget s disease, the elevation depending on the extent and activity. It is a useful marker of response to treatment Liver ALP is increased in cholestatic conditions Occasionally in childhood we see benign hyperphosphatasia
17 OTHER TISSUES - 3 The transaminases are important in the complex cyclic reactions that transfer reducing equivalents (NADH) and acetate from cytoplasm to mitochondria, so they are present in many tissues, of greatest interest to us heart and skeletal muscle The best way to interpret their plasma concentrations is to relate these to the creatine kinase (CK) level, which is present in high concentration in heart and muscle Liver has been discussed, CK is essentially absent or very low In heart, the AST/CK ratio is usually between 10-15%, and the ALT/CK ratio is lower at about 5%, and CK is rarely > 5000 units In skeletal muscle AST/CK is about 2% and ALT/CK is somewhat <1% (and LD/CK is about 5%), but as CK can be in the hundreds of thousands with severe muscle damage, significant rises in these other enzymes can be found. The pattern is most helpful
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