Value of Liver Function Tests in Cirrhosis

Abstract

The liver comprises both parenchymal and non-parenchymal cells with varying functions. Cirrhosis is often complicated by the development of portal hypertension and its associated complications. Hence, assessment of liver in cirrhosis should include assessment of its structural, function of both hepatic and non-hepatic tissue and haemodynamic assessment of portal hypertension. There is no single test that can evaluate all functions of liver and assess prevalence and severity of portal hypertension. Commonly available tests like serum bilirubin, liver enzymes (alanine [ALT] and aspartate aminotransferase [AST], serum alkaline phosphatase [ALP], gamma glutamyl transpeptidase [GGT]), serum albumin and prothrombin time for assessment of liver functions partly assess liver functions. quantitative liver functions like indocyanine clearance tests [ICG-K], methacetin breath test [MBT] were developed to assess dynamic status of liver but has its own limitation and availability. Imaging based assessment of liver by transient elastography, MRI based 99 mTc-coupled asialoglycoprotein mebrofenin scan help the clinician to assess liver function, functional volume of liver left after surgery and portal hypertension [PH]. Hepatic venous pressure gradient still remains the gold standard for the assessment of portal hypertension but is invasive and not available in all centres. Combinations of blood parameters in form of various indices like fibrosis score of 4 [FIB-4], Lok index, scores like model for end stage liver disease (MELD) and Child-Turcotte Pugh score are commonly used for assessing liver function in clinical practice.

Incidence of chronic liver disease (CLD) and its associated morbidity and mortality has increased over time and this had led to significant burden on health care system.^1,2 Alcohol-associated liver disease, non-alcoholic fatty liver disease (NAFLD) and viral hepatitis like hepatitis B and C are the commonest causes of liver disease.3, 4, 5 Other causes include autoimmune liver disease, haemochromatosis, primary biliary cholangitis (PBS), primary sclerosing cholangitis (PSC) and Wilson disease. Majority of chronic liver disease develops slowly and there may be no signs or symptoms in the initial phase. Patients present only when complications of end-stage liver disease (ESLD) develop like jaundice, ascites and other features of portal hypertension like variceal bleed. Natural history of cirrhosis is characterised by an early asymptomatic phase, termed ‘compensated’ cirrhosis followed by the development of complications of portal hypertension and/or liver dysfunction, termed ‘decompensated cirrhosis’. Patients with compensated cirrhosis live significantly longer than that of decompensated patients with median survival times of >12 years and <2 years, respectively.^6,7

The liver comprises both parenchymal and non-parenchymal cells with varying functions. Common functions of liver include glucose, fat, protein metabolism, coagulative functions and metabolism and excretions of number of substances, immune functions and regeneration of diseased liver.⁸ Cirrhosis is often complicated by the development of portal hypertension and its associated complications. So ideally assessment of liver by blood test should include assessment of its structural, function of both hepatic and non-hepatic tissue and haemodynamic assessment of portal hypertension (Figure 1). With so many functions, the liver has traditionally been difficult to assess by non-invasive means. Liver biopsy till now considered as gold standard for assessment of structure but it is not feasible in clinical practise. Liver biopsy has its own limitations and subject to inter and intra observer variability and it gives no information of functional component of liver, nor does it give any information about portal hypertension.^9,10 Therefore, tests for liver functions should also incorporate tests that evaluate portal hypertension. Liver assessment by simple test done routinely may miss chronic liver disease in its initial stages as majority of tests done for it may be normal or reveal subtle changes in total bilirubin, liver enzymes and serum albumin which may be overlooked.¹¹ However; in late stages of liver disease liver function tests—bilirubin, albumin, international normalised ratio (INR) and platelet count may be abnormal in majority of these patients. Ahmed et al¹² evaluated the stage of fibrosis with aspartate aminotransferase (AST), alanine aminotransferase (ALT), international normalised ratio (INR), blood urea nitrogen (BUN), creatinine, platelets, alkaline phosphatase, bilirubin, and albumin in a large retrospective observational study on 771 liver biopsies. They concluded that these markers remained near normal till late in patients with cirrhosis and cannot differentiate early cirrhosis stage versus late except AST.

Figure 1.

Assessment of liver.

The term “liver function tests” is not appropriate as it does not evaluate vast functions of liver rather it only evaluates liver injury (AST/ALT), synthesis (prothrombin ration, albumin) and cholestasis [serum bilirubin, serum alkaline phosphatase(ALP) and gamma glutamyl transpeptidase (GGT)]. These biochemical liver functions are objective, indirect markers, static and are subject to variation due to variables like acute infection, sepsis, biliary tract obstruction and deficient vitamin K state. Dynamic tests of liver function have emerged to provide objective, reliable and simple means of assessing parenchymal liver function and its reserve both before and after surgery. However, these tests are also subject to variation according to hepatic blood flow and are not easily available.¹³ Injury to liver causes alteration in both functional as well as structural organisation. Hence a liver function test should be able to identify the presence of liver parenchymal or biliary, assess its severity and should provide prognostic information. These functions should be able to assess dynamic changes in structure and function at the level of cellular change and its synthetic activity.

Assessment of functions of non-parenchymal cells which includes clearance of pathogens and pathogen associated molecular patterns, antigen presentations, cytokines release and clearances of inflammatory mediators, sinusoidal perfusion abnormalities etc. has not been evaluated by most of liver blood tests done in clinical setting.^14,15 Hence no single test can evaluate complete liver functions and a battery of tests can only partly evaluate liver functions. Optimal requirement of an ideal liver function tests for a clinician is that it should be sensitive to mild abnormalities of liver functions, easy, affordable, can be performed at the point of care, non-invasive, provides real-time results, does not require patient cooperation and expertise from the provider, good correlation with hepatic venous pressure gradient (HVPG) and liver biopsy and should correlates with long term clinical endpoints in patient care.

What are liver blood tests?

Liver blood tests are a battery of test which are easily available in biochemical laboratory and varies from hospital to hospital and country to country.^14,15 Traditionally called as liver function tests (LFTs) in majority of biochemical labs but in true sense these tests partly evaluate liver function. Hepatobiliary enzymes like aspartate aminotransferase (AST) and alanine aminotransferase (ALT) convey information on hepatocytes injury, whereas bilirubin, albumin and international normalised ratio (INR) convey information on liver function; and platelets number gives information on the degree of portal hypertension (PHT) with varying sensitivity and specificity.¹⁵

Serum Bilirubin

Bilirubin is a breakdown product of the haem, and it exists in two forms, unconjugated and conjugated. Bilirubin in its insoluble unconjugated form is transported to liver loosely bound to albumin, where it is converted into soluble conjugated bilirubin for excretion in bile. Hence bilirubin measurement estimates excretion and conjugation capacity of liver. Bilirubin production increases in haemolysis, ineffective erythropoiesis and resorption of a large haematoma and inherited disorders of conjugation. Unconjugated form of bilirubin in such cases does not reflect underlying liver injury by itself. Disproportionately isolated, unconjugated hyperbilirubinaemia with low serum alkaline phosphatase (ALP) may also be seen in Wilson disease. Conjugated hyperbilirubinaemia characteristically occurs in parenchymal liver disease and biliary obstruction.^15,16

Clinical relevance of bilirubin in cirrhosis

Total bilirubin level contributes to various prognostic models in the assessment of chronic liver disease like Child –Turcotte Pugh (CTP) and model for end-stage liver disease (MELD). Elevated bilirubin level>5 mg/dL is included in the definition of acute-on-chronic liver failure (ACLF) as per Asian pacific association for the study of liver (APASL) guidelines. Elevated bilirubin level is also included in the guideline of definition of organ failure (hepatic) by both APASL and European association for study of liver (EASL).^17,18 Total bilirubin alone can predict short-term mortality in patients with ACLF, acute liver failure, alcoholic hepatitis, and primary biliary cholangitis (PBC). It has a linear relationship with 90-day mortality in patients waiting for liver transplantation.19, 20, 21 Early change in bilirubin level at day 7 is a very simple predictive factor for identifying non-responders to steroids in patients with severe alcoholic hepatitis.²² Predominant increase in conjugated bilirubin with elevated alkaline phosphatase (ALP) and gamma-glutamyl transferase (GGT) suggests chronic liver disease secondary to primary biliary cholangitis (PBC) and primary sclerosing cholangitis (PSC) or overlap syndrome between autoimmune hepatitis (AIH) with either PSC or PBC.^15,16

Aminotransferase

Persistent elevation in aspartate aminotransferase (AST) and alanine aminotransferase (ALT) is a marker of hepatocyte injury and is the commonest abnormality seen in patients with underlying liver disease.¹⁵ ALT is considered a more specific marker to liver disease than AST as AST may be elevated due to muscle injury or cardiac injury, the concentration of AST may be a more sensitive indicator of liver injury in conditions such as alcohol-related liver disease and in autoimmune hepatitis (AIH).Elevated aminotransferase levels suggest hepatocytes injury however their absolute value do not predict prognosis in patients with cirrhosis though aspartate aminotransferase to alanine aminotransferase ratio (AAR) is a simple and reliable marker to assess liver function and hepatic fibrosis as well as to predict outcomes in patients with primary biliary cholangitis (PBC).^23,24

Clinical Relevance of AST/ALT Elevation in Cirrhosis

There is poor correlation between ongoing liver injury as assessed by liver biopsy and elevation in the level of in these enzymes.²⁵ AST and ALT are still used in the initiation of treatment, monitoring of response, remission and relapse, and in aiding decisions to discontinue treatment when enzymes have normalised for a prolonged period in patients with hepatitis B, autoimmune hepatitis and overlap syndrome.^26,27 Persistently raised transaminase level despite treatment, indicate a high risk of disease progression. The AST/ALT ratio as an indicator of cirrhosis in patients.²⁸High level>1000 IU/L are normally not seen in patients with cirrhosis and if seen then ischaemic liver injury, acute viral hepatitis, and drug- or toxin-induced liver injury and occasionally in acute obstructing choledocholithiasis should be ruled out.^14,15

A gamma-glutamyl transpeptidase (GGT) level of twice the normal with an AST/ALT ratio of 2:1 or more is highly suggestive of alcohol abuse as cause of cirrhosis. AST:ALT ratio >1 suggests cirrhosis when used in conjunction with a platelet count of <150,000/mm³ and other variables, such as prothrombin time, leucopenia, and low albumin. AST and ALT are used in combination with other variables in APRI (AST: platelet ratio index) and FIB-4 to predict fibrosis and portal hypertension with varying sensitivity and specificity. An AST to platelet ratio index (APRI) > 1.5 AND fibrosis score of 4 (fib-4) > 3.25 has been used to signify moderate-to-severe fibrosis.29, 30, 31, 32

In a meta-analysis by Denget al³³ the area under the summary receiver operating characteristic curve (AUROC) of APRI(AST/platelet ratio), AAR(AST/ALT ratio) and FIB-4 (age, AST, ALT and platelet) scores for the prediction of varices were 0.6774, 0.7275 and 0.7755 respectively; and those for the prediction of large varices were 0.7278, 0.7448 and 0.7095 respectively.

Alkaline phosphatase and gamma glutamyl transpeptidase

Alkaline phosphatase (ALP) and gamma glutamyl transpeptidase (GGT) reflect cholestasis and are commonly used for the diagnosis in patients with liver disease. Common causes of raised levels of ALP and GGT in patients with cirrhosis include extrahepatic biliary obstruction, PBC, PSC, infiltrative diseases, Ig4-associated cholangitis, and malignancy which could be primary or metastatic. Drugs-induced elevation in ALP and GGT should always be kept in mind and common drugs include anabolic steroids, carbamazepine, phenytoin, and barbiturates and certain herbal medicines. Low level of ALP in patients with cirrhosis could be due to Wilson disease and deficiency of protein, zinc and magnesium. 14, 15, 16

Clinical Relevance in Cirrhosis

Monitoring of ALP and GGT along with aminotransferase is commonly used for initiation and treatment monitoring in patients with PBC and PSC.³⁴ A raised GGT with raised transaminase and a ratio of AST to ALT of 2:1 or more would suggest alcohol-related liver disease. A raised GGT in patients with chronic hepatitis B and cirrhosis should evoke a high suspicion of hepatocellular carcinoma (HCC), and it should be ruled out by doing relevant radiological imaging.³⁵ In a study by Zhang et al.³⁶ who studied 414 patients of HCC, GGT and ALT/AST was positively correlated with the tumour size, tumour volume, tumour volume percentage; tumour, nodes, and metastases (TNM) system stage; 1,3 and 5-year survival rate.

Markers of liver functions

Serum Albumin

Albumin is synthesised in liver and its concentration in blood depends upon its synthetic capacity by liver. Approximately 10 g of albumin is synthesised by liver and secreted daily. Serum albumin levels fall with the progression of liver disease and its value below 3.5 g/dL should raise high suspicion of cirrhosis if total protein levels are normal.¹⁶ Serum concentration of albumin depends upon several other factors such as the nutritional, hormonal factors, sepsis, systemic inflammatory disorders, urinary and gastrointestinal losses. These should be considered when interpreting low albumin levels in patients with chronic liver disease.³⁷

Clinical Significance in Cirrhosis

Low albumin suggests underlying poor synthetic activity of liver and thereby underlying liver disease. Albumin is an integral part of CTP score to assess prognosis in patient with cirrhosis. In a retrospective cohort study (VOCAL-Penn models) for post operative mortality in patients with cirrhosis, preoperative albumin alone is an important component in assessing post operative mortality.³⁸

Patients with decompensated cirrhosis showed improve survival, less liver related complications, eases the management of ascites and reduces hospitalisations with long term albumin infusion hence cost-effective in these patients. Albumin level and its infusion post bleed reduces chances of rebleed in patients with cirrhosis and is used in the management of hepatorenal syndrome-acute kidney injury (HRS-AKI), spontaneous bacterial peritonitis and large volume paracentesis.^39,40

Prothrombin Time (PT)

One of the important functions of liver is coagulation factors synthesis (except factor VIII) and its assessment is one of the key elements of LFT. The PT measures the rate of conversion of prothrombin to thrombin (requiring factors II, V, VII, and X). International normalised ratio (INR) is more often tested now instead of or along with prothrombin time, in order to standardise the reporting of PT results.^15,41 Deficiency of these factors due to poor synthetic activity reflects disease progression in patient with cirrhosis and hence it is important parameter in various prognostic models of cirrhosis like CTP and MELD score.⁴² PT may also get prolonged in vitamin K deficiency as it is required for the gamma carboxylation of these factors, warfarin therapy, consumptive coagulopathy, cholestasis with fat malabsorption.

Clinical Relevance of PT

Vitamin K will reduce a prolonged prothrombin time due to vitamin K deficiency but not due to intrinsic liver disease and this Principle is often used in clinical practice to differentiate these two conditions. PT time/INR is an integral part of various prognostic scores of cirrhosis and ACLF though it does not always reflect bleeding tendency in these patients.^17,18 Patients with compensated liver cirrhosis have a rebalanced coagulation profile. The PT assessment is not a reliable measure of bleeding risk in patients with cirrhosis, because it assesses the activity of procoagulant clotting factors, not anticoagulants such as protein C and antithrombin. The production of these anticoagulants is also reduced in cirrhosis and hence may counterbalance the delicate system. The anticoagulation system (low protein C and protein S) and compensatory higher levels of FVIII may result in a hypercoagulable state despite increased INR. Progression of chronic liver failure is associated with derangement of coagulative balance, and the balance shifts from a procoagulant to anticoagulant phenotype.43, 44, 45Global viscoelastic (VE) test which include thromboelastography (TEG), rotational thromboelastometry (ROTEM), and Sonoclot does more physiological assessment of coagulation in patients with cirrhosis.⁴⁶

Bile Acid Levels

Bile acids are synthesised from cholesterol in hepatocytes, conjugated to glycine or taurine, and secreted into bile. Maintenance of normal serum bile acid concentrations depends on hepatic blood flow, hepatic uptake, secretion of bile acids, and intestinal transit. Serum bile acids are sensitive but non-specific indicators of hepatic dysfunction. It also allows some quantification of functional hepatic reserve.⁴⁷

Yan et al.⁴⁸ evaluated the potential diagnostic value of total bile acid-to-cholesterol ratio (TBA/TC) for liver fibrosis in patients with hepatitis B. The area under the curve (AUC) of TBA/TC (0.87) was almost equivalent to APRI, AUC = 0.84 and fibrosis 4 score (FIB-4, AUC = 0.80), and the optimal cut-off value for TBA/TC to diagnose cirrhosis was 2.70.

Platelet Count

Thrombocytopenia is the most common haematological abnormality found in cirrhosis patients and suggests advanced liver disease. Causes of thrombocytopenia include decreased production, splenic sequestration and increased destruction.¹⁶

Clinical Significance

Platelet count alone neither predicts oesophageal varices either at admission or during follow up nor it predicts variceal bleed during follow up of 4 years.^49,50 Kim et al.⁵¹ found that the index named P2/MS, (PLT)2/[monocyte fraction (%) × segmented neutrophil fraction (%)], was negatively associated with the presence of oesophageal varices in patients with hepatitis B-related cirrhosis, and the patients with P2/MS ≥ 25 had lower risk of bleeding oesophageal varices. Platelet count is an integral part of AST to platelet ratio index (APRI), fibrosis score of 4 (FIB-4), Fibro-index, Fibrometer and Forns which is often used to predict advanced fibrosis, presence and severity of portal hypertension in predicting oesophageal varices. According to Baveno VI guidelines, endoscopy is not necessary in patients with cirrhosis due to viral aetiology if liver stiffness <20 kPa and platelets >150,000/μL.⁵²

In a systemic review, Pang et al.⁵³ evaluated that out of the 4163 patients with hepatocellular carcinoma (HCC) who were recruited, 2746 (66.0%) experienced recurrence and had low platelet count before therapy. (HR = 1.53, 95%CI: 1.29–1.81). Sheng-Nan Lu et al.⁵⁴ also demonstrated that platelet count less than <150 × 10^(3)/mm⁽³⁾ in patients with hepatitis C virus (HCV)-related cirrhosis has higher risk of development of HCC. Platelet count need consideration during procedures like paracentesis, endoscopy, colonoscopy, variceal band ligation, liver biopsy, chemoembolisations, transjugular intrahepatic portosystemic shunts (TIPSs), and biliary procedures. In an analysis of bleeding complications after liver biopsy in patients with HCV with cirrhosis, 11% of biopsies could not be performed in 2 years because of thrombocytopenia, and there was a significantly increased risk for bleeding in patients with a platelet count of less than 60,000/μL.^15,16,55

Quantitative Liver Tests

Quantitative function tests were developed for better evaluating the excretory or detoxification capacity of the liver. These tests lead to improved sensitivity, but poor specificity and often cumbersome methodology has limited their widespread acceptance among clinician. Cirrhosis is state of altered liver structural and functional anatomy due to loss of absolute cell mass and enzyme function. Portal hypertension and associated portosystemic shunts and changes in arterial blood flow also influences clearances of many substances used in quantitative liver tests. Biliary obstruction decreased protein synthesis and binding capacity and secondary renal impairment all affects liver functional capacity to deal with various substances. The degree of functional impairment can be assessed by the use of clearance tests, which for the particular compounds are variably dependent upon: (1) cell mass and its enzymatic capacity; (2) its blood flow; (3) uptake of compounds from the blood into the liver; and (4) its excretion^56,57 (Table 1).

Table 1.

Liver Function Tests in Cirrhosis.

Static tests	Test	Indication and limitation
Parameter assessed
Excretion	Bilirubin Bile acids	Elevated in obstructive/cholestasis liver disease, isolated indirect bilirubin in Gilbert's Limitation: Affected by inflammatory states, biliary obstruction, and haemolysis
Cholestasis	Alkaline phosphatase Gamma Glutamyl transpeptidase 5 nucleosidase	Elevated in obstruction, intrahepatic cholestasis, drug injury, infiltrative liver disease and sepsis Limitation: Not specific to liver Elevated in alcohol abuse and in NAFLD
Liver injury	AST/ALT	Elevated in viral hepatitis, NAFLD, alcohol, metabolic liver disease and in drug injury to liver Limitation: Elevated in muscle and cardiac injury
Synthetic activity	Albumin Prothrombin time/ international normalised ratio (INR) Clotting factors	Determines synthetic function of liver and helps in prognostic score of acute liver failure and chronic liver disease Limitation: Affected in acute-phase reactant, nutrition dependant, nephrotic syndrome Affected in vitamin K deficiency Inherited disease
Metabolic functions Carbohydrates Amino acids Protein	Blood sugar Lactate urea	Helps in evaluating metabolic liver disease (glycogen storage disorder, urea cycle defect) Limitation: Affected in late stage liver disease Elevated in dehydration, ischaemia, systemic inflammatory response syndrome (SIRS) Poor nutrition, muscle mass, overhydration, and urea cycle disorders.
Dynamic tests or qualitative tests
Clearance	Indocyanine green CaffeineBromsulfthalein 99mTc-GSA	Assess metabolic function of liver (clearance and elimination, helps in dynamic assessment of liver) Limitation: Portal flow and collaterals Quantification of caffeine Fasting and availability in different centres Need MRI and special programme
Elimination	Galactose	Fasting and liver regeneration and wide applicability
Metabolic functions	Lidocaine (MGX) Methacetin breath test Aminopyrine	Availability, limited studies, cost
Assessment of portal hypertension
Liver stiffness Direct portal pressure Fluid parameters Platelet count AST/ALT APRI,FIB4,LOK index	Fibroscan, ARFI, MRI elastography HVPG Platelet count/AST/ALT/age/INR, NGAL, cystatin, cytokeratin 18	Affected by bilirubin, raised enzymes, obesity Best test to assess portal hypertension and prognosis in cirrhosis but is invasive and not widely available Non-invasive tests to assess fibrosis and portal hypertension Lack sensitivity and specificity and need more data

AST: aspartate aminotransferase, ALT: Alanine aminotransferase, 99mTc-GSA: technetium-99m galactosyl human serum albumin, INR:International normalised ratio, ARFI: Acoustic Radiation Force Impulse, HVPG: Hepatic venous pressure gradient.

Metabolite Testing Tests

Principle of Breath Tests

Hepatic breath testing was among the first dynamic tests. Substances are labelled with ¹³C or ¹⁴C isotopes of carbon, administered either orally or parenterally and are metabolised primarily in the liver. The labelled CO₂ diffuses into the systemic circulation and is collected through exhalation⁵⁶ (Table 2).

Table 2.

Breath Tests for the Evaluation of Liver Functions.

Test name	Metabolite	Procedure details	Assess	limitations
Aminopyrine breath test (ABT)56	13C-Aminopyrine	75 mg dissolved in warm water and breath assessed every 10 min for 2 h	cytochrome P-450 monooxygenases	Not easily available Genetic variation in Cyt P450 Biliary obstruction
Methacetin breath test (MBT)56	13C -Methacetin	75 mg dissolved in warm water and breath assessed every 10 min for 2 h	cytochrome P450 1A2 enzyme (CYP1A2)	Depend on hepatic blood flow and rate affected by genetic variation of (CYP1A2)
LiMAx test (maximum liver function capacity)63	13C –Methacetin	Intravenous 2 mg/kg 13C –Methacetin and breath sample collected at baseline 10 min prior to injection and 60 min after injection (40 breaths tests recorded)	cytochrome P450 1A2 enzyme (CYP1A2)	Less dependent on oral absorption of drug
Methionine breath test(MeBT)58	13C-methionine	Oral 13C-methionine and breath sample collected	Hepatic mitochondrial function	Affected by alcohol and aspirin
Caffeine breath test57	3-methyl13C-caffeine	breath samples at 30, 60, 90 and 120 min following the caffeine dose (200 mg)	cytochrome P450 system	Affected by blood flow and coffee intake, smoking
Galactose Breath Test58	14C-d-galactose	breath samples at 30, 60, 90 and 120 min following the galactose dose	cytochrome P450 system	Oral absorption affected in cirrhosis, genetic variation in Cyp450
Total Overnight Salivary Caffeine Assessment (TOSCA) 58	caffeine	Salivary samples (overnight fasting at baseline and then after caffeine consumption at 4 h and 12 h)	mediated by the cytochrome P450 1A2 enzyme (CYP1A2)	Calculation of daily caffeine, amount present in different substance, absorption
C14 Erythromycin breath test57	[N-methyl-14C] erythromycin	Breath samples collected every 10 min for 4 h	CYP3A4 a system	Need intravenous radiolabel drug, avoid alcohol, grapefruit juice and smoking

Hepatic breath testing offers a dynamic assessment of liver function and results may depend on the serum kinetics of bicarbonate pools. Carbon dioxide levels fluctuate at base line in few peoples depending on an individual's metabolic state at that particular moment hence resting the patient for 30min to 1 h is advisable. Few breath tests require fasting state, avoidance of smoking, tea, coffee, and heavy breakfast as it may impair the absorption and test results. Genetic polymorphisms of enzymes among individuals may also change the results. These discrepancies must be kept in mind when interpreting breath testing results.⁵⁹

Clearance Tests

Functional impairment of the liver can be assessed using clearance tests (Table 3). Clearance tests for a particular compound depends upon: (1) cell mass and its enzymatic capacity; (2) its blood flow; (3) uptake of compounds from the blood into the liver; and (4) its excretion. Clearance of any test substance is therefore primarily a function of hepatic perfusion and its functional capacity.⁵⁶

Table 3.

Common Clearance Tests.

Test name	Metabolite	Procedure details	Assess	Limitations
Indocyanine green tests(ICG)61	ICG(0.5 mg/kg) intravenous	Venous sample every 5 min interval for 15 min. ICG: PDR (plasma disappearance rate>15%/min or ICG-R15 < 15% at 15 min)	Hepatocyte mass and excretion	Affected by portal vein thrombosis, excessive bilirubin level, portosystemic shunting, SIRS
Galactose elimination capacity56	Galactose (0.5 mg/kg)	Blood sample after 60 min of infusion. < 10 mol/min/kg abnormal	Hepatocellular cell capacity	Prolonged fasting, liver regeneration time
Sulfobromophthalein (BSF)56	BSF (g 5 mg of BSF/kg body weight)	Venous sample at 30 min and 45min. > 5% retention is abnormal	Hepatocyte mass and excretion	Serum albumin concentration and drugs affects this test
Monoethylglycine xylidide (MEGX)62	MEGX (1% lidocaine solution (4 mL)	Venous sample after 15 min <MEGX levels below 10 kg/L	P-450 isoenzyme CYP3A4	Portal shunting and portal vein thrombosis
Antipyrine test60	15 mg/kg antipyrine	1 ml Saliva collected after 4◦ h and 24◦ h	Microsomal oxidases	Drug interactions, cannot be repeated within 1 week

Common clearance tests used are listed in Table 3.

Antipyrine Clearance

Antipyrine clearance is a good test of metabolic liver function. It is metabolised by mixed function oxidases and its elimination follows first order kinetics. It is a weak base and has the advantage of being rapidly absorbed (more than 90%) after oral administration. Its elimination is not dependent upon liver blood flow. Antipyrine clearance showed a significant negative correlation with Child-Pugh scores, total protein, the international normalisation ratio (INR) of prothrombin time and globulin, and a positive correlation with albumin and albumin-to-globulin ratio⁶⁰.

Clinical Significance

Studies had shown that quantitative liver functions tests can be utilised in differentiating the severity of fibrosis, evaluation of risk of decompensation in patients with early cirrhosis for any surgery, graft functioning after liver transplantation in early phase and in patients with acute liver failure to know about regeneration.60, 61, 62, 63 Quantitative methods have not yet been adopted to routine clinical practice in most of the countries because of their lack of specificity and dependence on specialised equipment, time needed to complete the tests, patient cooperativeness and thus are mainly used in research centres.

Assessment of Portal Systemic Shunt and Severity of Liver Disease by Quantitative Liver Function Tests

Dual Cholate Tests

Cholate is an endogenous bile salt and is synthesised in the liver from cholesterol. Its estimated pool is 1 to 3 g and is maintained by hepatic and intestinal transporters. Cholate has a high first-pass hepatic extraction (80% to 90%). It is not metabolised and can estimate liver blood flow and perfusion.

Cholates have been administered safely by oral and intravenous routes. In the dual test, [2,2,4,4-D]cholate (D4-cholate)is administered orally and, [24-13C]cholate (13C-cholate) is administered intravenously, simultaneously. The ratio of 13C (IV) to D4 (oral) cholate clearance estimates portal-systemic shunt. This test quantifies three parameters of liver function – clearance from the systemic circulation and portal circulation, and portal-systemic shunting.⁶⁴

Disease Severity Index

Portal hepatic filtration index, Systemic hepatic filtration index, and SHUNT are related to fibrosis stage and clinical stages of cirrhosis. Disease severity index (DSI) is based on modelling this index in an individual patient. The HepQuant SHUNT test uses the hepatic clearance of cholate from both systemic and portal circulations to assess global liver function and quantify portal-systemic shunting.⁶⁵ The test involves simultaneous administration of 20 mg 13C-cholate IV and 40 mg d4-cholate PO, and subsequent collection of 3 mL blood samples at 5, 20, 45, 60, and 90 min. Clearances are expressed as systemic and portal hepatic filtration rate. Portal-systemic shunting (SHUNT), a disease severity index (DSI), and an estimate of DSI (STAT) are calculated from the clearances.

Fallahzadeh et al.⁶⁶ reported the predictive value of the novel Hepquant SHUNT test among 70 patients with cirrhosis. These patients were followed for a median of 4.2 years. The test uses the estimates of cholate clearance from the systemic and portal circulation to assess liver function, portal blood flow and shunting. Test results have been shown to correlate with HVPG significantly. In this cohort, a DSI ≥24 was associated with a relative risk for clinical outcome of 3.45 (95% CI 1.89–6.29). In this study DSI was superior to and independent of MELD to predict outcome. This is especially relevant as many risk stratification tools may underestimate mortality risk in patients with preserved liver function but severe portal hypertension(Table 4).

Table 4.

Upcoming Tests to Assess Liver Function.

Name	Principle	Assess	Clinical use
HepQuant SHUNT65	Hepatic clearance of cholate (disease severity index, DSI)	Global liver function and portal shunting	Prediction of liver decompensation and complications if DSI>24
Urokinase plasminogen activator receptor (uPAR)68	Upregulated in pathological conditions	Differentiate cirrhosis with non-cirrhosis patients	suPAR levels >9 ng/mL predicts higher short- and long- term mortality
technetium-99m (99mTc)-mebrofenin hepatobiliary scintigraphy (HBS)74	Mebrofenin is taken up into hepatocytes through the action of OATP1B1 and OATP1B3 transporters	Liver volume and quantitative liver function	Correlates with ICG clearance and give functional liver anatomy so helpful in patients undergoing surgery
Technetium-99m–labelled galactosyl-human serum albumin (99mTc-GSA)77	Binds to asialoglycoprotein receptors on viable hepatocytes	Liver volume and quantitative liver function	Helpful for assessing functional liver after surgery and predicts post hepatectomy liver failure
Serum hyaluronic acid67	Marker of fibrosis	Liver injury	Helpful in predicting severity of fibrosis and post operative decompensation if level is > 100 ng/mL on day 1 of operation
Hepatocyte growth factor114	Helps in liver regeneration	Marker of regeneration	Increased level after liver surgery predicts increased regeneration post operatively.

Fluid Biomarkers in Patients with Cirrhosis

Fluid biomarkers have recently been evaluated to assess portal hypertension and prognosticate patients with cirrhosis. These simple, non-invasive, and reproducible markers either alone or in combination with other prognostic models like Child –Turcotte Pugh (CTP) or model for end-stage liver disease (MELD) scores help the physician in managing patients.

Few markers like red blood cell distribution width (RDW), C-reactive protein (CRP), neutrophil-to-lymphocyte ratio (NLR), Aspartate transaminase (AST)/PLT (APRI), serum albumin, serum sodium, Urokinase plasminogen activator receptor (uPAR), Neutrophil gelatinase-associated lipocalin (NGAL), M30, M65, and M65EpiDeath belong to cytokeratin 18 (CK-18), autotoxin, Urinary IL-18, Interleukin-22 (IL-22), myostatin etc. have been studied. However, lack of well randomised studies and availability outside research centres precludes their widespread use by clinicians.⁶⁷

Urokinase plasminogen activator receptor (uPAR): is a membrane glycoprotein and is upregulated in patients with tumour, inflammation, and other pathological conditions. Soluble form of Upar (suPAR) and a promising prognostic biomarker. Zimmermann et al.⁶⁸ detected serum suPAR levels in 159 patients with chronic liver disease (98 cirrhosis) and demonstrated that serum suPAR levels were significantly higher in cirrhosis patients than those in non-cirrhotic patients, and the patients with suPAR levels >9 ng/mL had significantly higher mortality during a median follow-up period of 478 days (RR: 8.5) and is independent of MELD score(Table 4).

Utility in Cirrhosis

Fluid biomarkers are measured objectively and had shown to predict both short- and long-term survival, complication of cirrhosis, and disease progression along with other scores in patients with cirrhosis. Some biomarkers are ‛multifunctional’ and predict mortality as well as development of acute kidney injury (AKI) in cirrhosis for example, CysC and neutrophil gelatinase-associated lipocalin (NGAL). Fluid biomarkers as found in blood, urine, and saliva fluids can be collected easily, repetitively, and noninvasively which is convenient to patient and clinician. However, these results need to be confirmed by more studies.

Fluid Markers to Improve CTP and MELD Score in Cirrhosis

Model for end-stage liver disease (MELD) score and Child –Turcotte Pugh (CTP) score are commonly used to prognosticate patients with cirrhosis. However, these scores have limitation of subjectivity and not involving complication of cirrhosis and portal hypertension. Many scores have been found to be better than MELD alone as MELD-Na score and combination of CRP and procalcitonin (PCT) with MELD score to predict 30-day mortality.^69,70

NGAL is found only in proximal renal tubule epithelial cells, while absent in the distal renal tubule and glomerulus and has shown to predict survival and occurrence of complications in cirrhosis and its combination with MELD has been studied in predicting survival.⁷¹ Similarly MELD-Cystatin Score showed a better performance than the MELD score and CTP score in predicting the probability of survival during a year follow-up.⁷²

Imaging-based Liver Function Tests

Radiological imaging is used primarily for liver resection planning to assess total liver volume and the postoperative liver volume (FLR – future liver remnant). Recently imaging method has been studied as quantitative liver function test along with volumetric assessment. The asialoglycoprotein receptor is expressed exclusively on hepatocytes and substances that bind to these receptors will be taken up by hepatocytes and then excreted in bile. The synthetic ^{99 m}Tc-coupled asialoglycoprotein mebrofenin is the most important substance with highest specificity for hepatocytes.⁷³ With the help of nuclear medicine or radiological methods, the drug concentration is measured via planar images (gamma camera) or cross-sectional imaging (single-photon emission computed tomography [SPECT]/ computed tomography [CT] or magnetic resonance imaging [MRI]). This can be performed at a fixed point in time or dynamically to determine kinetics. When combined with functional testing, image-based testing can help predict the hepatic reserve prior to hepatectomy.⁷⁴

In a study by Olthof et al.⁷⁵they assessed the value of preoperative assessment of liver function using^99mTc-mebrofenin hepatobiliary scintigraphy (HBS) in 116 patients of perihilar cholangiocarcinoma to predict post hepatectomy liver failure (PHLF). The cut-off of 8.5%/min can help to select patients for portal vein embolisation and might help to reduce postoperative liver failure. Galactosyl human serum albumin (GSA) receptor expression strongly correlates with liver function. Following injection with 99mTc-GSA, hepatocytes uptake depends on functional mass in the liver. Utilising single-photon emission computed tomography (SPECT), assessment of volume and liver function can be made at the same time. These tests are of use in conditions when the liver is not homogeneous, such as in unilateral cholestasis or after portal vein embolisation prior to surgery.^76,77In a study by Kato et al.⁷⁸ 100 patients who underwent ^99mTc-GSA scintigraphy and subsequent hepatectomy were retrospectively analysed. The blood clearance ratio (HH15), hepatic uptake ratio (LHL15), and maximal removal rate (R_max) of ^99mTc-GSA (GSA-R_max) were calculated as scintigraphic parameters for the total liver. Technetium-99m–labelled GSA scintigraphy is useful for predicting post hepatectomy liver failure, particularly for applying an accurate resection line on GSA-SPECT images(Table 4). An alternative approach is gadoxetic acid-enhanced magnetic resonance (MR) imaging to assess hepatic function. Following injection of gadoxetic acid, agent is taken up by hepatocytes and excreted into the biliary system.⁷⁶ Nuclear imaging now plays a definite role in assessing liver function along with volume assessment. These tests are expensive and are not available in most of the centres. These tests in combination with clearance tests like indocyanine green (ICG) probably will help in decreasing post operative morbidity and mortality in patients with chronic liver disease.

Assessment of Portal Hypertension as Part of Liver Function Test

Invasive Assessment by HVPG

Assessment of cirrhosis is not completed without assessing the presence and severity of portal hypertension. In clinical practice, presence of ascites, varices and thrombocytopenia suggests significant portal hypertension. However, response to therapy like beta blockers, antiviral treatment and antifibrotic treatment during follow up need objective assessment. Presently the best way to assess portal venous pressure is hepatic venous pressure gradient (HVPG) that is the gradient between the portal vein and the hepatic vein measurement. HVPG score >6 mmHg indicates portal hypertension and HVPG >10 mmHg represents clinically significant portal hypertension (CSPH). Patients with no significant portal hypertension that is HVPG ≤10 mmHg reveal a 90% chance of remaining free of varices and decompensation however, this risk increases to 22% at 24 months. Patients with cirrhosis and an HVPG >16 mmHg or HVPG >20 mmHg are an important predictor of poor outcome after variceal bleed. HVPG measurement is increasingly used these days in many centres as HVPG is a robust surrogate marker for diagnosis (to differentiate cirrhotic versus non-cirrhotic portal hypertension), risk stratification for variceal bleed (early transjugular intrahepatic portosystemic shunt [TIPS] versus standard treatment), identification of patients with hepatocellular carcinoma who are candidates for liver resection (HVPG<10 mmHg), monitoring of the efficacy of medical treatment by non-selective beta-blockers, and assessment of progression of portal hypertension79, 80, 81, 82 (Table 5).

Table 5.

Clinical Correlation of HVPG with PHT and Cirrhosis Complications.

HVPG (Hepatic venous pressure gradient)	Clinical end points
<5 mmHg	normal
5–10 mmHg >6 mmHg	Mild PHT Chronic hepatitis progression and post liver transplantation means recurrence of disease
>10 mmHg	Clinically significant portal hypertension (CSPH)
>10 mmHg	High risk of development of varices, ascites, HCC development, decompensation after liver surgery
>12 mmHg	Risk of variceal bleed
>16 mm Hg	High mortality after any superadded liver insult
>20 mm Hg	Failure to control bleed, think of early TIPSs
>22 mm Hg	High mortality after bleed, in alcoholic hepatitis poor predictor

Patients who had a reduction in HVPG of ≥20% or to ≤12 mmHg in response to drug therapy are defined as responders to beta-blockers. Responders have a markedly improved survival and reduce risk of rebleeding, ascites and spontaneous bacterial peritonitis (SBP).^81,82

Non-invasive Assessment of Clinically Significant Portal Hypertension in Cirrhosis

Rockey at el⁸³ in a retrospective cohort study had shown that that platelet count, APRI, AST/ALT ratio and Lok index were helpful in predicting the presence of varices prior to endoscopy and in predicting the likelihood of variceal bleeding. However, it has got poor sensitivity and specificity and normally not practised by clinicians. Liver stiffness as assessed by Fibroscan (transient elastography) has been correlated with HVPG. In a study of 150 patients undergoing liver biopsy, HVPG and transient elastography, liver stiffness had an AUROC of 0.95 in detecting significant portal hypertension (HVPG ≥10 mmHg) at cutoff of 21 kPa. Splenomegaly is a part of portal hypertension and its stiffness as measured by transient elastography is less affected by changes in aminotransferses level, congestive heart failure (CHF) and bilirubin level. Hence its stiffness along with liver stiffness has also shown to be correlated with clinically significant portal hypertension (CSPH) in many studies.^84,85 Baveno VI consensus guidelines stress on liver stiffness and platelet count to select cirrhotic patients due to hepatitis for varices screening. Spleen stiffness has still not been validated by Baveno.⁵²For patients with liver stiffness <20 kPa and normal platelet count(>150,000), the risk of bleeding and large varices is minimal, and the non-invasive tests may be repeated annually.

Fluid Biomarkers as Non-invasive Assessment of Portal Hypertension

Various blood parameters like osteopontin level had shown to distinguish CSPH at 75% sensitivity and 63% specificity, using a cut-off value of 80 ng/mL.⁸⁶ Von Willebrand factor was also proposed to be a clinically significant non-invasive predictor of clinically significant portal hypertension (CSPH).⁸⁷ Using a cut-off value of ≥241%, the area under receiver operating curve (AUROC) for detection of CSPH in cirrhosis was 0.85, and mortality prediction is comparable to the model for end-stage liver disease (MELD) score. Recently the VITRO score (Von Willebrand Factor Antigen/Thrombocyte Ratio), had shown to be a possible marker for detecting CSPH and better than von Willebrand factor, and the APRI and ELF score, but lower than that of transient elastography. Fibroscan (transient elastography) is thus the most common and easy method of assessment of portal hypertension in clinical practice⁸⁸ (Table 6).

Table 6.

Serum Tests Used to Assess Fibrosis and Complications of Cirrhosis in Various Studies.

Indirect serum tests	Components	Predicting fibrosis	Predicting portal hypertension and its complication
Platelet count	Platelet count	+++	++
AST/platelet ratio index	AST and platelet count	+++	–
AST/ALT ratio	AST and ALT	+++	–
Lok	Platelet count, AST, ALT, and INR	+++	+
FIB4(Fibrosis-4)	Platelet count, AST, ALT, and age	+++	+
Forns	Platelet count, GGT, age and cholesterol	+++	–
Fibroindex	Platelet count, AST, and gamma globulin	+++	+
Direct fibrosis markers
Fibrometer	a2-macroglobulin, ALT, AST, GGT, urea, prothrombin index, platelet count	+++	–
Fibrotest	a2-macroglobulin, GGT, total bilirubin, haptoglobin, apolipoprotein A1	+++	+
Hepascore	a2-macroglobulin, hyaluronic acid, GGT, total bilirubin, age, sex	+++	–
Hyaluronic acid	Hyaluronic acid	+++	–
Enhanced liver fibrosis	Hyaluronic acid, tissue inhibitor of metalloproteinase-1, amino-terminal propeptide of type III collagen	+++	–
FibroSpect	a2-macroglobulin, hyaluronic acid, tissue inhibitor of metalloproteinase-1	+++	–

AST: Aspartate Aminotransferase, ALT: alanine aminotransferase, GGT: Gamma-glutamyl transferase.

Liver Tests of prognostic Value

Liver tests in patients with cirrhosis should be able to enlighten the clinician regarding the overall prognosis of the patient, risk stratification in case of any surgery, need of liver transplantation, and regeneration of liver after any insult and response to treatment in patients with viral hepatitis, autoimmune hepatitis, cholestasis liver disease and storage disorders of liver. No hepatologist needs a new test to conclude that an asymptomatic Child's class A cirrhotic patient does not need to undergo transplantation at present, or to conclude that transplantation is best recommended for a Child's class C cirrhotic to prolong his survival unless there is no contraindication. Hence clinician used combination method for assessing overall prognosis which include CTP, MELD score along with HVPG and qualitative liver tests wherever these are available.

Scores in Cirrhosis

No single liver test has shown to have prognostic value due to diverse functions of liver and no single tests can evaluate all functions of liver. Prognosis in cirrhosis depends upon stage of cirrhosis, degree of portal hypertension, coexisting comorbidities, renal and circulatory dysfunctions, hepatocellular carcinoma and overall performance status in these patients. Hence, scoring systems like Child –Turcotte Pugh (CTP), model for end-stage liver disease (MELD) and its modifications like MELD-sodium came into picture. D'Amico et al.⁷ systemically reviewed 118 studies in patients with cirrhosis (23,797 patients) and found that most consistent and ‘robust’ predictor of death in cirrhosis is the Child–Pugh score and/or its components (albumin, bilirubin, ascites, encephalopathy, and prothrombin time. In patients with decompensated cirrhosis associated factors like circulatory dysfunction, serum creatinine, bleeding and associated HCC also are important predictors of mortality. CTP score does not take into consideration the renal impairment which has prognostic value in patients with decompensated cirrhosis. It is in this group it is not surprising that the MELD score (which incorporates creatinine in addition to markers of liver dysfunction) has become a valuable method to know prognosis. On the contrary, it is predictable that the MELD score would not be useful to predict survival in patients with compensated cirrhosis.⁴²

MELD and its modification MELD-sodium score has replaced the CTP score for stratifying patients on liver transplant waiting lists, it is still debated as to whether it is better than the CTP in settings of chronic liver disease. In a recent review of all data comparing the accuracy of MELD score to the CTP score, MELD score performance is not rated to be better than the CTP score in non-transplant settings.^89,90

Scores for Listing Patients for Liver Transplantation

The Child –Turcotte Pugh (CTP) and model for end-stage liver disease (MELD) scores are the two most common scores used to assess severity of cirrhosis and listing patients for liver transplantation (LT). There are several disease-specific scoring systems but are uncommonly used due to unavailability of large trials. MELD score is the standard score used across all liver diseases. Evaluation for LT should be considered once a patient with cirrhosis has experienced an index complication such as ascites, hepatic encephalopathy, or variceal haemorrhage or hepatocellular dysfunction results in a MELD Score ≥15.Many modification were done to improve predictability of allocation of organs in which MELD-sodium has been in use in many countries.^69,70 It has been shown that increase in mortality by 5% for each millimole decrease in serum sodium between 125 and 140 mmol/L.⁹¹ Other modification includes addition of ascites to MELD, MELD without international normalised ratio (INR), MELD to sodium ratio etc. Another score which is used is CTP score and a score of >7 indicates decompensation and used as a criterion for listing the patient for LT.^92,93

Patients with alcoholic hepatitis and Maddrey discriminant function [4.6 x (prothrombin time prolongation) + (serum bilirubin in mg/dL) > 32] represent severe disease and are an indication for corticosteroid therapy if patients satisfy other criteria. Other scores used to define severe alcoholic hepatitis and predict mortality include ABIC (age, bilirubin, INR, and creatinine score) and Glasgow alcoholic hepatitis. If patient has not responded to steroid according to Lille score (dynamic model assessed after 1 week of corticosteroid therapy) depicts poor survival and can be listed for LT if other criteria fit as assessed by a multidisciplinary team.⁹⁴

The Mayo score for primary biliary cholangitis formerly known as primary biliary cirrhosis [PBC] (0.039∗age (years) + 0.871∗log e bilirubin (mg/dL)—2.53∗log e albumin (g/dL) +2.9∗log e prothrombin time (s) +0.859∗oedema) has been shown to predict survival without transplantation. Mayo score of 7.8 predicts survival of 63% and 39% at 1 and 2 years, respectively so the ideal window for liver transplantation in primary biliary cirrhosis (PBC) is recommended between scores of 6–7.8.Patients with PBC and decompensation and PBC with severe itching affecting daily life even without decompensation is an indication for LT.⁹⁵ Mayo risk score for primary sclerosing cholangitis [PSC] [(0.0295∗age (years) + 0.5373∗log e bilirubin (mg/dL)—0.8389∗log e albumin (g/dL) + 0.5380∗log e alanine amino-aspartate (IU/L) + 1.2426∗(variceal bleeding)] allows the identification of three groups at low (score <0), intermediate (0 ≤ score >2) or high (score ≥2) risk. Five-year survival is <40% in patients at high risk and should be considered for LT.⁹⁶ LT is an effective therapy for decompensated liver disease due to PSC or in patients with PSC with recurrent cholangitis and sepsis.

Quantitative Liver Tests

Quantitative liver tests have not been shown to be superior over CTP score or MELD score in various earlier heterogeneous studies as enrolled patient had low fibrosis or compensated cirrhosis. Some recent studies had shown that these tests may predict decompensation, survival better than CTP and MELD score and are important complements to prevailing score. Indocyanine green (ICG) -r15, monoethylglycine xylidide (MEGX) and ¹³C-based liver function test (LiMAx) have shown to predict the occurrence of liver decompensation and survival in compensated patients with cirrhosis better than Child –Turcotte Pugh (CTP) score in cirrhosis.60, 61, 62, 63, 64, 65 MEGX has been shown to complement the CTP score in patients undergoing evaluation for liver transplantation. In a study of 165 patients with cirrhosis, the methacetin breath test (MBT) was found to be better than MELD in predicting liver -related death especially in patients with low MELD score when followed up for 1 year. MBT also predicted the risk of liver-related complications, particularly the risk of development/exacerbation of ascites, which was not achieved by MELD.⁹⁷Limitation of quantitative liver tests are poor availability, need patient cooperation, expertise, time consuming and lack of long term follow up studies. However, they may serve a very good complement test with CTP and MELD score to better prognosticate survival in patients with cirrhosis.

Liver Stiffness Measurement

Foucher et al.⁹⁸ concluded that liver stiffness was significantly correlated with clinical, biological, and morphological parameters of liver disease. With an negative predictive value (NPV) > 90%, the cut off values for the presence of large oesophageal varices, Child-Pugh B or C cirrhosis, history of ascites, hepatocellular carcinoma (HCC), and variceal bleeding were 27.5, 37.5, 49.1, 53.7, and 62.7 kPa, respectively. Kim et al.⁹⁹ in a prospective, longitudinal study assess the abilities of TE and LS-spleen diameter to platelet ratio score(LSPS) to predict the first event of hepatic decompensation in patients with cirrhosis. LS values (13–18 and ≥18 kPa) had significantly higher risks of developing hepatic decompensation compared to those with LS values (<13 kPa). Similarly, the risk of hepatic decompensation was increased in patients with higher LSPS values (1.1–2.5 and ≥2.5) compared to lower values (<1.1). Masuzaki et al.¹⁰⁰ found that in addition to age, male gender, and clinical cirrhosis, the LS value by transient elastography was an independent risk factor for HCC development. Compared to patients with LS values ≤ 10 kPa, patients with higher LS values had a significantly increased risk of developing HCC (liver stiffness [LS] values, 10.1–15 kPa, HR = 16.7; and LS values > 25 kPa, HR = 45.5).

A metanalysis included 17 studies, reporting on 7058 patients with chronic liver diseases (CLDs). Baseline liver stiffness measurement (LSM) was associated significantly with risk of hepatic decompensation (6 studies; RR, 1.07; 95% CI, 1.03–1.11), HCC (9 studies; RR, 1.11; 95% CI, 1.05–1.18), death (5 studies; RR, 1.22; 95% CI, 1.05–1.43), or a composite of these outcomes (7 studies; RR, 1.32; 95% CI, 1.16–1.51). Liver stiffness is a useful, easy objective parameter to assess future risk of liver-related complications which include variceal bleed, HCC and overall mortality.¹⁰¹

HVPG

Hepatic venous pressure gradient (HVPG) measurements provide one of best independent prognostic information on survival in patients with cirrhosis and the risk of decompensation in these patients. Clinically significant pulmonary hypertension [PHT] (clinically significant portal hypertension [CSPH], HVPG ≥10 mmHg) is normally seen in patients with oesophageal varices, its bleeding, and the development of decompensation. In decompensated patients with cirrhosis, HVPG provides predictive information about the risk of mortality in the future. If available HVPG provides best prognostic variable for survival and portal hypertension related complications and should be done in patients with cirrhosis.102, 103, 104

Preoperative Assessment in Cirrhosis Patient

Assessment of liver function is a critically important for patients undergoing any hepatic surgery especially in patients with underlying liver disease. Overall assessment in terms of remnant liver volume, its function, presence of significant portal hypertension, risk of postoperative decompensation and associated comorbidities all are important before taking up a patient with cirrhosis to surgery. Risk of decompensation also depends upon type of surgery whether related to liver or non-liver surgery like hernia repair etc. No single liver test can predict postoperative morbidity and mortality so a combination of prognostic models has been developed and validated.

Score System

In general, patients with Child-Pugh class A cirrhosis may undergo surgery in the absence of thrombocytopenia or clinically significant portal hypertension (CSPH); only selected patients with Child-Pugh class B cirrhosis are elective surgical candidates while patients with Child-Pugh class C cirrhosis are not considered candidates for elective surgery. Northup and colleagues¹⁰⁵ found a 1% increase in mortality for each MELD point until 20 and a 2% increase in mortality with each MELD point after 20. A MELD score of 5–15 had mortality of 5%–11%, as compared with mortality of 26%, 50%, and 67% when the MELD reaches the value of 25, 35, and 45, respectively. D'Amico et al.⁷ in a systemic review of 118 studies, suggested that Child –Turcotte Pugh (CTP) classification may be better in assessing the risk in compensated cirrhosis, whereas MELD is better predictor in decompensated cirrhotic state.

Platelet Count, Transient Elastography and HVPG

Platelet count of <100,000/mL suggests presence of significant portal hypertension and HVPG >10 mmHg signifies significant morbidity and mortality after any liver surgery and these patients should avoid liver-related surgery. Liver stiffness measurement for hepatic surgical resection has been proposed but optimal cutoff values have not been validated in large studies. In one series, a liver stiffness measurement below 22 kPa was the cutoff level for selecting patients for surgery.^106,107

Role of Quantitative and Imaging Test for Preoperative Decision

Liver quantitative and imaging test offer attractive means to supplement the CTP assessment. Of these, the most widely used is indocyanine green (ICG) retention. Clearance is impaired when 15% or more of the dye remains within the plasma after 15 min of injection. Thus, patients with CTP A and ICG 15 of greater than 14% are the ‘‘bad risk’’ patients compared to patients with <14%. It also correlates strongly with the rate of disappearance of tagged asialoglycoproteins in imaging studies.¹⁰⁸If ICGR15 is between 0 and 10% then patient has a good liver function and bisectorectomy can be done, if between 10 and 19% then left sided hepatectomy or right side sectionectomy can be done.¹⁰⁹

In combination with functional testing, image-based testing can help to predict hepatic reserve prior to hepatectomy. Galactosyl human serum albumin (GSA) receptor expression strongly correlates with liver function and helps in correctly identifying patients of cirrhosis undergoing any surgery.¹¹⁰ Hence combination of clinical assessment, CTP, or MELD score in combination with transient elastography, quantitative liver tests and functional imaging tests help the surgeon in estimating the risk of surgery in patients with cirrhosis.

Liver Function Tests to Estimate Liver Regeneration

Reliable assessment of hepatic function during liver regeneration is crucial after acute liver failure, drug-induced liver injury, viral hepatitis, liver surgery and post liver transplantation. Various functional imaging tests like (99m)Tc-galactosyl human serum albumin ((99m)Tc-GSA) scintigraphy, (99m)Tc-mebrofenin hepatobiliary scintigraphy (HBS) and quantitative liver tests like indocyanine green (ICG) clearance test, and the galactose elimination capacity (GEC) could be of use. There are not many studies evaluating these modalities in human however in a rat model it has been shown that functional regeneration is impaired, compared with volumetric regeneration, in the early phase of liver regeneration. In later stages of liver regeneration, (99m)Tc-GSA uptake underestimates hepatic regeneration compared to liver volume and (99m)Tc-mebrofenin uptake. GEC is of less value and is influenced by factors not related to liver function. (99m)Tc-mebrofenin HBS provides both visual and quantitative information regarding both uptake and excretory liver function. Hence not much data is available for estimating liver regeneration in humans.¹¹¹ Hepatocyte growth factor (HGF) is a hepatocyte mitogen that binds to HGFR/c-MET expressed in parenchymal and non-parenchymal liver cells. In humans, HGF estimation has shown some promising results in the context of liver regeneration in recipient and in donor. However its level in patients with cirrhosis after hepatic and non-hepatic injury has not been studied well.^112,113 After hepatectomy, HGF level rises rapidly following IL-6 and TNF stimulation and peaks on the first postoperative day (POD1), reaching 0.5–2 ng/mL and stays elevated for 7–14 days after the resection¹¹⁴. It is unclear if HGF correlates with remnant liver volume. Matsumoto et al.¹¹³ extensively analyzed correlations between HGF levels and liver volume, and reported an inverse correlation between remnant liver volume per body weight on POD0 to serum HGF on POD1¹¹³. Various factors which probably have a role in liver regeneration can be assessed like epidermal growth factors, vascular endothelial growth factor, insulin like growth factor, fibroblast growth factor, platelet- derived growth factors but these has not been properly validated in human studies.¹¹⁵ In a small cohort of patients, Starlinger et al.¹¹⁶ identified the miRNA signature, which consisted of circulating miRNAs 151a5p, 192-5p, and 122-5p, as a potential prognostic tool for predicting postoperative liver dysfunction, morbidity, and even mortality.

Hyaluronic acid (HA) is a glycosaminoglycan present in the extracellular matrix and has been established as a serum biomarker for severe hepatic fibrosis in chronic liver disease. In the liver, hepatic stellate cells synthesise HA which subsequently is degraded by sinusoidal endothelial cells. High levels of serum HA however are indicative of liver damage and fibrosis. Preoperatively increased serum HA may be indicative of an undiagnosed liver damage prior to hepatectomy which explains higher post-hepatectomy liver failure rates (PHLFs) in this group. Postoperatively high serum HA was shown to be predictive for post-hepatectomy liver failure (PHLF) rate with high sensitivity (100%) and specificity (93.75%). An important aspect is that serum HA was measured on the first postoperative day in most studies. These findings suggest that HA is a clinically relevant parameter to be used as a diagnostic criterion for PHLF as well as a preoperative assessment of underlying liver disease.¹¹⁷

Liver performs variety of functions, which include metabolic, immune regulation, synthetic and excretory functions. No single test can assess these functions and hence we need a battery of tests for its assessment. Cirrhosis is a condition which affects systemic and portal circulation and hence affects circulatory and renal functions. Liver functions tests hence include a battery of tests which include static blood tests, quantitative blood tests like metabolic and clearance tests, imaging modality like functional MRI, transient elastography and various scores like Child –Turcotte Pugh (CTP) and MELD. Traditional tests like serum bilirubin, AST, ALT, PT, and albumin only partly assess liver injury, secretory or synthetic activity and hence have limitation of not assessing portal hypertension and complication of cirrhosis. HVPG is still considered a gold standard for assessment of portal hypertension but is invasive and is not available in majority of centres. non-invasive assessment of portal hypertension as assessed by transient elastography and various scores like APRI, FIB-4 and Forns have varying sensitivity and specificity.ICG-15 and Methacetin breath test are tests which are increasing being used in various transplant centres in supplementation with CTP and MELD score for various liver related surgery in predicting post operative complications. We do not have any reliable tests for assessing liver regeneration after liver insult. Need of a test or a battery of tests that can reliably assess all functions of liver which is easy to use, reproducible, easily available is still required.

Credit authorSHIP CONTRIBUTION statement

Praveen Sharma is the sole author of this review article, and he is responsible for all statement and is fully responsible for this review article.

Conflicts of interest

The author has none to declare.

Funding

No grant received.