Autoimmune Liver Serology

Autoimmune liver serology is a group of autoantibodies used to diagnose autoimmune hepatitis (AIH), primary biliary cholangitis (PBC) and primary sclerosing cholangitis (PSC), and overlap syndromes. Testing is done by indirect immunofluorescence on rodent tissue (kidney, liver, stomach) or HEp-2 cells, and by immunoassay or line blot for specific targets.

The panel includes antinuclear antibody (ANA), anti-smooth muscle antibody (SMA, with F-actin pattern), anti-liver kidney microsomal antibody type 1 (anti-LKM1), anti-soluble liver antigen/liver pancreas (anti-SLA/LP), antimitochondrial antibody (AMA, with M2 subtype), PBC-specific antinuclear antibodies (anti-gp210, anti-sp100, anti-centromere), and serum IgG.

Autoimmune hepatitis affects about 1 to 2 in 100 000 people each year, mainly women (female to male ratio 3.5 to 1), with peaks in adolescence and the fifth and sixth decades. Primary biliary cholangitis affects 1 in 1000 women over 40 years. Untreated autoimmune hepatitis leads to cirrhosis within years, and a response to immunosuppression is excellent when diagnosed early, so a positive autoantibody in unexplained hepatitis is a prompt for biopsy.

Indications. Unexplained raised transaminases or acute hepatitis after viral causes, drugs and alcohol are excluded. Chronic cholestasis with raised alkaline phosphatase and gamma glutamyl transferase. Pruritus and fatigue in a middle aged woman. Hypergammaglobulinaemia with liver disease. Children with unexplained liver disease. Suspected overlap or drug induced autoimmune like hepatitis. Acute liver failure of unknown cause. Assessment before and during immunosuppression.

Specimen. Serum 1 to 2 mL, no fasting. Stable for days at 2 to 8 °C. Collected before steroid treatment where possible.

Autoimmune hepatitis markers.

• ANA. Positive in 50 to 70% of type 1 AIH (titre 1 in 80 or more in adults, 1 in 20 in children), with a homogeneous or speckled pattern. Not specific, and also positive in 20% of healthy older women.
• Anti-smooth muscle antibody (SMA). Positive in 50 to 70% of type 1 AIH (titre 1 in 40 or more in adults, 1 in 20 in children). The F-actin (anti-microfilament) pattern is more specific.
• Anti-LKM1. Defines type 2 AIH (children and young women, 10 to 15% of AIH), directed against CYP2D6. Also positive in some hepatitis C patients.
• Anti-SLA/LP. Highly specific (over 95%) for AIH, found in 10 to 30% of cases, both seronegative and seropositive, and linked to severe disease and relapse.
• Anti-LC1 (liver cytosol type 1). Type 2 AIH and a marker of activity in children.
• Serum IgG. Raised (over 16 g/L) in 85% of AIH, and its fall tracks response to treatment.

Cholestatic disease markers.

• AMA (M2). Positive in 90 to 95% of primary biliary cholangitis, specific (over 95%) at titre 1 in 40 or more. Anti-M2 immunoassay is more specific than immunofluorescence.
• PBC-specific ANA. Anti-gp210 (perinuclear rim) and anti-sp100 (multiple nuclear dots) in 20 to 50% of AMA-negative PBC and predict severe disease. Anti-centromere indicates portal hypertension risk.
• PSC. No specific antibody. Atypical p-ANCA (perinuclear ANCA directed against nuclear antigens) is positive in 50 to 80%, and ANA and SMA in 20 to 50%. Diagnosis is by MRCP. Serum IgG4 is measured to exclude IgG4-related sclerosing cholangitis.

Scoring and pattern interpretation.

Critical values and findings needing immediate action.

• ALT over 10 times the upper limit with jaundice, raised INR and positive autoantibodies: acute severe AIH or acute liver failure, urgent hepatology referral and consideration of high dose steroids and transplant listing.
• Prolonged INR over 1.5 with encephalopathy in any patient with liver disease: acute liver failure.
• IgG markedly raised with cirrhosis, ascites or variceal bleeding: decompensated autoimmune liver disease.
• New jaundice and itching in a middle aged woman with AMA: PBC, start ursodeoxycholic acid.

Interferences and pitfalls.

• Autoantibodies are supportive but not diagnostic, and the diagnosis rests on biopsy (interface hepatitis, plasma cells, rosettes), raised IgG and exclusion of viral hepatitis, drug injury, Wilson disease, haemochromatosis, alpha-1 antitrypsin deficiency and alcohol.
• ANA and SMA at low titres are common in healthy people, in non-alcoholic fatty liver disease (up to 30%), viral hepatitis and drug induced liver injury, so low titres in these settings do not indicate AIH.
• Seronegative AIH occurs in 10% of cases, and anti-SLA/LP and biopsy are valuable.
• Drug induced autoimmune like hepatitis (nitrofurantoin, minocycline, methyldopa, hydralazine, infliximab, statins) mimics AIH with positive antibodies, and the response to drug withdrawal confirms it.
• AMA can be positive in 1 to 2% of healthy people, and in AMA-positive patients with normal alkaline phosphatase, annual follow up is needed because 15% develop PBC.
• Antibody titres do not track disease activity, and transaminases and IgG are used to monitor treatment.
• Antibody patterns on immunofluorescence need an experienced reader.
• Immunosuppression, including steroids, can lower or clear titres.

Special groups.

• Children: titres of 1 in 20 are significant, and type 2 AIH is more frequent. Sclerosing cholangitis overlaps with AIH in 50% (autoimmune sclerosing cholangitis).
• Pregnancy: AIH flares in the postpartum period, and treatment continues in pregnancy (azathioprine, steroids), while mycophenolate is stopped.
• Older adults: AIH in over 60 years is not rare and is often diagnosed late at cirrhosis stage.
• Regions: viral hepatitis is the main differential in high prevalence countries, and viral markers are always requested alongside.

Follow up tests. Liver function tests, INR, immunoglobulins with IgG subclasses (IgG4), viral hepatitis A to E serology, caeruloplasmin and urinary copper, ferritin and transferrin saturation, alpha-1 antitrypsin, ultrasound with Doppler, MRCP, transient elastography, liver biopsy, thiopurine methyltransferase before azathioprine, bone density scan, and screening for associated autoimmune disease (thyroid, coeliac).