ABCB4 gene sequence variation in women with intrahepatic cholestasis of pregnancy.

Intrahepatic cholestasis of pregnancy (ICP), also known as obstetric cholestasis, is a liver disease of pregnancy that complicates 0.7% of pregnancies in the UK. 2 ICP causes maternal pruritus and hepatic impairment and can cause fetal death, spontaneous prematurity, and sudden intrauterine death. A diagnosis of ICP is made by the demonstration of abnormal liver function test results, and in particular the serum bile acids are raised. This is thought to be a consequence of abnormal bile transport across the hepatocyte canalicular membrane. Clinical features are heterogeneous and the aetiology is likely to be complex. Insights to the genetic aetiology of ICP have come from studies of the childhood liver disease progressive familial intrahepatic cholestasis (PFIC), a condition which is divided into three subtypes. Children with PFIC1 and 2 have low concentrations of biliary bile acids and low to normal gammaglutamyl transpeptidase (GGT) in the serum. PFIC3 patients have high serum levels of GGT and bile which lacks phospholipid but has a normal biliary bile acid concentration. Homozygous mutations of the ABCB4 (also called MDR3 or mdr2 in the mouse) gene have been described in pedigrees with PFIC3. The ABCB4 protein is a member of the ATP binding cassette (ABC) family of membrane transporters. One of the normal functions of ABCB4 is to transport phosphatidylcholine across the hepatocyte canalicular membrane. The fact that expression is not only found in hepatocytes but also in B lymphocytes, heart, and muscle suggests that it may also transport other substrates. However, homozygous knockouts of the homologous (>90% identity at the amino acid level) murine mdr2 only had hepatic effects. Several heterozygous mothers of children with PFIC3 have symptoms consistent with ICP. In a large consanguineous pedigree with coexisting PFIC3 and ICP, three of the six mothers with ICP had pregnancies complicated by unexplained intrauterine death. Four of the six women were investigated and shown to be heterozygotes for the ABCB4 mutation for which the proband was a homozygote. In a recent study of 31 patients with PFIC3, ABCB4 mutations were found in 17 cases, and three heterozygous mothers were either diagnosed or suspected of having ICP. More frequently, ICP occurs in women with no family history of PFIC. Based on the observation that children with PFIC3 and ABCB4 gene mutations have a raised serum GGT, we previously investigated a subgroup of eight women with ICP and a raised GGT and found a missense mutation in the ABCB4 gene in one case. This mutation resulted in lack of functional protein at the cell surface. Both homozygous and heterozygous ABCB4 mutations have also been described in three of six cases of women with cholesterol gallstones in association with ICP, and with no family history of PFIC3. In an attempt to elucidate the contribution of sequence variants in ABCB4 in the pathogenesis of ICP, we sequenced the complete coding region from 14 women with ICP and raised GGT. The frequencies of variants found in these cases and in eight cases that were previously sequenced were compared in 184 ICP subjects and controls. In the majority of ICP cases in this cohort, the GGT level had not been measured. We also examined the cohort of 184 ICP patients for the presence or absence of known variations in the ABCB4 gene and analysed three SNPs within the coding region for disease association. Previous studies have investigated the role of ABCB4 mutations in specific small subgroups of ICP cases, that is, a small number of cases with a raised GGT, three women with coexistent cholesterol gallstones with atypical clinical features, or the minority of cases with children affected by PFIC3. This is the first study of ABCB4 variants in a larger group of ICP cases with and without a known raised GGT.