Prenatal diagnosis of bilirubin-UDP-glucuronosyltransferase deficiency in rats by genomic DNA analysis

• Published in: Hepatology (Baltimore, Md.) Vol. 16; no. 3; p. 756
• Main Authors: Huang, T J, Chowdhury, J R, Lahiri, P, Yerneni, P C, Bommineni, V R, Arias, I M, Chowdhury, N R
• Format: Journal Article
• Language: English
• Published: United States 01-09-1992
• Subjects: Animals; Base Sequence; DNA - analysis; Genetic Carrier Screening; Genotype; Glucosyltransferases - deficiency; Glucosyltransferases - genetics; Heterozygote; Homozygote; Molecular Sequence Data; Phenotype; Polymerase Chain Reaction; Prenatal Diagnosis - methods; Rats; Rats, Gunn; Rats, Inbred Strains; RNA, Messenger - analysis
• ISSN: 0270-9139
• DOI: 10.1002/hep.1840160323

Hepatic bilirubin excretion requires UDP-glucuronosyltransferase-mediated glucuronidation. Patients with type I Crigler-Najjar syndrome and mutant rats (Gunn strain) inherit deficiency of UDP-glucuronyltransferase activity toward bilirubin as an autosomal recessive trait and, as a result, exhibit marked nonhemolytic unconjugated hyperbilirubinemia throughout postnatal life. Heterozygous carriers of the trait have normal serum bilirubin levels. Because of placental excretion of unconjugated bilirubin, type 1 Crigler-Najjar syndrome patients and Gunn rats are not jaundiced in utero, making prenatal diagnosis difficult. Here we report a diagnostic method in Gunn rats based on genomic DNA analysis for prenatal recognition of deficiency of UDP-glucuronyltransferase activity toward bilirubin in Gunn rats and identification of heterozygous carriers. We and others have shown that two distinct messenger RNA species (UDP-glucuronyltransferase activity toward bilirubin and the 3-methylcholanthrene-inducible phenol-UDP-glucuronyltransferase messenger RNA) in Gunn rat liver contain identical deletions of a single guanosine residue in their common 3' regions. Loss of the restriction site for the endonuclease BstNI, which results from this deletion, was used as the basis for a diagnostic test. Female heterozygous Gunn rats were mated with male homozygous Gunn rats. Genomic DNA was extracted from the chorionic aspect of placenta of 17-day fetuses or from leukocytes from normal rats, obligate heterozygotes and homozygous Gunn rats. The DNA was sequentially digested with the restriction enzymes EcoRI and BstNI and subjected to Southern-blot analysis with a double-stranded DNA probe for the common region of UDP-glucuronyltransferase activity toward bilirubin and the 3-methylcholanthrene-inducible UDP-glucuronyltransferase messenger RNAs.