The surgery of biliary atresia

One hundred thirty-one consecutive infants with biliary atresia were operated on during the 15-year period between 1973 and 1988. Six patients did not have biliary reconstruction because of advanced cirrhosis or transplant preference. The other 125 infants had excision of all nonpatent extrahepatic bile ducts; biliary drainage was provided by a gallbladder-common bile duct conduit in 14 patients and by a Roux-en-Y portoenterostomy in 111 infants (including the seven patients with correctable biliary atresia). The bilioenteric conduit was temporarily exteriorized and, for the past 2 years, a conduit intussusception valve was incorporated. Immediate postsurgical bile drainage was achieved in 103 infants (82%). Reoperation during the first 6 postoperative weeks restored bile flow in 14 of 18 infants who had shut down. Seventy-two patients (57%) had sustained (more than 1 year) relief of biliary obstruction. Postoperative morbidity was substantial. The six children not having corrective surgery died within 19 months. Three patients were lost to follow-up. Sixty-eight patients having Kasai's operation died, 55 from complications of liver disease, 1 from a coexisting malformation, and 12 after liver transplantation. Fifty-seven patients are alive, 13 by virtue of liver replacement, 9 with mild-to-moderate hepatic sequelae, and 35 (28%) with normal to near-normal liver function. Although none is considered "cured," the 35 children are anicteric, have normal growth and development, and participate in full school activities (including contact sports). Average follow-up is 85.8 months (range 1 to 15 years).     

High throughput parallel analysis of hundreds of patient samples for more than 100 mutations in multiple disease genes

As more mutations are identified in genes of known sequence, there is a crucial need in the areas of medical genetics and genome analysis for rapid, accurate and cost-effective methods of mutation detection. We have developed a multiplex allele-specific diagnostic assay (MASDA) for analysis of large numbers of samples (> 500) simultaneously for a large number of known mutations (> 100) in a single assay. MASDA utilizes oligonucleotide hybridization to interrogate DNA sequences. Multiplex DNA samples are immobilized on a solid support and a single hybridization is performed with a pool of allele-specific oligonucleotide (ASO) probes. Any probes complementary to specific mutations present in a given sample are in effect affinity purified from the pool by the target DNA. Sequence-specific band patterns (fingerprints), generated by chemical or enzymatic sequencing of the bound ASO(s), easily identify the specific mutation(s). Using this design, in a single diagnostic assay, we tested samples for 66 cystic fibrosis (CF) mutations, 14 beta-thalassemia mutations, two sickle cell anemia (SCA) mutations, three Tay-Sachs mutations, eight Gaucher mutations, four mutations in Canavan disease, four mutations in Fanconi anemia, and five mutations in BRCA1. Each mutation was correctly identified. Finally, in a blinded study of 106 of these mutations in > 500 patients, all mutations were properly identified. There were no false positives or false negatives. The MASDA assay is capable of detecting point mutations as well as small insertion or deletion mutations. This technology is amenable to automation and is suitable for immediate utilization for high-throughput genetic diagnostics in clinical and research laboratories.      

Mouse homologues of the human AZF candidate gene RBM are expressed in spermatogonia and spermatids, and map to a Y chromosome deletion interval associated with a high incidence of sperm abnormalities

An RNA-binding motif (RBM) gene family has been identified on the human Y chromosome that maps to the same deletion interval as the 'azoospermia factor' (AZF). We have identified the homologous gene family (Rbm) on the mouse Y with a view to investigating the proposal that this gene family plays a role in spermatogenesis. At least 25 and probably >50 copies of Rbm are present on the mouse Y chromosome short arm located between Sry and the centromere. As in the human, a role in spermatogenesis is indicated by a germ cell-specific pattern of expression in the testis, but there are distinct differences in the pattern of expression between the two species. Mice carrying the deletion Yd1, that maps to the proximal Y short arm, are female due to a position effect resulting in non-expression of Sry ; sex-reversing such mice with an Sry transgene produces males with a high incidence of abnormal sperm, making this the third deletion interval on the mouse Y that affects some aspect of spermatogenesis. Most of the copies of Rbm map to this deletion interval, and the Yd1males have markedly reduced Rbm expression, suggesting that RBM deficiency may be responsible for, or contribute to, the abnormal sperm development. In man, deletion of the functional copies of RBM is associated with meiotic arrest rather than sperm anomalies; however, the different effects of deletion are consistent with the differences in expression between the two species.      

L1 knockout mice show dilated ventricles, vermis hypoplasia and impaired exploration patterns

L1 is a neural cell adhesion molecule mainly involved in axon guidance and neuronal migration during brain development. Mutations in the human L1 gene give rise to a complex clinical picture, with mental retardation, neurologic abnormalities and a variable degree of hydrocephalus. Recently, a transgenic mouse model with a targeted null mutation in the L1 gene was generated. These knockout (KO) mice show hypoplasia of the corticospinal tract. Here we have performed further studies of these KO mice including magnetic resonance imaging of the brain, neuropathological analysis and behavioral testing. The ventricular system was shown to be abnormal with dilatation of the lateral ventricles and the 4th ventricle, and an altered shape of the Sylvius aqueduct. Additionally, the cerebellar vermis of the KO mice is hypoplastic. Their exploratory behavior is characterized by stereotype peripheral circling reminiscent of that of rodents with induced cerebellar lesions.