Biomedical subjects
Publications and source records attributed to R Ohi.
Although the prognosis of biliary atresia has been dramatically improved in the era of liver transplantation, the Kasai operation is still the first line of surgical treatment. Successful hepatic portoenterostomy depends on early diagnosis and surgery, adequate surgical technique, prevention of cholangitis, and precise postoperative management.
Neuroblastoma (NB) is a well-known malignant disease in infants, but its molecular mechanisms have not yet been fully elucidated. To investigate the genetic contribution of abnormalities on the long arm of chromosome 14 (14q) in NB, we analysed loss of heterozygosity (LOH) in 54 primary NB samples using 12 microsatellite markers on 14q32. Seventeen (31%) of 54 tumours showed LOH at one or more of the markers analysed, and the smallest common region of allelic loss was identified between D14S62 and D14S987. This region was estimated to be 1-cM long from the linkage map. Fluorescence in situ hybridization also confirmed the loss. There was no statistical correlation between LOH and any clinicopathologic features, including age, stage, amplification of MYCN and ploidy. We further constructed a contig spanning the lost region using bacterial artificial chromosome and estimated this region to be approximately 1.1-Mb by pulsed-field gel electrophoresis. Our results will contribute to cloning and characterizing the putative tumour-associated gene(s) in 14q32 in NB.
The conserved CDC5 family of Myb-related proteins performs an essential function in cell cycle control at G(2)/M. Although c-Myb and many Myb-related proteins act as transcription factors, herein, we implicate CDC5 proteins in pre-mRNA splicing. Mammalian CDC5 colocalizes with pre-mRNA splicing factors in the nuclei of mammalian cells, associates with core components of the splicing machinery in nuclear extracts, and interacts with the spliceosome throughout the splicing reaction in vitro. Furthermore, genetic depletion of the homolog of CDC5 in Saccharomyces cerevisiae, CEF1, blocks the first step of pre-mRNA processing in vivo. These data provide evidence that eukaryotic cells require CDC5 proteins for pre-mRNA splicing.
In eukaryotes, G2/M progression is mediated by activation of mitosis promoting factor (MPF). To ensure faithful chromosome segregation, the activity of key mitotic inducers and inhibitors are coupled with chromosome replication, spindle pole duplication, morphogenesis, and DNA damage. Evidence gathered in the past two years has underscored the importance of positioning MPF and its regulators in the proper place at the proper time to ensure orderly progression through the G2/M transition. Altering the spatial organization of G2/M regulators also contributes to prevention of mitosis following DNA damage.
Gastric adenocarcinoma is one of the most common gastrointestinal (GI) malignancies, especially among Japanese adults, but represents only 0.05% of all malignant pediatric GI tumors. We report a case of gastric adenocarcinoma arising at the esophageal gastric junction of an 11-year-old girl. The tumor was polypoid, measuring 3.0 x 3.0 x 1.2 cm and was light gray and partially red in color with a stalk. Light microscopic examination of the lesion demonstrated adenocarcinoma of variable degrees of both architectural and nuclear atypia with invasion into the submucosa. Immunohistochemical findings of cytokeratin subtypes revealed positive immunoreactivity for cytokeratin subtypes 8, 19 and 20 and negative for 5/6/18, 7, 13 and 14, which is consistent with those of gastric adenocarcinoma. The patient was alive and well 12 months postoperatively.
The Arp2/3 complex is an essential component of the actin cytoskeleton in yeast and is required for the movement of actin patches. In an attempt to identify proteins that interact with this complex in the fission yeast Schizosaccharomyces pombe, we sought high-copy suppressors of the S. pombe arp3-c1 mutant, and have identified one, which we have termed asp1(+). The asp1(+) open reading frame (ORF) predicts a highly conserved protein of 921 amino acids with a molecular mass of 106 kD that does not contain motifs of known function. Neither asp1(+) nor its apparent Saccharomyces cerevisiae ortholog, VIP1, are essential genes. However, disruption of asp1(+) leads to altered morphology and growth properties at elevated temperatures and defects in polarized growth. The asp1 disruption strain also is hypersensitive to Ca+ ions and to low pH conditions. Although Asp1p is not stably associated with the Arp2/3 complex nor localized in any discrete structure within the cytoplasm, the asp1 disruption mutant was synthetically lethal with mutations in components of the Arp2/3 complex, arp3-c1 and sop2-1, as well as with a mutation in actin, act1-48. Moreover, the vip1 disruption strain showed a negative genetic interaction with a las17Delta strain. We conclude that Asp1p/Vip1p is important for the function of the cortical actin cytoskeleton.
Myb-related cdc5p is required for G(2)/M progression in the yeast Schizosaccharomyces pombe. We report here that all detectable cdc5p is stably associated with a multiprotein 40S complex. Immunoaffinity purification has allowed the identification of 10 cwf (complexed with cdc5p) proteins. Two (cwf6p and cwf10p) are members of the U5 snRNP; one (cwf9p) is a core snRNP protein. cwf8p is the apparent ortholog of the Saccharomyces cerevisiae splicing factor Prp19p. cwf1(+) is allelic to the prp5(+) gene defined by the S. pombe splicing mutant, prp5-1, and there is a strong negative genetic interaction between cdc5-120 and prp5-1. Five cwfs have not been recognized previously as important for either pre-mRNA splicing or cell cycle control. Further characterization of cwf1p, cwf2p, cwf3p, and cwf4p demonstrates that they are encoded by essential genes, cosediment with cdc5p at 40S, and coimmunoprecipitate with cdc5p. We further show that cdc5p associates with the U2, U5, and U6 snRNAs and that cells lacking cdc5(+) function are defective in pre-mRNA splicing. These data raise the possibility that the cdc5p complex is an intermediate in the assembly or disassembly of an active S. pombe spliceosome.
Gastric peristaltic contractions are controlled by an intrinsic electrical pacemaker located in the mid-body along the greater curve. This study was undertaken to investigate gastric motility in long-term survivors of neonatal gastric rupture who were surgically deprived of their original pacemaker. Four patients, 1 boy and 3 girls, aged between 6 and 12 years were studied. Physiological activity of the gastric remnant was assessed in terms of electrical as well as peristaltic functions by means of electrogastrography and video-recorded barium swallow study. Electrical and mechanical pacing activities were classified into normogastria or dysrhythmia (brady- or tachygastria) according to their frequencies. In these patients, ectopic pacemakers were found to be arising just distal to the site of resection along the greater curve. Electrophysiologically, one patient was diagnosed as having normogastria, and other 3 patients were found to have dysrhythmia (2, bradygastria; 1, tachygastria) on the basis of electrogastrographic analyses. In two of three patients studied further by fluoroscopy, electrical activity agreed well with peristaltic activity. In one patient, however, electrical tachygastria was associated with peristaltic bradygastria. In conclusion, an ectopic pacemaker arises in the stomach that does not remain silent after neonatal surgical loss of its own pacemaker. Noninvasive electrogastrography seems useful in assessing electrical potentials generated by the ectopic pacemaker.
Biliary atresia (BA), which is thought to result from progressive destruction of the bile ducts by a necroinflammatory process, is the most common cause of obstructive jaundice in infancy. Abnormalities in the cell turnover of remodelling ductal plates are considered one of the important aetiological factors in this disorder, but little work has been done on this topic. Programmed cell death or apoptosis was therefore examined by TdT-mediated dUTP biotin nick end labelling (TUNEL) and cell proliferation by Ki67 immunostaining in 34 cases of BA. The results were compared with normal control liver (five cases) and congenital dilatation of the bile ducts (CDB, five cases) in order to study the cell turnover or tissue dynamics of BA. The TUNEL labelling index (LI) in bile ducts (48.9 +/- 13.2 per cent) was significantly higher than that of the control normal liver (3.6 +/- 2.8 per cent) and of CDB (2.5 +/- 5.1 per cent). The Ki67 LI in the bile ducts of BA (15.0 +/- 5.57 per cent) was also significantly higher than that of CDB (8.6 +/- 5.4 per cent). No significant differences of the TUNEL and Ki67 LIs in hepatocytes were, however, observed between BA, CDB, and normal liver. The TUNEL LI was significantly higher than the Ki67 LI in the bile ducts of BA. BA is therefore associated with increased and disorganized cell turnover of the bile ducts, which is related to malformation of the ductal plate or abnormal bile duct development.
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Biliary atresia (BA) still remains one of the most intractable gastrointestinal diseases in infancy despite the concerted efforts of pediatric surgeons all over the world. The introduction of liver transplantation has revolutionized the protocols for the treatment of this condition. In this editorial, the role of hepatic portoenterostomy (the Kasai procedure) in the surgical treatment of BA in the "transplantation era" will be discussed.
Schizosaccharomyces pombe cdc5p is a Myb-related protein that is essential for G2/M progression. To explore the structural and functional conservation of Cdc5 throughout evolution, we isolated Cdc5-related genes and cDNAs from Saccharomyces cerevisiae, Caenorhabditis elegans, Drosophila melanogaster, and Homo sapiens. Supporting the notion that these Cdc5 gene family members are functionally homologous to S. pombe cdc5(+), human and fly Cdc5 cDNAs are capable of complementing the temperature-sensitive lethality of the S. pombe cdc5-120 mutant. Furthermore, S. cerevisiae CEF1 (S. cerevisiae homolog of cdc5(+)), like S. pombe cdc5(+), is essential during G2/M. The location of the cdc5-120 mutation, as well as mutational analyses of Cef1p, indicate that the Myb repeats of cdc5p and Cef1p are important for their function in vivo. However, we found that unlike in c-Myb, single residue substitutions of glycines for hydrophobic residues within the Myb repeats of Cef1p, which are essential for maintaining structure of the Myb domain, did not impair Cef1p function in vivo. Rather, multiple W-to-G substitutions were required to inactivate Cef1p, and many of the substitution mutants were found to confer temperature sensitivity. Although it is possible that Cef1p acts as a transcriptional activator, we have demonstrated that Cef1p is not involved in transcriptional activation of a class of G2/M-regulated genes typified by SWI5. Collectively, these results suggest that Cdc5 family members participate in a novel pathway to regulate G2/M progression.
The "touchdown" polymerase chain reaction (PCR) technique has been applied to analyze expression of the neuron-specific protein, PGP9.5, and tyrosine hydroxylase (TH) genes for detection of minimal residual neuroblastoma cells in bone marrow and peripheral blood. PGP9.5 and TH gene products were not detected in any normal samples (n = 72) examined. However, in patients more than 1 year of age with stage III and IV neuroblastoma PGP9.5 mRNA was detected in six of seven bone marrow samples and in four of eight peripheral blood samples, and TH mRNA in four of seven and three of eight, respectively. The detection sensitivity was up to 10(-6) to 10(-7) micrograms of total cellular RNA for PGP9.5 and 10(-4) micrograms for TH. Among forty bone marrow specimens from nineteen patients with neuroblastoma both PGP9.5 and TH mRNAs were detected in six, and only PGP9.5 mRNA was detected in ten. Since detection of PGP9.5 and TH gene transcripts by the "touchdown" PCR was highly specific and sensitive, it might be most informative at present to carry out both PGP9.5 and TH mRNA assays for minimal residual neuroblastoma cells in blood and bone marrow.
We report a nine-month-old boy with stage III B hepatoblastoma of caudate lobe origin. Surgical resection was attempted following six courses of chemotherapy, but viable tumor cells remained microscopically at resection margins. Subsequently, he received peripheral blood stem cell transplantation (PBSCT), whose preparative regimen being consisted of carboplatin, etoposide, tetrahydropyranyl adriamycin, and melphalan. Since then, the patient shows no relevance of local relapse or distant metastasis without any chemotherapy. PBSCT for patients with post-operative residue may improve the outcome of advanced hepatoblastoma and worth of a further clinical investigation.
Hepatic and serum bile acids in five patients with biliary atresia were preoperatively determined by microanalysis using gas chromatography-mass spectrometry with negative ion chemical ionization detection. The hepatic content of total bile acids was markedly elevated (3079+/-711 nmol/g protein), most of which were primary bile acids. Accumulation of unconjugated bile acids (2.93% to 4.62% of the total) was observed in the liver tissue of these patients, although only trace amounts were detected in their sera. The ratio of glycine-conjugated to taurine-conjugated bile acids was 0.44+/-0.18 in liver tissue and 0.79+/-0.52 in serum and these values were significantly lower than those of controls. This study has shown that the composition of bile acids in serum does not reflect that in liver tissue faithfully. The accumulation of these hydrophobic bile acids may contribute to initiating or exacerbating liver injury in infants with cholestatic liver diseases.
The clinical and postsurgical TNM classifications (cTNM and pTNM) for neuroblastoma (NB), nephroblastoma (WT) and soft tissue sarcomas were presented in 1982 by the TNM Committee in UICC in collaboration with SIOP. The Japanese TNM Committee proposed new pTNM systems (J-pTNM) for NB and WT, and new cTNM and pTNM system for primary liver carcinoma in infants and children (HT). These pTNMs were based on the staging systems developed by the Malignant Tumor Committee of the Japanese Society of Pediatric Surgeons. The proposal of subdivision of M category in NB was presented for testing the new telescopic ramifications of TNM. The TNM for HT was added as a new classification recommended for testing. The effectiveness of these TNM systems was assessed using NB, WT and hepatoblastoma (HB) cases which were registered in collaborating institutes. The analyses suggested that pTNM, especially the J-pTNM system in NB, WT and HT were effective for the assessment of prognoses, although cTNM systems were not enough to assess the extent of the disease.
The profile of fecal bile acids was examined in 13 children with short bowel syndrome; 7 of the 13 did not have diarrhea and the other 6 had intractable diarrhea. In children without diarrhea, no severe fat malabsorption was recognized, and the content of total bile acids in the feces was within the normal range or slightly higher. The ratio of primary to total bile acids showed various patterns. In children with intractable diarrhea, in contrast, fat malabsorption was observed and the fecal content of total bile acids in these patients was more than ten times higher than that of the control group, primary bile acids accounting for more than 95% of the total bile acids and taurine- or glycine-conjugated bile acids for 10%. In the children with intractable diarrhea, the values for the D-xylose absorption test were lower than the normal range. These results suggested that, in children with short bowel syndrome with diarrhea, the loss of bile acids was strongly associated with a decrease in the actual absorptive surface area of the residual small intestine, and the growth of the normal bacterial flora was disturbed in the residual intestine. Some children with or without diarrhea also had hyper bile acidemia. Ursodeoxycholic acid was not effective for the treatment of hyper bile acidemia or fat malabsorption.
Eleven children, 4 males and 7 females, with biliary atresia receiving living related liver graft were studied. The mean age was 1.8 years and the mean body weight was 10.3 kg. The donors were 4 fathers and 7 mothers. The graft was the lateral segment or left lobe. ABO blood group matching was compatible in 9 and incompatible in 2. All patients except one were crossmatch negative. Immunosuppression at induction was triple therapy (cyclosporine, azathioprine and steroid) or FK506 plus steroid. Acute rejection episodes were treated with pulse steroids. When the signs of rejection persisted despite steroid pulse therapy, 15-deoxyspergualin (DSG) was added. The survival rate of the patients was 73%. Three patients died of portal vein thrombosis, hepatic artery thrombosis and sepsis respectively. Other major complications included hyperbilirubinemia, bile duct stenosis, bile leakage and portal vein anastomosis narrowing. Complications of the donor were sepsis in one, and liver dysfunction in two. Although there are some complications related to graft size mismatch and operative procedure, living related partial liver transplantation is an effective therapy in countries where donor source is restricted.