Liver cell membrane antibody detected by protein A and isolated rabbit liver plasma membrane in sera of patients with chronic liver diseases

Using the plasma membrane fraction isolated from rabbit liver (RLPM), we detected non-species specific IgG antibody against liver cell surface membrane in the sera from the patients with chronic liver diseases. The sea were treated with dithiothreitol and iodoacetamide, and absorbed with sufficient amount of actin isolated from rabbit striated muscles. The antibody was detected by incubation of RLPM with the treated and absorbed sera and subsequent determination of IgG bound to RLPM by 125I-staphylococcal protein A. It was found mainly in the patients with autoimmune hepatitis (12 of 28) and liver cirrhosis (eight of 24). It occurred more frequently in HBsAg negative liver cirrhosis than in HBsAg positive forms (six of 13 vs two of 11). The frequency of the antibody was low in chronic hepatitis except autoimmune hepatitis, and primary biliary cirrhosis. Thus the antibody against RLPM was an immunological marker of autoimmune hepatitis and HBsAg negative liver cirrhosis. The occurrence did not correlate with those of anti-smooth muscle antibody, anti-nuclear antibody and anti-mitochondrial antibody. In two cases of autoimmune hepatitis, the antibody against RLPM decreased with clinical improvement induced by corticosteroids.     

Determination of plasma bromvalerylurea and its main metabolite by a simple high-performance liquid chromatographic method and quantitation of bromide by energy dispersive X-ray spectrometry in carbon tetrachloride-intoxicated rats

In the present study, small volumes of plasma were used for the measurement of bromvalerylurea (BVU), its metabolite, 3-methylbutyrylurea (MVU), and bromide in carbon tetrachloride (CCl4)-treated rats by HPLC-UV and energy dispersive X-ray spectrometry. A liquid-liquid extraction system was also investigated. BVU and MVU were extracted from 100 microl plasma samples in a single-step involving deproteination with 1 M hydrochloric acid using ethenzamide as internal standard. Samples were separated by HPLC in an acetonitrile-8 mM potassium dihydrogenphosphate buffer (35:65, v/v) mobile phase at a flow-rate of 0.4 ml/min on a 15 cm octadecylsilyl column at room temperature. Analytes were detected at a wavelength of 210 nm. The limits of quantitation for BVU, MVU and bromide are 0.1, 0.1 and 50 microg/ml, respectively. The intra-day accuracies over the range of concentrations were 95.8 to 121.1%, 97.2 to 119.7% and 96.2 to 105.8% for BVU, MVU and bromide, respectively. The inter-day accuracies were 97.7 to 115.1%, 98.3 to 111.6% and 98.3 to 102.9% for BVU, MVU and bromide, respectively. The absolute recoveries using tert.-butyl methyl ether are 96-98% for BVU and 95-98% for MVU. The decline in the plasma concentrations of BVU in olive oil-treated rats fitted a one-compartment model and the plasma MVU level reached a peak at around 1.5-2 h and then decreased gradually. The elimination of BVU in CCl4 (1 ml/kg)-treated rats was delayed and MVU production was less than that in the olive oil-treated group. However, there was no difference in the plasma levels of bromide between CCl4-treated rats and control rats. rights reserved.     

P16-immunostaining pattern as a predictive marker of lymph node metastasis and recurrence in early uterine cervical cancer.

OBJECTIVES: We investigated the relationship between P16-immunostaining patterns and clinicopathological factors in early uterine cervix cancers and assessed whether P16-immunostaining patterns predict the prognosis of the patients with early uterine cervix cancers. METHODS: Twenty-nine early squamous cell carcinoma (SCC) specimens of the uterus were examined using immunohistochemistry for P16 expression. The P16-immunostaining pattern was classified into two groups: the homogeneous type and the heterogeneous type. P16-immunostaining patterns were evaluated in different parts of the carcinoma in situ (CIS): the center of the tumor and the front interface of the infiltrating tumor. RESULTS: All specimens were of the homogeneous type in CIS. The P16-immunostaining pattern was significantly of the heterogeneous type in the front interface of the infiltrating tumor with lymphatic invasion, vascular invasion, lymph node metastasis, and recurrence. Regarding the P16-immunostaining patterns in the front interface of the infiltrating tumor, the patients with the heterogeneous type showed a significantly worse prognosis than the patients with the homogeneous type. CONCLUSIONS: The prognosis of patients with early uterine cervical SCC may be predicted by evaluating the P16-immunostaining pattern in the front interface of the infiltrating tumor.     

Two regions of homozygous deletion clusters at chromosome band 9p21 in human lung cancer

We examined 149 lung cancer cell lines for homozygous deletions using 24 DNA markers, which were mapped and ordered in chromosome band 9p21, to define the target regions for 9p21 deletions in human lung cancer. Homozygous deletions were detected in 39 (26%) cell lines and clustered at 2 independent regions. One was the region containing the p16/CDKN2A tumor suppressor gene, and this region was deleted in 32 (21%) cell lines. The other was the region containing D9S171, which is the locus approximately 3 Mb proximal to the CDKN2A locus. This region, designated as the D9S171 region, was deleted in 18 (12%) cell lines. Seven of the 18 cell lines had identical minimum deletions of a 17,036 bp sequence located 20 kb distal to the D9S171 locus. However, such a deletion was also observed in the corresponding B-lymphoblastoid cell line from 1 of the 7 cell lines and in 5 (16%) of 32 noncancerous tissues, suggesting that the deletion was a genetic polymorphism. By considering this polymorphism, 11 (7%) cell lines still had deletions at the D9S171 region. Two NSCLC cell lines showed deletions at the D9S171 region and retentions of the CDKN2A locus. Furthermore, an NSCLC cell line showed discontinuous deletions including either the CDKN2A or D9S171 locus. Therefore, the region surrounding the D9S171 locus was defined as another target region for the 9p21 deletions. It is possible that unknown tumor suppressor gene(s) are present in this chromosomal region. Genes Chromosomes Cancer 27:308-318, 2000.     

Multi-center study of seasonal affective disorders in Japan. A preliminary report

A multi-center study on seasonal affective disorder (SAD) was conducted from the autumn of 1988 to the spring of 1989 with the cooperation of 16 facilities in Japan. Forty-six SAD patients were identified among 1104 respondents to our advertisements in mass media, or patients seen at the outpatient clinics. Essentially similar findings to other previous reports were obtained in terms of onset age of the first episode, duration of episode, high proportion of depression in first-degree relatives and atypical vegetative symptoms. However, a nearly equal sex ratio, together with a high proportion of unipolar depression, is characteristic of the present study. Increased appetite and carbohydrate craving were predominant only in female patients, whereas hypersomnia was prominent in both sexes. Effective response to light therapy was found in 17 SAD patients. However, a controlled study on a large number of patients is required to allow final conclusions on the efficacy of light therapy in Japanese SAD patients.     

Transgenic rabbits with increased VEGF expression develop hemangiomas in the liver: a new model for Kasabach-Merritt syndrome

Clinical studies have provided ample evidence that high (either systemic or local) levels of vascular endothelial growth factor (VEGF) are associated with several pathophysiological disorders, including hemangiomas. To investigate whether elevated VEGF expression could directly affect these disorders, we created a transgenic (Tg) rabbit model with increased hepatic expression of the human VEGF(165) transgene under the control of the human alpha-antitrypsin promoter. Tg rabbits exhibited marked hepatomegaly, with livers 2.5-fold heavier than those of control rabbits. Histological analysis revealed that the livers of Tg rabbits showed prominent dilation of the sinusoids and formed various-sized blood vessel networks, a feature of diffuse hemangiomas. Immunohistochemical staining revealed that the hepatocytes produced VEGF(165), whereas plasma VEGF(165) was not detected. Furthermore, Tg rabbits suffered from hemolytic anemia, thrombocytopenia and splenomegaly, which was associated with marked extramedullary hematopoiesis. The manifestations of Tg rabbits mimic many of the features of hemangiomatous disorders in humans such as the Kasabach-Merritt syndrome, and therefore this model may be potentially useful for the study of the pathogenesis and complications of hemangiomas as well as the investigation of angiogenesis inhibitors.     

Single nucleotide polymorphisms in TNFSF15 confer susceptibility to Crohn's disease

The inflammatory bowel diseases (IBDs), Crohn's disease (CD) and ulcerative colitis, are chronic inflammatory disorders of the digestive tract. The pathogenesis of IBD is complicated, and it is widely accepted that immunologic, environmental and genetic components contribute to its etiology. To identify genetic susceptibility factors in CD, we performed a genome-wide association study in Japanese patients and controls using nearly 80,000 gene-based single nucleotide polymorphism (SNP) markers and investigated the haplotype structure of the candidate locus in Japanese and European patients. We identified highly significant associations (P = 1.71 x 10(-14) with odds ratio of 2.17) of SNPs and haplotypes within the TNFSF15 (the gene encoding tumor necrosis factor superfamily, member 15) genes in Japanese CD patients. The association was confirmed in the study of two European IBD cohorts. Interestingly, a core TNFSF15 haplotype showing association with increased risk to the disease was common in the two ethnic groups. Our results suggest that the genetic variations in the TNFSF15 gene contribute to the susceptibility to IBD in the Japanese and European populations.     

A novel action of stargazin as an enhancer of AMPA receptor activity

Stargazin (γ-2) is disrupted in the ataxic and epileptic mutant mouse, stargazer (stg). The striking defect in the stg cerebellum is the lack of functional AMPA receptors on granule cells. Recently, it has been reported that γ-2 and its related molecules are crucial for the surface expression, synaptic targeting and recycling of AMPA receptors, being termed collectively as the transmembrane AMPA receptor regulatory proteins (TARPs). However, it is still unclear whether TARPs directly modulate AMPA receptor activity. Here we report that coexpression of GluR1 (GluR1) with γ-2 using HEK293 cells and Xenopus oocytes markedly enhanced glutamate-induced currents. This effect was far beyond the increase of AMPA receptor surface expression and accompanied by increased glutamate affinity and subunit cooperativity. Other member of TARPs (γ-3, γ-4, and γ-8) also enhanced the current response through the AMPA receptors. The enhancing effect by γ-2 coexpression was further observed for homomeric GluR2 (GluR2) channels, which, when expressed alone, are known to produce only a small or negligible current response. These results suggest that γ-2 not only promotes AMPA receptor surface expression but also directly modulates AMPA receptor activity.     

Three novel mutations of the fibrillin-1 gene and ten single nucleotide polymorphisms of the fibrillin-3 gene in Marfan syndrome patients

Marfan syndrome (MFS) is an autosomal dominant disorder of the extracellular matrix. Allelic variations in the gene for fibrillin-1 (FBN1) have been shown to cause MFS. To date, over 550 mutations have been identified in patients with MFS and related connective tissue diseases. However, about a half of MFS cases do not possess mutations in the FBN1 gene. These findings raise the possibility that variants located in other genes cause or modify MFS. To explore this possibility, firstly we analyzed FBN1 allelic variants in 12 Japanese patients with MFS, and secondly we analyzed fibrillin-3 gene (FBN3) in patients without FBN1 mutations using conformation sensitive gel electrophoresis (CSGE) and direct sequencing analysis. We identified three novel FBN1 mutations and ten FBN3 single nucleotide polymorphisms (SNPs). In this report, we could not detect a responsible mutation of the FBN3 gene for MFS. Although the number of the cases in this report is small, at least these results suggest that disease-causing mutations in exon regions of the FBN3 gene are very rare in MFS.Nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under the accession numbers: AB177797, AB177798, AB177799, AB177800, AB177801, AB177802, AB177803