Neuroendocrine changes in colon of mice with a disrupted IL-2 gene

Neuroendocrine peptides have a variety of physiological functions in the gastrointestinal tract. This study was carried out to investigate the impact of IL-2 deficiency on the neuroendocrine system in normal colon, and the neuroendocrine changes during colonic inflammation. Mice with homozygous disrupted IL-2 gene (IL-2-/-) spontaneously developed a bowel disease with similarities to human ulcerative colitis. Different types of colonic endocrine cells and myenteric nerves were analysed in the IL-2-/- mice using immunomorphometry. The neuropeptide contents in the colonic tissues were determined by radioimmunoassay. Age-matched healthy IL-2+/- and IL-2+/+ mice served as controls and the colonic IL-2 levels were compared between these two groups of mice by ELISA. Our data showed that less than half the amount of IL-2 was synthesized in the colon of IL-2+/- mice compared with the IL-2+/+ wild-type mice. Two major differences in the neuroendocrine colon were found between the mice with an intact and disrupted IL-2 gene. One was age-related. The frequencies of various endocrine cells and myenteric nerves increased with age in the IL-2+/+ mice. However, no such increases were seen in the mice with a disrupted IL-2 gene. Instead, the volume densities of enteroglucagon, serotonin cells and substance P (SP), vasoactive intestinal polypeptide (VIP) and total myenteric nerves were lower in the older IL-2+/- and IL-2-/- mice compared with the wild type. The other was disease-related. Polypeptide YY (PYY) cells and tissue levels of PYY, SP and VIP were significantly decreased in the IL-2-/- mice during the course of bowel inflammation compared with the healthy IL-2+/- and IL-2+/+ controls. These findings indicate that colonic neuroendocrine alterations did occur in the mice with a disrupted IL-2 gene and diminished local IL-2 level, suggesting a role of IL-2 in the regulation of the neuroendocrine system and a prevalent interaction between the immune and neuroendocrine systems in normal colon. On the other hand, there were some changes that seemed to correlate with the bowel inflammatory process. They might be associated with the impaired function in inflamed gut and contribute to the development and/or prolongation of disease.     

A psoriasis vulgaris protective gene maps close to the HLA-C locus on the EH18.2-extended haplotype

We determined the molecular haplotypes of the HLA-A, HLA-C and HLA-B loci and the MHC class I-B-related (MIB) microsatellite in 179 unrelated psoriatic patients (72 familial cases) and in 120 controls. The HLA-A*3002-Cw*0501-B*1801-MIB1 haplotype showed a strong negative association with psoriasis vulgaris (PV) and in particular with familial PV, revealing the presence of a PV-protective gene. Analysis of association and linkage disequilibrium of the single alleles and the various two-three-four-locus segments of this haplotype indicated the presence of a protective gene telomeric to the HLA-C locus. This finding was confirmed in 13 informative multiplex PV families, in which at least one parent carried the EH18.2 haplotype. In two families, an affected sibling presented HLA-A/C recombination on the EH18.2 haplotype. A study of 12 polymorphic microsatellites in all members of the informative families, 145 PV patients, 120 controls and 32 EH18.2 homozygous healthy individuals demonstrated that the protection conferred by the EH18.2 haplotype lies within a 170 kb interval between the C143 and C244 loci, most probably in a 60 kb segment between the C132 and C244 loci.     

Detection of antigen receptor gene rearrangements in lymphoproliferative malignancies by fluorescent polymerase chain reaction

Abstract: Monoclonal rearrangements of antigen receptor genes in lymphoproliferative diseases are characterized by the specific sequence and the length of their junctional region, which can be used as markers of the proliferating clone. PCR techniques have greatly simplified routine detection of monoclonal rearrangements. But on the one hand, identification of the sequences requires sequencing methods and on the other hand, sizing of rearrangements by conventional analysis of PCR products on agarose or nondenaturing polyacrylamide gels may be uncertain. We have developed an approach based on amplification of rearranged IGH, TCRG and TCRD locus by fluorescent PCR associated to a computerized analysis of generated PCR products allowing their objective sizing. We tested this method on DNA samples from patients with acute lymphoblastic leukemia and chronic lymphocytic leukemia, whose pattern of IGH and TCRG rearrangements had been previously identified by Southern blot techniques. TCRG-PCR assay allowed detection of 100% of rearranged samples. No false-negative results were found but a high rate (60%) of Southern-negative and PCR-positive samples were identified. TCRD PCR-assay detected VD1JD1 or VD2-D2/3 rearrangements in both acute lymphoblastic leukemia and chronic lymphocytic leukemia samples. IGH PCR assay permitted detection of all known rearranged samples. The sensitivity of these three different PCR assays (1% leukemic cells) was equivalent to that of other published PCR protocols. These results show the validity and reliability of the fluorescent PCR method for routine detection of IGH, TCRG and TCRD rearrangements. Sizing of PCR products by computerized analysis was also validated. It provides additional information on rearrangement patterns in lymphoproliferative diseases, as clonal rearrangements can be recognized by their size. This can be of great interest in various circumstances, particularly for detection and follow-up of oligoclonality.     

Follicular fluid renin concentration and IVF outcome

Total renin protein concentration (TRC) was measured in stored follicular fluid (FF) samples from 42 women. Samples were selected according to their origin from follicles either without recovered ova ('empty', n = 38) or fertilized but with failed implantation ('failed', n = 36) or successful deliveries ('deliveries', n = 71). Ratios of number of embryos transferred to number of infants delivered were 2:1, 3:1 or 4:2 but 1:1 was not available. Non-parametric testing was applied to FF-TRC, volume and outcome. TRC was significantly higher in the delivery than the failed (P = 0.001) or empty (P = 0.002) categories. Assuming that the range of renin in failed follicles can identify the sub-population of unsuccessful follicles in the delivery category, then elevated FF-TRC was clearly associated with successful outcome. For individual women, the odds of infant delivery increased 17-fold as a function of average FF-TRC between 10,000 and 25,000 microIU/ml. For failed and delivery but not empty follicles, higher renin levels occurred in the smaller follicles, consistent with a burst of renin synthesis associated with the presence of an oocyte. The results suggest that FF-TRC relates to ovum viability with ovarian hyperstimulation and may have predictive use in IVF programmes.     

A PCR–restriction fragment length polymorphism assay to genotype human metapneumovirus

Human metapneumovirus (hMPV) genotypes A and B show epidemiological and probably clinical differences. This report describes a fast and simple PCR–restriction fragment length polymorphism (PCR-RFLP) assay, involving digestion of the fusion protein gene with Tsp 509I, that allows lineages A1, A2, B1 and B2 to be distinguished. The assay should help in elucidating the epidemiology of hMPV, and possibly in predicting the severity of clinical infection.     

IV型II类反式激活因子基因新变异体的克隆、鉴定和功能测定

为获取有功能的IV型II类反式激活因子基因 (CIITA IV ) ,诱导肿瘤细胞表达MHCII类分子 ,从IFN γ刺激的THP 1细胞中以RT PCR获得CIITA IV ,将其连接到pGEMT easy载体。对所构建的pcDNA3 1 CIITA IV型表达载体进行反复测序后发现 ,所获得的CIITA IV基因存在结构变异 ,在 2 87位插入了 3个核苷酸TAG ,使 2 86 2 88位的AAG改变成为ATAGAG(2 86 2 90 ) ,并引起其他 8个座位核苷酸 (及推导的氨基酸残基 )发生改变。将表达载体转入原先不表达MHCII类分子的HeLa细胞中 ,检测到所获得的IV型CIITA变异体具有诱导人II类分子HLA DR表达的能力。空载体和CIITA IV基因导入的HeLa细胞中 ,DR阳性细胞百分率分别为 0 0 1 %和 37 6 4 %。该基因已从GenBank得到登录号 ,表明这是一个具有诱导HLA DR分子表达功能的IV型CIITA新基因。     

Gene polymorphisms and serological markers of patients with active Crohn's disease in a clinical trial of antisense to ICAM-1

Serological profiles for anti-Saccharomyces cerevisiae antibodies (ASCA)/ perinuclear antineutrophil cytoplasmic antibodies (pANCA) and gene polymorphisms in tumour necrosis factor (TNF)-alpha and intercellular adhesion molecule-1 (ICAM-1) are associated with occurrence and/or outcome in Crohn's disease. The aim of the study was to characterize the ASCA/pANCA profile, soluble ICAM-1 expression and single nucleotide gene polymorphisms (SNPs) in TNF-alpha and ICAM-1 genes. Crohn's patients with moderate disease activity were enrolled in a clinical trial of Alicaforsen (ISIS 2302). Peripheral blood samples were collected prospectively for serum studies and for potential analysis of gene polymorphisms. A multivariate analysis was performed to compare treatment effect with the biomarkers studied. Serological testing for ASCA/pANCA was obtained for 257 patients at baseline: 37% were ASCA(+)/pANCA(-) (Crohn's pattern), 9% had both markers, 15% were ASCA(-)/pANCA(+) and 39% had neither marker. When the data were analysed by multiple regression analysis, a trend was found within the Alicaforsen-treated groups for greater rates of remission in the ASCA(+)/pANCA(-) subgroup versus all other serological profiles (25 versus 14%, P = 0.068), but not versus the placebo remission rate (18.8%). Gene polymorphisms were assessed in 64 patients, 21 from the placebo group. ICAM-1 assessment revealed no over-representation. However, three unique TNF-alpha SNPs were identified that correlated significantly with remission; sites 290 (P = 0.0253), -2735 (P = 0.0317) and -3090 (P = 0.0067). Although the overall clinical trial was negative, we have identified a trend towards clinical remission with Alicaforsen therapy in a subgroup of patients with Crohn's disease expressing ASCA(+)/pANCA(-). Furthermore, we have identified three TNF-alpha SNPs that may also predict a positive therapeutic outcome.     

Comparative study on deletions of the dystrophin gene in three Asian populations

The frequency and distribution of deletions of 19 deletion-prone exons clustered in two hot spots in the proximal and central regions of the dystrophin gene were compared in three populations from Singaporean, Japan, and Vietnam. DNA samples obtained from 105 Singaporean, 86 Japanese, and 34 Vietnamese Duchenne muscular dystrophy patients were examined by polymerase chain reaction amplification. Deletions of the examined exons were found in 51.2% of Japanese patients but in 40.0% or less of the Singaporeans and Vietnamese. About two thirds of the deletions were localized in the central region and the remaining deletions were clustered at the proximal region. The most commonly deleted exons at the central deletion hot spot were exon 50 in the Singaporean, exons 49 and 50 in the Japanese, and exon 51 in the Vietnamese population. At the proximal deletion hot spot, the most commonly deleted exons were exons 6 and 8 in the Singaporeans, exons 12 and 17 in the Japanese, and exons 8 and 12 in the Vietnamese. Two cases each from Singapore and Japan had large-scale gross mutations spanning both deletion hot spots. Our results suggest that, although the presence and frequency of the two deletion hot spots may be similar in the three Asian populations analyzed, the distribution and frequency of deletions among the different exons can vary as a result of population-specific intronic sequences that predispose individuals to preferential deletion breakpoints. Received: May 20, 2002 / Accepted: July 1, 2002     

华南、华北人群HLA-DQB1等位基因多态性

目的从基因水平调查了中国华南、华北地区人群HLA-DQB1等位基因频率，并研究比较两地区人群HLA-DQB1多态性分布。方法采用深圳益生堂生物企业有限公司研制开发的“HLA-DQB1低分辨率分型基因芯片检测试剂盒”，应用聚合酶链反应．序列特异性引物＋序列特异性寡核苷酸探针芯片检测技术，对700名南方地区的中国人和320名北方地区的中国人进行基因分型。结果鉴定了10个HLA-DQB1等位基因，获得了一组准确、科学的统计数据。结论得到了中国华南、华北地区人群HLA-DQB1等位基因频率差异的数据，证明中国人群HLA-DQB1＊02，05，0601，0602，0603的分布南北差异有统计学意义（P〈0．05），为疾病相关性研究、人文科学研究提供了可靠的遗传学数据。