日本血吸虫基因表达序列标签的获得和分析

为寻找日本血吸虫新基因并进行血吸虫基因功能研究,随机挑选日本血吸虫（中国大陆株）成虫cDNA库克隆,培养后提取噬菌体DNA作模板,进行PCR反应扩增,将扩增产物部分测序产生表达序列标签（EST）,并将EST输入GenBank和EMBL,运用分子生物学 软件比较其同源性, 推导其蛋白序列并进行分析,结果得到75个未曾登录的新基因,并对其中一些基因进行功能推测研究,认为表达序列材人是快速有效寻找基因的方法,生物信息学软件的应用提供了新的医学研究途径。     

Primary cutaneous diffuse large B-cell lymphoma (leg type) after renal allograft: case report and review of the literature

We report a case of a 58-year-old man who presented with a rapidly growing proliferative lesion on the left lower limb, clinically resembling a soft tissue sarcoma 3 years after renal allograft. There was no evidence of systemic involvement on bone marrow needle aspiration and computed tomography (CT) scans of the chest and abdomen. The lesion turned out to be primary cutaneous diffuse large B-cell lymphoma, leg type (PCLBCL LT), as defined in the recent World Health Organization–European Organization for Research and Treatment of Cancer (WHO–EORTC) classification of cutaneous lymphomas by skin biopsy. Immunosuppression reduction, chemotherapy with CHOP regimen and local radiotherapy induced complete remission of the tumor.     

Inhibitory effects of genistein on metastasis of human hepatocellular carcinoma

AIM: To investigate the inhibitory effects of genistein on metastasis of MHCC97-H hepatocellular carcinoma cells and to explore the underlying mechanism.METHODS: MHCC97-H hepatocellular carcinoma cells were exposed to genistein. A cell attachment assay was carried out in a microculture well pre-coated with fibronectin. The invasive activity of tumor cells was assayed in a transwell cell culture chamber, and cell cycle and apoptosis were evaluated by a functional assay. In addition, the expression and phosphorylation of FAK were detected by Western blotting. In situ xenograft transplantation of hepatocellular carcinoma was performed in 12 nude mice and lung metastasis of hepatocellular carcinoma was observed.RESULTS: Genistein significantly inhibited the growth of MHCC97-H cells in vitro. Adhesion and invasiveness of MHCC97-H cells were inhibited in a concentrationdependent fashion, and the inhibitory effect of genistein was more potent in the 10 μg/mL and 20 μg/ mL genistein-treated groups. Genistein caused G0/G1 cell cycle arrest, an S phase decrease, and increased apoptosis. The expression and phosphorylation of FAK in MHCC-97H cells were significantly decreased. In situ xenograft transplantation of hepatocellular carcinoma was also significantly suppressed by genistein. The number of pulmonary micrometastatic foci in the genistein group was significantly lower compared with the control group (12.3 ± 1.8 vs 16.6 ± 2.6, P < 0.05). CONCLUSION: Genistein appears to be a promising agent in the inhibition of metastasis of hepatocellular carcinoma.     

不同脂肪含量饮食对小鼠脂肪细胞肥胖相关基因表达的影响

目的 探讨饮食中的脂肪含量对小鼠营养性肥胖症的作用.方法 将40只C57BL/6小鼠随机分为4组,分别喂养含量不同的猪油餐.第1～4组猪油所提供热量分别为O%、20%、40%和60%,单位重量饲料的总热量不变.第7周末测定小鼠的血糖、血脂等指标,并取肝脏和肾脏作病理切片,取白色脂肪组织做基因芯片分析.结果 第4组小鼠的体重、血糖、血脂、胰岛素、肥胖指数与第1组小鼠比较有统计学差异;肝细胞和肾近曲小管细胞有大最脂质沉积;脂肪细胞中的肥胖相关基因有13个的表达是2倍以上上调,有8个的表达是2倍以下下调.结论 高脂饮食通过调节小鼠脂肪细胞中刺激食欲的基因、抑制食欲的基因和能量消耗相关基因使小鼠发生肥胖、高胰岛索血症和脂质代谢异常.     

Interferon therapy in chronic hepatitis B reduces progression to cirrhosis and hepatocellular carcinoma: a meta‐analysis

Summary. Interferon (IFN) has been used in the treatment of chronic hepatitis B for decades. Beneficial effects including hepatitis B e antigen/HBV DNA seroclearance have been documented. However, the effect of treatment on the prevention of cirrhosis and hepatocellular carcinoma (HCC) development remains controversial. We conducted a meta‐analysis of available literature to evaluate whether IFN reduces the incidence of liver cirrhosis and HCC in patients with chronic hepatitis B. Twelve clinical controlled trials, including 2082 patients and comparing IFN with no treatment, were selected. Data on the incidence of liver cirrhosis and HCC in IFN treated and untreated patients were extracted from each study. The evaluation of preventive effectiveness was performed with an intention‐to‐treat method. The relative risk (RR) and 95% confidence interval (CI) of the main outcomes as a measure of efficacy were used. Meta‐analysis was performed using fixed‐effect or random‐effect methods, depending on absence or presence of significant heterogeneity. Analyses were performed with stata version 9.0 and Review Manager Version 4.2. Five studies including the data on development of liver cirrhosis, and eleven studies including the data on development of HCC were analyzed. There was no evidence for publication bias on the funnel plot or by Egger’s test, and the heterogeneity test indicated that the variation of trial‐specific RR was not statistically significant. A different incidence of liver cirrhosis and HCC was observed between treated and untreated patients. The RR of liver cirrhosis and HCC was 0.65 (95% CI: 0.47, 0.91) and 0.59 (95% CI: 0.43, 0.81), respectively. In conclusion, the results of this meta‐analysis indicate that IFN prevents or delays the development of liver cirrhosis and HCC in patients with chronic hepatitis B.     

急性肠系膜动脉缺血性疾病的诊治分析——附27例报道

目的:探讨急性肠系膜动脉缺血的诊断和治疗方法。方法:回顾性分析自2002年5月至2008年12月我院共收治肠系膜动脉缺血性疾病患者27例。急诊行手术治疗21例,其中单纯行肠系膜上动脉取栓术9例,肠系膜上动脉切开取栓加肠切除吻合术8例,单纯肠切除吻合术3例,肠切除加肠造瘘术1例。保守治疗6例。结果:手术治疗21例,1例于死于感染性休克,5例出现短肠综合征,经胃肠外营养等对症治疗后症愈出院,6例行保守治疗者均症状减轻,好转出院。结论:彩超是早期诊断肠系膜动脉缺血的重要手段,及时手术和加强术后监护是提高疗效的关键。     

Liver transplantation outcomes in 1,078 hepatocellular carcinoma patients: a multi-center experience in Shanghai, China

Purpose  To evaluate current selection criteria for patients undergoing liver transplantation (LT) in response to hepatocellular carcinoma (HCC), and to analyze the prognostic factors for successful transplantation. Methods  We evaluated the outcome of 1,078 consecutive patients with HCC from the Shanghai Multi-Center Collaborative LT Group who underwent LT over a 6-year period. Clinicopathologic data for these patients were evaluated. The prognostic significance was assessed using Kaplan–Meier survival estimates and log-rank tests. Multivariate study with Cox’s proportional hazard model was used to evaluate the prognosis-relative aspects. Results  We determined that expansion of Milan criteria to include: a solitary lesion ≤9 cm in diameter, no more than three lesions with the largest ≤5 cm, a total tumor diameter ≤9 cm without macrovascular invasion, lymph node invasion and extrahepatic metastasis (referred to as the “Shanghai criteria”), resulted in overall survival (OS) and disease-free survival (DFS) rates that were similar to the Milan criteria. Multivariate analysis using the Cox proportional hazards regression model showed that the Child-Pugh-Turcotte classification (P = 0.010, 0.000), tumor differentiation (P = 0.001, 0.000), tumor size (P = 0.000, 0.000) and number (P = 0.014, 0.016), macrovascular invasion (P = 0.022, 0.000) and alpha-fetoprotein (AFP) levels (P = 0.031, 0.003) were independent predictors of OS and DFS, while post-LT chemotherapy (OS, P = 0.000) and tumor encapsulation (DFS, P = 0.038) were independent predictors of OS or DFS. Conclusion  Shanghai criteria expanded the current criteria while maintaining similar survival. J. Fan, G.-S. Yang, Z.-R. Fu, Z.-H. Peng, Q. Xia, C.-H. Peng, J.-M. Qian, J. Zhou and Y. Xu contributed equally to this work. This is an original work by all the authors from the Shanghai Multi-center Collaborative Liver Transplantation Group.     

Allosteric modulation of Ca2+ channels by G proteins, voltage-dependent facilitation, protein kinase C, and Ca(v)beta subunits

N-type and P/Q-type Ca(2+) channels are inhibited by neurotransmitters acting through G protein-coupled receptors in a membrane-delimited pathway involving Gbetagamma subunits. Inhibition is caused by a shift from an easily activated "willing" (W) state to a more-difficult-to-activate "reluctant" (R) state. This inhibition can be reversed by strong depolarization, resulting in prepulse facilitation, or by protein kinase C (PKC) phosphorylation. Comparison of regulation of N-type Ca(2+) channels containing Cav2.2a alpha(1) subunits and P/Q-type Ca(2+) channels containing Ca(v)2.1 alpha(1) subunits revealed substantial differences. In the absence of G protein modulation, Ca(v)2.1 channels containing Ca(v)beta subunits were tonically in the W state, whereas Ca(v)2.1 channels without beta subunits and Ca(v)2.2a channels with beta subunits were tonically in the R state. Both Ca(v)2.1 and Ca(v)2.2a channels could be shifted back toward the W state by strong depolarization or PKC phosphorylation. Our results show that the R state and its modulation by prepulse facilitation, PKC phosphorylation, and Ca(v)beta subunits are intrinsic properties of the Ca(2+) channel itself in the absence of G protein modulation. A common allosteric model of G protein modulation of Ca(2+)-channel activity incorporating an intrinsic equilibrium between the W and R states of the alpha(1) subunits and modulation of that equilibrium by G proteins, Ca(v)beta subunits, membrane depolarization, and phosphorylation by PKC accommodates our findings. Such regulation will modulate transmission at synapses that use N-type and P/Q-type Ca(2+) channels to initiate neurotransmitter release.     

Three-dimensional structure of a mammalian thioredoxin reductase: implications for mechanism and evolution of a selenocysteine-dependent enzyme

Thioredoxin reductases (TrxRs) from mammalian cells contain an essential selenocysteine residue in the conserved C-terminal sequence Gly-Cys-SeCys-Gly forming a selenenylsulfide in the oxidized enzyme. Reduction by NADPH generates a selenolthiol, which is the active site in reduction of Trx. The three-dimensional structure of the SeCys498Cys mutant of rat TrxR in complex with NADP(+) has been determined to 3.0-A resolution by x-ray crystallography. The overall structure is similar to that of glutathione reductase (GR), including conserved amino acid residues binding the cofactors FAD and NADPH. Surprisingly, all residues directly interacting with the substrate glutathione disulfide in GR are conserved despite the failure of glutathione disulfide to act as a substrate for TrxR. The 16-residue C-terminal tail, which is unique to mammalian TrxR, folds in such a way that it can approach the active site disulfide of the other subunit in the dimer. A model of the complex of TrxR with Trx suggests that electron transfer from NADPH to the disulfide of the substrate is possible without large conformational changes. The C-terminal extension typical of mammalian TrxRs has two functions: (i) it extends the electron transport chain from the catalytic disulfide to the enzyme surface, where it can react with Trx, and (ii) it prevents the enzyme from acting as a GR by blocking the redox-active disulfide. Our results suggest that mammalian TrxR evolved from the GR scaffold rather than from its prokaryotic counterpart. This evolutionary switch renders cell growth dependent on selenium.