药物性肝衰竭研究进展

药物性肝衰竭(DILF)比例高,损伤机制多样,多数固有型和部分特异质型DILF可能与线粒体功能障碍、氧化应激、胆汁酸稳态改变有关,特异质性DILF是患者自身适应性免疫攻击的结果。DILF预后较差,是死亡和肝移植的重要原因,目前较多研究致力于重症化的早期预测和预后判断,产生了不同预测模型和生物学标志。     

肝移植患者术后门静脉血栓发生病因及预后分析

目的 探讨肝移植术后患者门静脉血栓(PVT)发生的病因.方法 回顾性分析2015年11月至2019年2月解放军总医院第五医学中心401例肝移植患者临床资料,分析肝移植后发生PVT的病因.结果 401例肝移植患者中,15例出现PVT,发生率为3.7％,男性9例,女性6例,平均年龄50.8岁(35～63岁).术前乙肝肝硬化...     

1种血清戊型肝炎病毒荧光定量RT-PCR快速检测方法的建立

目的建立一种从血清样本中基于荧光定量PCR的快速检测戊型肝炎病毒的方法。方法 (1)从GeneBank获得包含我国流行的3个基因型的100条戊型肝炎病毒序列。选择合适的序列设计合成荧光定量引物和Tanqman探针。(2)将引物从血清模板中PCR扩增片段在体外转录产物作为荧光定量PCR标准品,同时引入了微量核酸裂解液。(3)取10份临床戊肝阳性患者血清样品,用建立的方法检测,进一步验证该方法。结果该检测技术可以有效检测I型和IV型戊型肝炎阳性患者的血清样本,而对其他病毒传染病患者血清检测为阴性结果,证实该RT-PCR检测技术的特异度较高、可靠性较好。对10份临床血清样品的验证检测进一步证实了该方法快速、便捷、灵敏且重复性好。结论建立了一种适用于中国人群戊型肝炎病毒主要基因型检测的荧光定量RT-PCR检测技术。可满足戊型肝炎病毒临床早期快速诊断的需求。     

三明治培养新生实验小型猪肝细胞的功能与形态

目的 探索双层胶原夹心猪肝细胞的方法，观察培养猪肝细胞的功能与形态特征。方法 将两步法分离的新生中国实验小型猪肝细胞接种于胶原被覆的培养板常规培养，24h后，肝细胞上再加一层胶原形成双层胶原夹心（三明治）培养，观察培养猪肝细胞的白蛋白分泌、形态学特征和乳酸脱氢酶（LDH）漏出量。结果 成功将新生实验小型猪肝细胞固定于双层胶原中，形成三明治状。培养1周内，用SDS／PAGE可检测出猪肝细胞分泌的白蛋白，倒置相差显微镜下观察到典型的形态特征，培养期内LDH漏出量较少。结论 可用双层胶原对新生乳猪肝细胞进行三明治样培养，为肝细胞提供更接近体内的培养环境，有助于维持特殊的肝功能和形态学特征。     

慢性重型乙型肝炎预后指标分析

目的：探讨慢性重型乙型肝炎血清甲胎蛋白（AFP）、胆碱酯酶（CHE）、凝血酶原活动度（PA）、总胆红素（TBIL）、重型肝炎分期等指标与预后的关系。方法：检测慢性重型乙型肝炎148例入院时AFP、TBIL、CHE、PA的水平,分析其与预后和肝炎分期的关系。结果：本组患者临床治愈好转率31．8％。治愈好转组与无效组入院时AFP、PA水平比较,差异显著（P〈0．05）;重型肝炎早期患者与晚期患者入院时AFP水平比较,差异显著（P〈0．05）。结论：AFP、PA、重型肝炎分期是判断重型乙型肝炎预后的重要及可靠指标,且AFP、PA的信息反应较早,更有助于进行早期判断预后。     

MicroRNA-A6增效的HEV体外瞬时转染HepG2复制体系的建立及鉴定

目的验证HEV编码的MicroRNA-A6对全长HEV瞬转HepG2细胞系中病毒复制的增效作用。方法按照已报道序列合成miR-A6,利用重组PCR、基因克隆以及体外转录技术获得单一高浓度的HEV RNA。HEV RNA与miR-A6按比例采用脉冲电流转染HepG2细胞,转染后24、48和96 h后观察细胞上清IgG分泌和HEV RNA表达载量,与不加miR-A6的HepG2细胞进行比较,分析miR-A6对HEV复制的作用,然后加入anti-A6抑制miR-A6表达,分析其对HEV表达和复制的影响。结果 MiR-A6∶HEV RNA质量比为0.5∶1和1∶1,共转染HepG2细胞后48 h,与单转染HEV RNA相比,lg值升高了1.57和2.44。HEV-IgG先于HEV RNA表达。体外实验显示,Ⅰ型和Ⅳ型HEV复制力没有明显差别,都在转染后48 h达到峰值,加入anti-A6抑制miR-A6后HEV-IgG抑制47.12%,HEV RNA抑制43.3%。结论 MiR-A6可以在体外显著增加HEV RNA瞬转HepG2细胞系中HEV病毒的复制力,为后续HEV分子病毒学和表型耐药研究奠定基础。     

Design and activity evaluation of deoxyribozymes specifically targeting hepatitis C virus RNA

Objective: To explore the cleaving and inhibitory activity of hepatitis C virus ( HCV)-specific deoxyribozymes (DRz) at both molecular and transgeneic cellular levels. Methods: According to the secondary structure of HCV 5′-noncoding region (5′-NCR) and the sites characterized with 5′…Y ↓ R…3′ ( Y = A/G, R = U/C), HCV-specific naive deoxyribozymes were designed and named DRz-232, DRz-127, DRz-84, DRzl, and the phosphorothioate deoxyribozymes (PSDRz) and mutated phosphorothioate deoxyribozymes (MPSDRz) were also designed. HCV RNA 5′-NCR was transcribed in vitro from linearized plasmid pHCV-neo and radiolabelled at its 5′-end. DRz, PSDRz or MPSDRz was respectively mixed with the substrate RNA and incubated under appropriate conditions, the cleaved products were displayed by 8% denaturated polyacrylamide gel electrophoresis (PAGE) and autoradiography, and the optical density of each band was measured to calculate cleavage rates. After that, every kind of DRz was added respectively to the cultured transgeneic HepG2 cells containing luciferase gene controlled by HCV 5′-NCR. The ceils were lysed at intended time points and the activity of luciferase was measured with chemiluminescence method for calculating inhibition rates. Results: After incubated for 90 rain in vitro, the cleavage rates of DRz-127, PSDRz-127, DRzl and PSDRzl reached 32.6%, 30.8%, 24.3% and 21.5%, respectively. No cleavage product was observed in any MPSDRz. DRz-127, PSDRz-127, DRzl and PSDRzl had an inhibitory rate of 53.2%, 50.6%, 44.7% and 43.3% respectively in transgeneic HepG2 cells in the first 24 h when the final dose of the DRz was 0.5 la.moL/L, higher than that of the corresponding MPSDRz. There was no significant difference between the inhibitory effect of each DRz and its PSDRz in HepG2 cells, but the inhibitory rate of DRz decreased more rapidly than that of the latter with the elapse of time. The results from transfection groups were significantly better than those of non-transfection groups. Conclusion:Rationally-designed HCV-specific deoxyribozymes are able to cleave target RNA at molecular level in vitro, and efficiently inhibit the expression of luciferase gene controlled by HCV 5′-NCR in transgeneic cells. Appropriate PSDRz may be more stable, and thus more suitable than the naive DRz in the application to cells. Introduction of the deoxyribozymes with transfection is more efficient than with direct delivering ways.     

肝移植后患者外周血Tfh细胞亚群的变化特征及意义

目的 探讨肝移植术前和术后患者外周血滤泡辅助性T细胞(Tfh)亚群的表达变化及其与预后的相关性.方法 对11例肝移植术后肝功能稳定的患者进行随访研究,采用流式细胞术检测移植前和移植后4周外周血中Tfh细胞亚群频率的变化.结果 与移植术前比较,移植术后4周患者外周血Tfh1 (CD4+CXCR5+CXCR3+CCR6)细胞频率较移植前有下降趋势,但差异无统计学意义(P＞0.05),Tfh2(CD4+CXCR5+CXCR3-CCR6)细胞频率较移植前明显升高(P＜0.05),Tfh1 7(CD4+CXCR5+CXCR3-CCR6+)细胞频率较移植前有上升趋势,但差异无统计学意义(P＞0.05).结论 Tfh2和Tfh17细胞可能与肝移植后患者肝功能的恢复有关.     

原发性肝淀粉样变性1例报告

淀粉样变性,又称淀粉样物质沉积症,是以淀粉样物质在各种组织和脏器细胞外间隙沉积为特征的一种进行性疾病,主要累及心、肝、肾、脾、胃肠、肌肉、皮肤、神经系统等部位,当发生于肝脏时,则称为肝淀粉样变性。     

生物人工肝临床应用的疗效与难点

各种原因导致的肝衰竭病死率高,人工肝不仅为肝衰竭患者等待肝移植提供过渡,还为部分患者延长生存时间、争取肝脏再生、避免肝移植提供可能.肝脏具有复杂的代谢、合成、转化等功能,生物人工肝支持系统引入肝细胞,从而部分替代肝脏的解毒与生物合成的功能.随着肝细胞及其体外分离、培养,纯化技术的深入研究,国内外许多类型的生物人工肝支持系统进入临床研究[1 2].