骨髓间充质干细胞通过抑制铁死亡改善大鼠重症急性胰腺炎相关肺损伤

目的:探讨骨髓间充质干细胞(BMSCs)对重症急性胰腺炎(SAP)大鼠肺组织内细胞铁死亡的抑制作用及其潜在机制。方法:选择3周龄SD雄性大鼠提取、培养BMSCs。将30只6周龄SD雄性大鼠随机分为5组：假手术组(Sham)、胰腺炎组(SAP)、PBS对照组(SAP+PBS)、干细胞治疗组(SAP+BMSCs)、干细胞联合Nrf2阻断剂ML385治疗组(SAP+BMSCs+ML385),每组6只。使用牛磺胆酸钠(NaT)构建大鼠SAP模型。于造模前2 h, SAP+BMSCs+ML385组腹腔注射ML385(5 mg/kg);于造模后6 h, SAP+BMSCs组和SAP+BMSCs+ML385组尾静脉注射BMSCs(1×10⁷/只),SAP+PBS组尾静脉注射等量的磷酸盐缓冲液(PBS)。3 d后处死大鼠,取胰腺、肺组织和血清。H&E染色观察组织损伤情况。检测血清中淀粉酶、脂肪酶活性和炎症因子水平,肺组织中铁含量、丙二醛(MDA)、谷胱甘肽(GSH)活性。检测肺组织铁响应元件结合蛋白2(IREB2)、铁蛋白重链1(FTH1)、长链脂酰CoA合成酶4(ACS...     

成人胰腺干细胞分离及转分化为胰岛的研究

目的 通过对成人胰腺干细胞分离和转分化为胰岛过程的研究以便更进一步了解及改进胰腺干细胞分离、培养、鉴定方法。方法 成人胰腺组织以胶原酶消化，密度梯度离心法获得纯化的胰腺外分泌细胞、导管上皮细胞和胰岛。导管上皮细胞在体外共培养27d，观察细胞形态学变化及干细胞特异性转录基因PDX—1，CK—19蛋白等的表达。结果 上述方法可获得大量胰腺导管上皮细胞。体外培养第1天即可见PDX—1，CK—19阳性细胞，胰腺导管上皮细胞迅速分裂增殖并转变为有分化能力的干细胞继而转分化为三维结构的胰岛细胞。培养27d后，平均每克胰腺组织可生成760个胰岛。结论 用改进的方法可获得大量成人胰腺导管上皮细胞，并可在体外转分化为大量具有内分泌功能的胰岛，可能为克服胰岛移植的供体短缺提供一条新途径。     

成人胰腺干细胞转分化为胰岛细胞过程中的形态学观察

目的：观察成人胰腺干细胞转化化为胰岛过程中的形态学变化，以便更深入了解有关机制。方法：人胰腺组织经胶原酶消化后，用梯度离心法将胰腺外分泌细胞和胰岛分离，混于外分泌组织中的导管上皮细胞即具有转分化潜能的干细胞。在体外经CMRL1066及不含血清的DMEM／F12加多种营养因子的培养液中培养27d，在培养的不同时间点取细胞作PDX－1、CK－19等单抗的免疫组化染色。光镜和电镜观察不同时间点的形态学变化。结论：上述方法可获得大量以往在胰岛分离时丢弃的胰腺导管上皮细胞。经体外一定条件的培养后，其形态学变化过程为：导管上皮细胞迅速分裂增殖转变为有分化能力的干细胞继而转分化为胰岛细胞。结论：成人胰腺的导管上皮具有干细胞潜能，并可在体外转分化为大量具有内分泌功能的胰岛。用此方法获得大量的胰岛可能为克服胰岛移植的供体短缺提供一条新的途径。     

骨髓基质干细胞及其来源的产胰岛素细胞移植对受体胰岛和新生血管的影响

Objective To investigate the proliferative impact of transplantation of bone marrow mesenchymal stem cells (MSCs) and insulin producing ceils (IPCs) from them on recipients' remnant islets and neovascularization around. Methods MSCs were induced to IPCs in vitro. The rats with type Ⅰ diabetic meilitus were randomly divided into control group, MSCs transplant group, and IPCs transplant group. When the blood glucose level of recipient rats was decreased to 10 mmol/L, the right kidney and pancreas were removed from each group simultaneously. All the thin tissue slices were estimated by immu-nohistochemistry staining of PCNA antibody,insulin antibody and CD31 anbibody. Results The passaged MSCs differentiated into IPCs. In the grafts, insulin-positive and CD31 -positive cells were observed in the right kidney. In control group, few proliferative cells located in the atrophic islets. In the transplant groups, a lot of proliferative islet cells were obsereved, around which few CD31 -positive ceils existed;Pancreatic proliferative islet cell rate in MSCs and IPCs transplant group was ( 20.84±3.48 ) % and ( 18.43± 2.84% ) ( P > 0.05 ) respectively. Conclusion Obviously, MSCs and IPCs promoted recipient neovascu-larization surrounding the islets, which enhanced the proliferation of remnant islet cells. MSCs differentiated to IPCs and vascular endothelial cells in the recipient transplant location.     

腹腔镜胆总管探查术与内镜下乳头括约肌切开取石术治疗胆总管结石中远期并发症的Meta分析

目的:评价腹腔镜胆总管探查术(LCBDE)与内镜十二指肠乳头括约肌切开术(EST)治疗胆总管结石的中、远期结石残留率与复发率。方法:检索PubMed、Embase、Cochrane Library、CBM、CNKI及Google Scholar数据库中关于LCBDE与EST治疗胆总管结石疗效比较的随机对照试验(RCT)。根据纳入标准与排除标准,由两名研究员独立进行数据提取并对文献质量进行评价;采用RevMan 5.3软件对两种术式的胆总管结石残留率与复发率进行Meta分析。结果:纳入12篇RCT文献,总样本量为1 414例,其中LCBDE组708例,EST组706例。Meta分析结果显示,两组术后胆总管结石残留率差异无统计学意义(OR=0.48,95%CI:0.23～1.03,P=0.06),术后胆总管结石复发率LCBDE组低于EST组,两组差异有统计学意义(OR=0.32,95%CI:0.19～0.53,P<0.0001)。结论:两种术式的术后结石残留率相近,但LCBDE术后胆总管结石复发率低于EST。     

经脐入路腹壁无疤痕腹腔镜胆囊切除术的临床观察

目的:探析经脐入路腹壁无疤痕腹腔镜胆囊切除术的临床疗效。方法:入选我院2010年5月～2012年5月间收治的60例具有胆囊疾患的患者,按随机原则分为治疗组和对照组各30例,分别进行单孔腹腔镜胆囊切除术和传统三孔法腹腔镜胆囊切除术,观察分析两组手术有效率、手术时间、出血情况、出院时间、并发症以及有无可见疤痕等。结果:治疗组30例患者有1例中转开腹手术手术成功,对照组30例患者全部手术成功,差异无统计学意义。治疗组平均手术时间2.1(h 1.75～2.67h),术中平均出血量5ml(2～8ml),术后无出血,术后1～3天出院,均未有胆总管损伤等并发症发生,平均随访2月(1.5～3月),未有腹壁可见疤痕;对照组平均手术时间1小时(0.5～1.5h),术中平均出血量4ml(2～6ml),术后1～3天出院,无并发症发生,平均随访2月(1.5～3月),上腹部腹壁有明显疤痕。结论:针对有胆囊疾患的患者实施经脐入路腹壁无疤痕腹腔镜胆囊切除术安全有效,无可见疤痕。     

肝细胞肝癌中RhoA基因表达及其与肿瘤临床病理特征的关系

目的 研究RhoA基因在肝细胞肝癌中的表达及其与肝癌临床病理特征的关系.方法 应用实时荧光定量RT-PCR与Western blot同步检测64例肝细胞肝癌及对应癌旁组织中RhoA mRNA及蛋白的表达情况,分析RhoA基因的表达与肝细胞肝癌临床病理参数的关系.结果 肿瘤组织中RhoA mRNA的表达量明显高于癌旁组织(t=3.445,P=0.0006),RhoA mRNA肝癌中的表达水平与肿瘤病理分期密切相关,在门静脉受侵犯(t=2.667,P=0.009)、有微卫星灶(t=2.172,P=0.038)和pTNM分期较晚(Ⅲ和Ⅳ期,t=2.551,P=0.013)的患者中明显升高.Western blot结果显示肿瘤组织中RhoA蛋白的水平明显高于癌旁组织(t=3.532,P=0.0002),RhoA蛋白的含量在门静脉受侵犯(t=2.087,P=0.042)、有微卫星灶(t=2.254,P=0.031)和pTNM分期较晚(Ⅲ和Ⅳ期;t=2.812,P=0.007)的患者中明显升高.RhoA在肝癌组织中高表达,其表达量与肿瘤侵袭能力、门静脉癌栓以及肿瘤分期相关.结论 RhoA基因可能成为一种新的肿瘤标志物,用于判断良恶性、肿瘤转移能力和估计预后.     

骨髓基质干细胞及其来源的产胰岛素细胞移植对受体胰岛和新生血管的影响

Objective To investigate the proliferative impact of transplantation of bone marrow mesenchymal stem cells (MSCs) and insulin producing ceils (IPCs) from them on recipients' remnant islets and neovascularization around. Methods MSCs were induced to IPCs in vitro. The rats with type Ⅰ diabetic meilitus were randomly divided into control group, MSCs transplant group, and IPCs transplant group. When the blood glucose level of recipient rats was decreased to 10 mmol/L, the right kidney and pancreas were removed from each group simultaneously. All the thin tissue slices were estimated by immu-nohistochemistry staining of PCNA antibody,insulin antibody and CD31 anbibody. Results The passaged MSCs differentiated into IPCs. In the grafts, insulin-positive and CD31 -positive cells were observed in the right kidney. In control group, few proliferative cells located in the atrophic islets. In the transplant groups, a lot of proliferative islet cells were obsereved, around which few CD31 -positive ceils existed;Pancreatic proliferative islet cell rate in MSCs and IPCs transplant group was ( 20.84±3.48 ) % and ( 18.43± 2.84% ) ( P > 0.05 ) respectively. Conclusion Obviously, MSCs and IPCs promoted recipient neovascu-larization surrounding the islets, which enhanced the proliferation of remnant islet cells. MSCs differentiated to IPCs and vascular endothelial cells in the recipient transplant location.     

大鼠β细胞素蛋白的表达、纯化及生物学活性鉴定

目的:获得大量重组大鼠β细胞素并检测其活性。方法:用PCR法从大鼠肾组织中扩增534bp的β细胞素基因片段,并按读框克隆到原核表达载体pET28a( )中。以构建的重组质粒pET28a-rBTC转化大肠杆菌BL-21(DE3)菌株,在IPTG诱导下表达β细胞素蛋白,表达产物用SDS-PAGE和Westernblot检测。采用镍柱亲和层析法纯化目的蛋白。结果:经IPTG诱导后,在大肠杆菌中可表达相对分子质量Mr为20000的目的蛋白。目的蛋白主要以包涵体的形式表达;表达量约占菌体蛋白总量的20%～30%。表达的目的蛋白与抗His标签抗体具有良好的反应性。结论:大鼠β细胞素基因在PET表达系统中得到高效表达;蛋白复性后能有效地促进NIH3T3细胞的体外增殖。