MRL/lpr鼠骨髓间充质干细胞对T淋巴细胞作用的研究

目的 探讨狼疮鼠（MRL／lpr）及正常鼠（C57BL／6）骨髓间充质干细胞（MSCs）体外生物学特性、对T淋巴细胞作用的影响。方法 采用直接贴壁筛选法分离培养扩增两种鼠系骨髓MSCs，流式细胞术鉴定MSCs表面标记物，观察MSCs生长状况并绘制生长曲线；取C57BL／6鼠脾脏细胞，经尼龙毛柱分选CD3T淋巴细胞．经刀豆蛋白A（ConA）刺激，取正常及狼疮鼠MSCs或MSCs培养上清与T细胞共培养72h，然后羧基荧光素二醋酸盐琥珀酰亚胺酯（CFSE）染色测定T细胞增殖，流式细胞术检测共培养前后T细胞活化及T细胞的凋亡率。结果 第2代（P2）后狼疮鼠MSCs生长增殖快于正常鼠MSCs；正常及狼疮鼠MSCs共培养均可抑制正常鼠T细胞的增殖（P〈0．05），与MSCs上清共培养对正常鼠T细胞增殖无抑制作用（P〉0．05）；与两种MSCs或上清共培养均可降低正常T细胞的活化（P〈0．05）及凋亡率（P〈0．05）。对正常T细胞增殖、活化、凋亡的影响，狼疮鼠MSCs与正常鼠MSCs间差异无统计学意义（P〉0．05）。结论 狼疮鼠MSCs在体外生长上存在异常．但对T细胞增殖、活化、凋亡免疫调节无异常。     

人肝细胞L02诱导MSCs分化为肝样细胞的观察

目的 探讨骨髓间充质干细胞(MSCs)在与肝细胞L02在体外共培养条件下诱导分化为肝细胞的可行性.方法 密度梯度离心联合贴壁筛选法获取MSCs.将接种于盖玻片上的MSCs和人肝细胞L02共置于培养皿中.在2、4、6、8 d时分别用免疫细胞化学检测AFP、细胞角蛋白(CK18),同时检测染色体核型.结果 成功分离培养出MSCs,其表型为CD29阳性,CD34阴性.共培养的MSCs可在2～8 d时,形态变为三角形、多角型或类圆形.第2天时,共培养细胞AFP表现为强阳性表达,以后减弱,第8天AFP的阳性表达很少.第2、4天检测出CK18均为阴性表达,第6、8天CK18为阳性表达.阳性对照组细胞CK18有较强的阳性表达,AFP弱阳性表达.阴性对照组均为阴性表达.共培养细胞染色体核型无融合现象.结论 人肝细胞L02能够诱导MSCs分化为肝样细胞.     

猪骨髓间充质干细胞对共培养肝细胞的作用及其机制

目的 探索细胞外基质(ECM)在猪肝细胞与骨髓间充质干细胞(MSCs)体外共培养中的表达与分布规律. 方法自中华实验猪髂前上棘抽取骨髓,采用密度梯度离心法分离单个核细胞,贴壁传代培养至第3代;原位两步胶原酶法分离猪肝细胞后与MSCs随机混合培养,观察肝细胞形态和功能的变化情况.用免疫细胞化学染色法观察ECM的表达和分布情况.RNA干扰MSCs后继续观察共培养肝细胞的功能变化情况.多组间比较采用单因素方差分析,两两比较采用LSD法.结果 第3代MSCs纯度>90%;肝细胞活率>95%,纯度>99%.共培养组肝细胞迅速黏附于MSCs表面,呈肝细胞岛样分布.共培养组肝细胞白蛋白分泌水平和尿素合成能力自共培养第1天起均明显高于单纯肝细胞组(P值均<0.01),并在第2天达到高峰,分别为(457.71士22.62)ng和(69.05±2.12)μg.免疫细胞化学检测结果显示共培养组MSCs表达多种ECMl RNA干扰实验进一步证实ECM的存在与共培养中肝细胞的白蛋白分泌和尿素合成功能有关.结论 骨髓MSCs通过分泌多种ECM改善共培养肝细胞的形态与功能.     

果糖诱导肝脂肪变性细胞模型建立及评价

目的建立果糖诱导L02肝细胞脂肪变性模型的方法,并对该细胞模型脂质合成进行评价。方法体外培养L02肝细胞,以不同浓度的果糖处理24 h诱导细胞脂肪变性。Cck-8法检测果糖对细胞的活性影响,检测培养液中ALT、AST含量变化以确定果糖对肝细胞的损伤。油红O染色观察胞内脂滴沉积情况,并测定胞内三酰甘油(TG)含量,确定最佳模型浓度;同时利用蛋白印迹检测碳水化合物反应元件结合蛋白(ChREBP)、固醇调节元件结合蛋白-1c(SREBP-1c)、乙酰辅酶A羧化酶1(ACC1)和硬脂酰辅酶A去饱和酶1(SCD1)蛋白表达,并与游离脂肪酸(FFA)干预相比较。结果 L02肝细胞在0~32 mmol/L果糖干预下活性无显著改变,细胞无显著损伤。果糖浓度为4 mmol/L时,L02肝细胞内即有大量脂滴形成。且细胞内TG含量显著增加,为正常对照的1.5倍。4 mmol/L果糖能显著上调ChREBP、SREBP-1和ACC1蛋白表达。相对于FFA,果糖对SCD1和ACC1蛋白表达上调更显著。结论采用4 mmol/L果糖可成功诱导L02肝细胞脂肪变性,该模型适用于高果糖饮食诱发的非酒精性脂肪性肝病脂质合成研究。     

系统性红斑狼疮患者骨髓间质干细胞对B淋巴细胞免疫抑制作用的初步研究

目的：探讨系统性红斑狼疮（SLE）患者骨髓间质干细胞（MSCs）体外对B淋巴细胞的免疫调节作用是否存在异常。方法：采用直接贴壁法分离培养正常人及SLE患者MSCs。免疫磁珠法分选正常人外周血B淋巴细胞,与MSCs共培养,流式细胞术检测B细胞凋亡及表面标志表达,3H掺入法检测B细胞增殖,酶联免疫吸附法（ELISA）检测培养上清中免疫球蛋白IgG、IgM和IgA水平。结果：①B细胞＋正常MSCs共培养组B细胞增殖（4307±308）cpm较单纯B细胞培养组（7059±346）cpm降低（P〈0.05）;②B细胞＋正常MSCs共培养组B细胞凋亡率（14±7）%低于单纯B细胞培养组（25±8）%（P〈0.05）;③B细胞＋正常MSCs共培养组B细胞表面CD86表达率（53±5）%较单纯B细胞培养组（66±10）%低（P〈0.05）;④与MSCs共培养后,B细胞分泌免疫球蛋白IgG、IgM和IgA较单纯B细胞培养显著减少（P〈0.05）;⑤B细胞＋狼疮MSCs共培养组与B细胞＋正常MSCs共培养组比较,B细胞增殖、凋亡、CD86表达和Ig分泌两组间差异无统计学意义。结论：MSCs在体外对B细胞有抑制作用,SLE患者骨髓MSCs与正常MSCs相比,对B细胞的表面刺激分子表达、增殖、Ig分泌有相似的抑制作用。     

复方甘草酸单铵和异甘草酸镁对化学损伤人肝细胞的保护作用

目的:研究2 类甘草甜素药物对半乳糖胺( D-GalN)和四氯化碳(CCl4)损伤培养人肝细胞(L-02)的保护作用及差异.方法:培养L-02, 用复方甘草酸单铵(compoundammonium glycyrrhizin, CAG)和异甘草酸镁(magnesium isoglycyrrhizinate, MI)分别进行保护后, 再经D-GalN或CCl4处理. 观察肝细胞生长状态、测定AST、LDH酶活力、及细胞内谷胱甘肽(GSH)含量. 从而评价CAG和MI对D-GalN和CCl4损伤L-02的保护作用差异.结果:浓度为1 g/L的CAG和MI均能提高细胞的存活率, 显著抑制D-GalN和CCl4所致的AST及LDH释放. CAG抑制D-GalN和CCl4所致的AST与LDH效果显著好于MI(均P<0.05);浓度为1 g/L的CAG和MI均能显著抑制2种化学损伤细胞内的GSH降低(CCl4损伤:7.59±1.27, 5.23±0.70 vs 3.33±0.40; D-GalN损伤:7.93±0.36, 5.40±0.52 vs 3.77±0.45, P<0.01或0.05), CAG效果明显好于MI.结论:1 g/L的CAG和MI 2种药物对D-GalN和CCl4致人肝细胞损伤均有保护作用, 其机制与抑制GSH降低相关. 2种甘草甜素药物相比较,CAG保护肝细胞效果好于MI.     

乙型肝炎病毒x影响肝细胞增殖与凋亡的初步研究

目的 观察乙型肝炎病毒x(HBx)导入正常肝细胞后对细胞周期及凋亡的影响,探讨HBx致肝细胞癌的发生机制.方法 应用脂质体转染重组质粒pcDNA3.1( )/v5-hisB-HBx后,G418筛选获得阳性克隆L02/HBx,分别用RT-PCR和Western blot鉴定其HBx mRNA与蛋白的表达.噻唑蓝比色法(MTT)及细胞生长曲线观察细胞增殖情况,流式细胞术分析其细胞周期特征与凋亡率.结果 构建的L02/HBx细胞中可稳定表达HBx mRNA及蛋白.与对照组比较,L02/HBx细胞增殖速度明显加快(P＜0.05);其G1期细胞比例明显减少,S期细胞比例显著增加,凋亡率明显降低(P＜0.05).结论 成功构建稳定表达HBx的转基因细胞株L02/HBx,初步显示HBx可促进肝细胞增殖,抑制细胞凋亡,从而加速细胞周期进程,可能参与肝细胞的恶性转化.     

氟对骨髓基质细胞增殖和成骨能力的影响

目的探讨氟对体外培养的大鼠骨髓基质细胞（MSCs）增殖能力与成骨能力的影响。方法体外培养大鼠MSCs。噻唑蓝（MTT）比色法分析不同染氟水平下MSCs的增殖能力；光、电镜下观察不同染氟水平下MSCs成骨转化能力和相应超微结构变化。结果与对照组比较，低水平染氟（0．02mg／L）96h后促进MSCs增殖（P〈0．05）；较高水平染氟（0．05～1．00mg／L）72h即可促进MSCs增殖（P〈0．05或〈0．01）；高水平染氟（10．00、20．00mg／L）对MSCs具有抑制增殖和促细胞凋亡作用（P〈0．01）。较高水平染氟（2．00～20．00mg／L）时MSCs成骨转化受抑（P〈0．01），较低水平染氟（0．01～1．00mg／L）时MSCs成骨转化增强（P〈0．01）。结论低水平染氟促进MSCs增殖。成骨转化能力增强；高水平染氟抑制MSCs增殖，促进细胞凋亡，抑制MSCs成骨转化能力。     

骨髓基质干细胞对H_2O_2诱导的肝细胞损伤保护作用的研究

背景:骨髓基质干细胞是一种多能干细胞,不但可分化为肝细胞,并且可通过旁分泌的方式分泌细胞因子,从而促进肝再生。目的:探讨骨髓基质干细胞对H_2O_2诱导的肝细胞损伤的保护作用。方法:以500μmol/L H_2O_2诱导人肝细胞株THLE-3损伤模型,24h后与骨髓基质干细胞共培养。倒置相差显微镜下观察细胞形态改变,检测细胞毒性、caspase-3/7活性以及YO-PRO-1阳性细胞率。结果:与骨髓基质干细胞共培养后,H_2O_2损伤的THLE-3细胞形态明显改善,细胞毒性显著减少(P0.05),caspase-3/7活性显著下降(P0.05),YO-PRO-1阳性细胞率显著降低(P0.05)。结论:骨髓基质干细胞通过降低细胞毒性、caspase-3/7活性和细胞凋亡而对H_2O_2诱导的THLE-3细胞损伤起保护作用。     

转基因细胞模型L02/HBx的构建及HBx对细胞周期的影响

目的:构建L02/HBx转基因细胞模型并研究HBx对肝细胞周期的影响.方法:运用脂质体转染和G418筛选获得L02/ HBx阳性克隆,并分别用RT-PCR和Western blot鉴定HBx mRNA与蛋白的表达.进一步用四唑蓝(MTT)比色试验、流式细胞仪检测L02/HBx的增殖、凋亡和细胞周期.结果:RT-PCR和Western blot分别检测到L02/ HBx细胞中HBx mRNA和蛋白的表达.MTT比色试验显示L02/HBx生长速度加快,流式细胞仪检测发现L02/HBx凋亡率低(0.09%±0.13% vs 3.74%±1.29%,P<0.05),G1期细胞比例减少(61.35%±0.82% vs 67.80±6.84%,P<0.05),S期细胞比例相应增加(36.59%±2.54% vs 22.37%±2.17%,P<0.05).经阿霉素(ADM)培养后,L02/HBx的凋亡率显著增加(34.91%±5.85% vs 0.09%±0.13%,P<0.05),G1期细胞比例明显增加但低于对照组(82.81%±6.48% vs 61.35%±0.82%,P<0.05;82.81%±6.48% vs 87.19%±1.92%,P<0.05),S期细胞比例降低但较对照组高(13.84%±6.16% vs 36.59%±2.54%,P<0.05;13.84%±6.16% vs 2.22%±1.26%,P<0.05).结论:L02/HBx构建成功,HBx能促进细胞周期进程,加快细胞的生长并抑制细胞的凋亡;转染HBx基因的肝细胞凋亡更易受凋亡因子所触发,表明HBx可能会增加正常肝细胞对诱导凋亡因素的敏感性.     

Macrophages and mesenchymal stromal cells support survival and proliferation of multiple myeloma cells

Multiple myeloma (MM) is characterized by almost exclusive tropism of malignant cells for the bone marrow (BM) milieu. The survival and proliferation of malignant plasma cells have been shown to rely on interactions with nonmalignant stromal cells, in particular mesenchymal stromal cells (MSCs), in the BM microenvironment. However, the BM microenvironment is composed of a diverse array of cell types. This study examined the role of macrophages, an abundant component of BM stroma, as a potential niche component that supports malignant plasma cells. We investigated the proliferation of MM tumour cell lines when cultured alone or together with MSCs, macrophages, or a combination of MSCs and macrophages, using the carboxyfluorescein succinimidyl ester assay. Consistently, we observed increased proliferation of MM cell lines in the presence of either MSCs or macrophages compared to cell line-only control. Furthermore, the combined co-culture of MSCs plus macrophages induced the greatest degree of proliferation of myeloma cells. In addition to increased proliferation, MSCs and macrophages decreased the rate of apoptosis of myeloma cells. Our in vitro studies provide evidence that highlights the role of macrophages as a key component of the BM microenvironment facilitating the growth of malignant plasma cells in MM.     

The anti-tumoral effect of PI3K inhibitor and MEK inhibitor combined with STI571 on chronic myeloid leukemia cells in a bone marrow stromal cell co-culture system

The goal of this study was to elucidate the functional roles of PI3K/AKT and MEK/ERK signaling on the proliferation and apoptosis of STI571-sensitive and -resistant CML cell lines in a co-culture system with human marrow stromal cells (MSCs), mimicking the bone marrow microenvironment. The phosphorylation of AKT and ERK was enhanced by co-culture with MSCs in both STI571-sensitive KBM-5 and STI571-resistant KBM-5/STI cells. In KBM-5 cells, the STI571 and PI3K inhibitor LY294002 combination was effective on apoptosis induction in the MSC co-culture system. In KBM-5/STI cells, treatment with LY294002 or PD98059 alone resulted in massive apoptosis, which was enhanced by co-culture with MSCs. These results provide a rationale for multi-molecular target therapy approaches based on a combination of signal transduction inhibitors with STI571 in CML.     

重组蛋白rh TRAIL55-281与GST-rh TRAIL55-281有肝细胞毒性

目的:检测重组蛋白rhTRAIL55-281与GST- rhTRAIL55-281的对肝细胞的毒性,以确定其是否可以开发为抗肿瘤药物．方法:采用胰酶消化与机械分离结合的方法, 获取原代成人肝细胞和胎肝细胞．用Factor- Xa切除重组蛋白GST-rhTRAIL55-281的GST 标签获取rhTRAIL55-281蛋白,然后分别利用 rhTRAIL55-281与GST-rhTRAIL55-281干预肝细胞株L-02、原代成人肝细胞、原代胎肝细胞及对照组正常人外周血单个核细胞(PBMC),最后利用流式细胞仪检测细胞的凋亡率．结果:获得的原代成人肝细胞和胎肝细胞细胞活力达95％以上．无GST标签的 rhTRAIL55-281蛋白纯度达到97％;rhTRAIL55-281 与GST-rhTRAIL55-281使肝细胞株L-02及原代成人肝细胞、胎肝细胞大量凋亡．在10 mL／L 的浓度下,48 h后凋亡率分别为79．1％,72．8％, 42．2％与80．3％,74．7％,47．2％,而对照组正常人PBMC基本不凋亡．结论:重组蛋白rhTRAIL 55-281与GST- rhTRAIL55-28I有肝细胞毒性．     

Exaggeration of acute liver damage by hepatic sympathetic nerves and circulating catecholamines in perfused liver of rats treated with D-galactosamine

Effects of electrical stimulation of the hepatic nerves on acute liver damage were examined using isolated rat liver perfused in situ, 24 hours after intraperitoneal injection with D-galactosamine (800 mg/kg). The leakage of lactate dehydrogenase (LDH) and aspartate aminotransferase (AST) from the liver was used as markers of acute liver damage. In perfused livers after treatment with galactosamine, nerve stimulation (20 V, 20 Hz, 2 ms) increased the leakage of LDH and AST about 3-fold over the basal level accompanied by the decrease in flow rate, whereas with control livers the leakage of LDH and AST into the effluent was almost undetectable throughout the perfusion. The rapid increase in the leakage of LDH and AST was observed during nerve stimulation even under conditions where perfusion flow was maintained constant. Such effects of hepatic nerve stimulation on galactosamine-treated livers were mimicked well by infusion of noradrenaline or phenylephrine, and inhibited by the alpha1-antagonist bunazosin. Artificial reduction of perfusion flow alone did not induce the rapid leakage of LDH and AST into the effluent. On the other hand, low concentration (10 nmol/L) of noradrenaline only minimally decreased the flow rate but apparently augmented liver cell damage. The acute liver damage augmented by noradrenaline was dependent on extracellular Ca²⁺. These results indicate that in the liver, already having been injured slightly, the activation of hepatic sympathetic nerves and circulating catecholamines exaggerates acute liver damage through an action on liver cells, which depends on the influx of extracellular Ca²⁺. (Hepatology 1996 Mar;23(3):524-9)     

刚地弓形虫细胞培养上清对人结肠癌细胞sw480增殖与凋亡的影响

目的观察刚地弓形虫RH株速殖子与人结肠癌sw480细胞系共培养上清对人结肠癌sw480细胞增殖的影响和诱导其凋亡与坏死的情况。方法取对数生长期的人结肠癌sw480细胞(1×106),建立弓形虫RH株速殖子数目分别为2×106、4×106、8×106、16×106的共培养模型,观察弓形虫速殖子在sw480细胞中的寄生,提取共同孵育72h后的培养上清,以新鲜培养基稀释为半量,体外作用人结肠癌sw480细胞12h、24h、48h、72h,CKK-8法检测吸光度(A450值)并计算抑制率;Annexin-v-FITC/PI染色细胞后上流式细胞仪检测细胞凋亡与坏死率;琼脂糖凝胶电泳观察细胞凋亡DNA条带;透射电镜和荧光显微镜观察细胞形态学改变。结果弓形虫RH株速殖子可在人结肠癌sw480细胞内寄生和增殖。CKK-8法检测结果显示上述共培养上清对sw480细胞增殖抑制率随上清作用时间延长均明显增大,48h最大抑制率达44.55%。流式细胞仪检测显示实验组sw480细胞早期凋亡率和晚期凋亡与坏死率随上清作用时间延长均明显增加,48h早期凋亡率达最高值(11.54%),48h以后早期凋亡率下降,晚期凋亡和坏死率显著上升,72h总死亡率可达46.11%,细胞杀伤效果明显;琼脂糖凝胶电泳显示典型的DNA云梯状条带;荧光显微镜和透射电镜观察到细胞凋亡和坏死典型形态。结论弓形虫RH株速殖子与人结肠癌sw480细胞共培养上清对体外培养的sw480细胞增殖有明显的抑制作用,并可诱导sw480细胞凋亡与坏死。     

Ethanol effect on cell proliferation in the human hepatoma HepaRG cell line: relationship with iron metabolism

Background: Alcoholism increases the risk of cirrhosis and/or hepatocellular carcinoma development. Iron, like ethanol, modulates the cell growth. However, the relationship between alcohol and iron toward hepatocyte proliferation has not been clearly elucidated. The purpose of this study was to evaluate, in the human HepaRG cell line model, the impact of ethanol on hepatocyte proliferation in relation to modulations of iron metabolism and the protective effect of iron metabolism manipulation by chelators in alcohol liver diseases. Methods: The human hepatoma HepaRG cell line model was used. Cell viability was determined by measuring succinate dehydrogenase activity, total protein level by the Bradford method. DNA synthesis was evaluated by [³H]‐methyl thymidine incorporation. Cytotoxicity was studied by release of lactate dehydrogenase (LDH), aspartate aminotransferase (AST), alanine aminotransferase (ALT) in culture medium and apoptosis by measuring caspase 3/7 activity. Gene expression was analyzed by RT‐qPCR. Total iron, soluble transferrin receptor, and ferritin levels were, respectively, measured by colorimetrical, immuno‐nephelometrical, and immuno‐turbidimetrical methods. Intracellular iron uptake and accumulation was examined by radionuclide ⁵⁵Fe (III) measurement and Perls staining. Results: Results showed that ethanol decreased all the parameters associated with HepaRG cell proliferation (cell viability, total protein levels, and DNA synthesis) in a dose‐ and time‐dependent manner. This effect was accompanied by cytotoxicity and apoptosis as evaluated by a significant increase in extracellular enzymes (LDH, AST, ALT) and caspase 3/7 activity, respectively. Ethanol exposure was accompanied by an increased cellular iron uptake, together with increased expression of genes involved in iron transport and storage such as l ‐ferritin, Divalent Metal transporter 1, transferrin, transferrin receptor 1, and ceruloplasmin. Ethanol impact was intensified by iron‐citrate and decreased by iron chelators when added to the culture medium. Conclusions: The results indicated that (i) ethanol‐induced iron metabolism dysfunction could be one of the underlying mechanisms of ethanol antiproliferative effect and (ii) exogenous iron may accentuate ethanol hepatoxicity. These data suggest that iron metabolism manipulation by chelators may be a useful therapeutic approach in alcohol‐related liver diseases.     

Staurosporine诱导小鼠心肌细胞凋亡模型的构建

目的 :Staurosporine可诱导不同细胞类型的细胞凋亡 ,但尚缺乏其对小鼠心肌细胞作用影响的报道。本研究旨在观察 Staurosporine是否也可引起小鼠心肌细胞的凋亡 ,并对其模型进行分子水平的鉴定。方法 :用 Stau-rosporine处理原代培养的小鼠心肌细胞的直接刺激后 ,观察心肌细胞的形态学变化、DNA片段、半胱天冬蛋白酶(caspase) -3活性、细胞存活率以及细胞膜完整性。结果 :1Staurosporine处理过的心肌细胞具有明显的凋亡形态学特征 :细胞的皱缩、核的浓缩及 DNA梯改变。2 Staurosporine4μmol/ L 对心肌细胞作用 8h后 ,与正常对照组比较细胞存活率降至 3 1.2 % (与对照组比 ,P<0 .0 1) ,且细胞存活率与 Staurosporine的作用浓度和作用时间呈依赖关系 ;同时测定细胞培养液中能反映细胞膜完整性的胞浆内容物乳酸脱氢酶 (L DH)水平 ,发现在 Staurosporine作用 1～ 8h的各个时间点 ,L DH的漏出水平最高未超过 10 % (与对照组相比 ,P>0 .0 5) ;这种高的细胞死亡率与低的细胞膜的损伤率 ,恰恰反映了细胞死亡的性质是凋亡性死亡。3 Staurosporine能激活心肌细胞内的 Caspese-3的活性 ,当使用广谱的 Caspese抑制剂 ZVAD-fmk预处理培养的小鼠心肌细胞后 ,能够有效的阻止上述细胞凋亡的发生 ,从而避免了 Staurospor     

Effect of co-culture of mesenchymal stem/stromal cells with pancreatic islets on viability and function outcomes: a systematic review and meta-analysis

The maintenance of viable and functional pancreatic islets is crucial for successful islet transplantation from brain-dead donors. To overcome islet quality loss during culture, some studies have co-cultured islets with mesenchymal stem/stromal cells (MSC). However, it is still uncertain if MSC-secreted factors are enough to improve islet quality or if a physical contact between MSCs and islets is needed. Therefore, we performed a systematic review and meta-analysis to clarify the effect of different culture contact systems of islets with MSCs on viability and insulin secretion outcomes. Pubmed and Embase were searched. Twenty studies fulfilled the eligibility criteria and were included in the qualitative synthesis and/or meta-analysis. For both outcomes, pooled weighted mean differences (WMD) between islet cultured alone (control group) and the co-culture condition were calculated. Viability mean was higher in islets co-cultured with MSCs compared with islet cultured alone [WMD = 18.08 (95% CI 12.59–23.57)]. The improvement in viability was higher in islets co-cultured in indirect or mixed contact with MSCs than in direct physical contact (P <0.001). Moreover, the mean of insulin stimulation index (ISI) was higher in islets from co-culture condition compared with islet cultured alone [WMD = 0.83 (95% CI 0.54–1.13)], independently of contact system. Results from the studies that were analyzed only qualitatively are in accordance with meta-analysis data. Co-culture of islets with MSCs has the potential for protecting islets from injury during culture period. Moreover, culture time appears to influence the beneficial effect of different methods of co-culture on viability and function of islets.     

去唾液酸糖蛋白-人端粒酶逆转录酶-胸苷激酶/丙氧鸟苷的靶向促HepG2细胞凋亡作用及旁观者效应

目的 观察去唾液酸糖蛋白(AF)-pGL3-人端粒酶逆转录酶(hTERT)-胸苷激酶(TK)对肝癌细胞株HepG2细胞生长及凋亡的影响.方法 培养细胞并构建pGL3-hTERT-TK质粒、AF脂质体复合物后,转染HepG2细胞和L02细胞,通过单光子液闪计数仪,脱氧核糖核苷酸末端转移酶介导的缺口末端标记法和流式细胞仪观察自杀基因对肝癌细胞生长和凋亡的影响,以及其对自杀基因的旁观者效应.结果 在肝癌细胞HepG2中,TK基因可以被hTERT启动子驱动高效的表达,而不影响正常肝细胞L02的生长,AF通过识别去唾液酸糖蛋白受体结合到HepG2细胞表面,其携带的TK基因更易进入肝癌细胞,同时增强自杀基因TK的高效表达,在旁观者效应机制的参与下,肝癌细胞总的凋亡率达85%±3%,而正常肝细胞则仅为16%±2%.结论 AF-pGL3-hTERT-TK可以靶向攻击肝癌细胞,对正常肝细胞几乎无影响,其基因传递系统具备靶向治疗肝癌的潜力.     

戊型肝炎病毒ORF3融合蛋白对L02细胞增殖和凋亡的影响

目的检测ORF3融合蛋白对L02细胞增殖与凋亡的影响。方法将重组质粒ORF3-pEGFP和ORF3-pXF2RH脂质体法转染L02细胞,用MTT比色法测定细胞增殖情况,用流式细胞仪碘化丙啶染色法检测细胞周期和细胞凋亡情况。结果转染重组质粒的L02细胞自第4d开始,明显增殖(P<0.05);L02细胞凋亡率由转染前的1.41%分别下降为0.83%和0.88%,差异有统计学意义(P<0.05)。结论戊型肝炎ORF3融合蛋白可减少L02细胞凋亡,而使L02增殖速度增加(P<0.05)。