Intrathymic inoculation of liver specific antigen protects hepatocytes from apoptosis after liver allotransplantation

AIM: To study the effects of intrathymic inoculation of liver specific antigen (LSA) on hepatocyte apoptosis after liver allotransplantation. METHODS: Orthotopic liver transplantation was used in this study. Group Ⅰ: syngenic control (Wistar-to-Wistar); Group Ⅱ: acute rejection (SD-to-Wistar); Group Ⅲ: thymus inoculation of SD rat LSA day 7 before transplantation. The observation of general situation and survival time, hepatocyte apoptosis and LAT expression in liver transplants were used to analyze immune state of animals in different groups. RESULTS: The general situation of group Ⅰ was very well after transplantation. Recipients of groupⅡ lost body weight progressively and all died within day 9 to day 13 post transplantation. As for group Ⅲ, the general situation of recipients was remarkably better than that in group Ⅱ. The positive cells of apoptosis in group Ⅲ detected by TUNEL were not significantly different from that in group Ⅰ, but was significantly lower than that in group Ⅱ. LAT was detected at any time in group Ⅱ with peak expression at day 5 and day 7 post transplantation. In contrast, LAT was not detected in any other groups. CONCLUSION: Intrathymic inoculation of LSA protects hepatocytes from apoptosis after liver allotransplantation.     

嗅鞘细胞对无血清培养诱导PC12细胞凋亡的作用

采用无血清培养诱导PC12细胞凋亡,研究嗅鞘细胞(Olfactory ensheathing cells,OECs)对去血清后诱导PC12细胞凋亡中的作用。通过MTT法检测细胞的活性,细胞形态学观察,以及结合流式细胞仪检测细胞的凋亡率,井对不同组细胞凋亡率进行比较。结果表明：①OECs培养上清对去血清后PC12细胞的存活有明显的促进作用;②从形态学上观察,OECs对去血清诱导PC12细胞凋亡有明显的抑制作用,使得细胞向死亡过渡受阻;②流式细胞对细胞周期分析,去血清诱导后的PC12细胞,大部分细胞滞留在G1期,细胞向S期过渡受阻,造成G1期细胞的堆积,它们所占比例分别为：73．6％,25．9％,0．5％;而OECs联合培养组所占比例：53％,42．3％,4．7％;OECs上清组：42．1％,52．2％,5．7％;正常有血清培养组：45,6％,41．4％,13％。凋亡细胞所占百分比分别为：60．3％,45,6％,37．4％,0．5％。     

二乙基己烯雌酚对成年仓鼠睾丸一氧化氮生成和生精细胞凋亡的影响

探讨了在二乙基己烯雌酚（DES）诱发成年动物生精细胞凋亡过程中睾丸一氧化氮（NO）生成和生精细胞一氧化氮合酶（eNOS和iNOs）表达的变化，以期为阐明DES诱发生精细胞凋亡机理的研究提供基础资料。成年雄性仓鼠皮下注射不同剂量DES（分别为0．01、0．1和1mg／kg体重），连续注射7d后取其睾丸，进行NO含量的测定和eNOS、iNOS免疫组化染色。电镜观察生精细胞超微结构的变化，并用TUNEL法检测睾丸中生精细胞凋亡的变化，苏丹Ⅲ染色法检测睾丸生精小管内脂滴分布的变化。结果显示：NO的生成与DES呈剂量依赖性。DES处理后，在1mg／kg体重剂量组，大量生精细胞表达eNOS和iNOS，并出现大量凋亡，退化的生精细胞胞浆内有大量髓样结构，并有大量脂滴分布于生精细胞内和细胞间。eNOS和iNOS阳性生精细胞与凋亡的生精细胞数量和类型基本一致，主要为精母细胞和圆形精子细胞。     

细胞色素C在生物医学方面的研究进展

综合论述了细胞色素C的研究简史和结构性质,探讨了细胞色素C与细胞线粒体呼吸链、生物进化细胞凋亡的关系,并对细胞色素C的应用前景作了展望。     

桦褐孔菌对肺癌细胞抗增殖和诱导凋亡观察

用MTT法测定桦褐孔菌抗突变活性成分Inotodiol,对人肺癌A549细胞株的增殖抑制作用,20mg.L-1～80mg·L^-1桦褐孔菌的抗突变活性成分Inotodiol对人肺癌A549细胞株有抑制增殖作用,且表现出浓度和时间依赖关系。在倒置显微镜下、HE染色光学显微镜下、AO/EB双染色荧光显微镜下、透射电子显微镜下观察,肿瘤细胞均有体积变小、有凋亡小体。桦褐孔菌的抗突变活性成分Inotodiol可抑制人肺癌A549细胞株的增殖和诱导肿瘤细胞凋亡。     

猪圆环病毒2型感染细胞凋亡研究进展

细胞凋亡是多种因素调节的细胞主动死亡过程,细胞凋亡的两条主要通路分别是死亡受体介导的凋亡途径和线粒体凋亡途径,但它们共同的凋亡效应物为胱冬酶;理化因素如放射线、紫外线和高温等及生物学因素如基因表达产物、酶和细胞因子等都影响细胞的凋亡。肿瘤坏死因子和胱冬酶在猪圆环病毒2型感染细胞凋亡中起着非常重要的作用;p53诱导环-H2J、un氨基端激酶,NF-κB等均参与细胞凋亡。生产中,可采用猪圆环病毒2型疫苗免疫,干扰RNA的使用和减少应激等措施降低PCV-2感染,抑制细胞凋亡。     

玉米赤霉烯酮对小鼠脾淋巴细胞凋亡的影响

采用MTT法、DNA ladder分析、流式细胞仪分析等方法,离体研究玉米赤霉烯酮对小鼠脾淋巴细胞增殖与凋亡的影响,结果发现:ZEA对离体培养LPS活化小鼠脾淋巴细胞增殖具有显著抑制作用(P<0.05),且抑制强度与其剂量和处理时间均呈依赖性关系;小鼠脾淋巴细胞经ZEA处理后,出现DNA断裂等典型细胞凋亡特征;ZEA对离体培养LPS活化小鼠脾淋巴细胞具有显著促凋亡作用(P<0.0 5),且促进强度与其剂量呈依赖性关系。这些结果表明,玉米赤霉烯酮对小鼠脾淋巴细胞有直接毒害作用。     

Study on Proliferation and Apoptosis of EBL Cells Induced by P48 Protein of Mycoplasma bovis

The aim of this study was to investigate the effect of P48 protein on proliferation and apoptosis of embryonic bovine lung (EBL) cells.In this study,samples which co-incubated with P48 protein and EBL cells in different concentrations at different time points were collected,the proliferation rate of the cells was detected by MTT method,and the changes in the nuclear morphology of EBL cells were observed by DAPI staining method.Meanwhile,flow cytometry was used to detect the apoptosis rate of EBL cells induced by P48 protein.Real-time quantitative PCR was used to detect the changes in mRNA level of apoptotic marker genes,and Western blotting tested the Bax and Beclin-1 protein expressions.The results showed that under the condition of 72 h and 10 μg/mL of protein concentration treatment,P48 extremely significantly inhibited EBL cells proliferation (P<0.01),while 0.1 and 0.5 μg/mL protein concentration had no inhibitory effect (P>0.05).The nuclear morphology showed no significant change after protein induction for 12 h,but wrinkled and condensed at 24 h.The nucleus was fragmented,and a sprouted apoptotic body was appeared at 48 and 72 h.Apoptosis related genes expression showed no obvious increase at 2 and 12 h at mRNA level,but gradually increased at 24,48 and 72 h,and it showed a time-dependent manner.Accordingly,the expression of apoptosis marker proteins Bax and Beclin-1 significantly increased.Flow cytometry analysis showed that the apoptosis rate of EBL cells induced by P48 protein was 48.44%.In conclusion,P48 recombinant protein of Mycoplasmas bovis inhibited the proliferation of EBL cells and promoted their apoptosis,which provided reference for revealing the pathogenic mechanism of Mycoplasmas bovis.     

日粮添加酵母硒对鸡肉线粒体氧化损伤及细胞凋亡的影响

旨在研究日粮中添加酵母硒对白羽肉鸡宰后成熟过程中线粒体氧化损伤及细胞凋亡的影响。以饲喂不同酵母硒添加量日粮肉鸡的胸肉为研究对象,测定宰后成熟过程中鸡肉线粒体氧化损伤程度、抗氧化酶活性、Caspase-3和Caspase-9活性及肉嫩度的变化。结果表明,宰后成熟过程中,酵母硒饲喂组活性氧（reactive oxygen species,ROS）水平、线粒体膜通透性转换孔（mitochondrial permeability transition pore,MPTP）开放程度、线粒体膜通透性、Caspase-3以及Caspase-9活性均显著低于对照组（P<0.05）;硒含量、胞浆细胞色素c （cytochrome c,Cyt-c）还原水平、线粒体膜电位及谷胱甘肽过氧化物酶（glutathione peroxidase,GSH-Px）活性显著高于对照组（P<0.05）。酵母硒饲喂组总抗氧化能力（total antioxidant capacity,T-AOC）在宰后成熟初期显著高于对照组（P<0.05）,而在其他时间点虽有提高但差异不显著（P>0.05）。此外,酵母硒饲喂组肌原纤维小片化指数（myofibrillar fragmentation index,MFI）在宰后成熟过程中显著低于对照组,相对应的剪切力显著高于对照组（P<0.05）。由此可见,饲喂酵母硒通过降低鸡肉线粒体的氧化损伤进而抑制了细胞凋亡,最终减弱了肌原纤维蛋白的降解。     

鸡软骨细胞体外分离培养鉴定以及过表达miR-15a对软骨细胞的影响

为分析miR-15a在肉鸡不同组织中的表达情况,并探究过表达miR-15a对体外培养鸡软骨细胞的影响。本研究首先通过倒置显微镜观察、PCR、凝胶电泳和甲苯胺蓝染色对体外分离培养的软骨细胞进行鉴定。通过实时荧光定量PCR检测miR-15a在肢体内外翻畸形(valgus-varus deformity, VVD)组和健康组肉鸡中(各3只)各组织的表达量。之后通过CCK8和EDU方法分析软骨细胞过表达miR-15a后细胞增殖情况。软骨细胞转染miR-15a mimics后,通过qPCR检测软骨细胞的标志基因Collagen-2、Aggrecan、Collagen-10,成熟分化基因Runx2、Sox9、VEGF、MMP9,炎性因子IL-1β、IL-6、IL-8、IL-10、TNF-α、TGF-β3以及凋亡基因Fas、FasL、Bcl-2的表达量。并构建FKBP5 3'UTR的野生型载体和突变型载体,通过双荧光素酶检测报告检测miR-15a与FKBP5的靶向关系。结果表明,本研究所用的胰蛋白酶、Ⅱ型胶原酶和透明质酸酶联合消化法成功分离得到状态良好的软骨细胞。荧光定量结果显示,miR-15a在各组织中均有表达,与健康组相比,miR-15a在VVD组的肝(P < 0.01)、脾(P < 0.05)、胸腺(P < 0.01)中的表达量显著升高,在心和胸肌组织中的表达量显著降低(P < 0.01)。CCK8与EDU分析结果显示,与NC组相比,过表达miR-15a组软骨细胞增殖速度显著降低(P < 0.01),增殖细胞数量显著减少(P < 0.01)。qPCR结果显示,与mimics NC组相比,miR-15a mimics组的软骨细胞标志基因Aggrecan、成熟分化基因Sox9、Runx2表达量显著降低(P < 0.05),Fas基因表达量极显著上升(P < 0.01),FasL基因和抗凋亡基因Bcl-2极显著下降(P < 0.01)。成功构建了FKBP5 3'UTR野生型和突变型载体,双荧光素酶检测报告结果显示预测的靶基因FKBP5与miR-15a没有靶向关系。本研究成功分离并鉴定了鸡软骨细胞,过表达miR-15a抑制鸡软骨细胞增殖、成熟和分化并促进细胞凋亡。