Tanshinone IIA prevents high glucose-induced human umbilical vein endothelial cell apoptosis

AIM: To investigate the effect of tanshinone IIA on the apoptosis of human umbilical vein endothelial cells(HUVECs) after high glucose treatment.METHODS: The cell viability was determined by MTT assay. The cell apoptotic rate was examined by flow cytometry with Annexin V-FITC/PI double staining. The expression of Bcl-2 and Bax, and the release of mitochondrial cytochrome C(Cyt C) were analyzed by Western blotting.RESULTS: Tanshinone IIA significantly inhibited high glucose-induced decrease in cell viability and increased the cell apoptosis. Additionally, after tanshinone IIA treatment, Bax expression and the release of mitochondrial Cyt C were significantly inhibited, while Bcl-2 expression was increased.CONCLUSION: Tanshinone IIA prevents high glucose-induced endothelial cell apoptosis via mitochondria-dependent pathway.     

scRNA-seq of ovarian follicle granulosa cells from different fertility goats reveals distinct expression patterns

The new technology of high-throughput single-cell RNA sequencing (10 × scRNA-seq) was developed recently with many advantages. However, it was not commonly used in farm animal research. There are few reports for the gene expression of goat ovarian follicle granulosa cells (GCs) during different developmental stages. In the current investigation, the gene expression of follicle GCs at different stages from two populations of Ji'ning grey goats: high litter size (HL; ≥3/L; 2 L) and low litter size (LL; ≤2 /L; 2 L) were analysed by scRNA-seq. Many GC marker genes were identified, and the pseudo-time showed that GCs developed during the time course which reflected the follicular development and differentiation trajectory. Moreover, the gene expression difference between the two populations HL versus LL was very clear at different developmental stages. Many marker genes differentially expressed at different developmental stages. ASIP and ASPN were found to be highly expressed in the early stage of GCs, INHA, INHBA, MFGE8 and HSD17B1 were identified to be highly expressed in the growing stage of GCs, while IGFBP2, IGFBP5 and CYP11A1 were found to be highly expressed in late stage. These marker genes could be used as reference genes of goat follicle GC development. This investigation for the first time discovered the gene expression patterns in goat follicle GCs in high- or low-fertility populations (based on litter size) by scRNA-seq which may be useful for uncovering the oocyte development potential.     

VEGF对绵羊卵泡颗粒细胞FSHR、E2R表达的影响

血管内皮生长因子(VEGF)可以促进颗粒细胞的增殖,但是否影响同样分布于颗粒细胞上的FSHR、E2R表达效果尚属未知.因此,通过Western blotting和荧光定量PCR分别对添加0、5、10、15、20、25 μg/L VEGF卵泡颗粒细胞中FSHR、E2R表达量及FSHR mRNA、E2R mRNA表达量进行检测.Western blotting蛋白检测结果表明,空白对照组和各个不同质量浓度VEGF处理组的颗粒细胞上均有FSHR和E2R蛋白表达,FSHR相对分子质量为75 000,E2R相对分子质量为56 000；荧光定量PCR检测结果表明,添加VEGF的各个处理组中FSHR mR-NA、E2R mRNA表达量均显著高于对照组(P＜0.05),各处理组间的FSHR mRNA之间差异不显著(P＞0.05),而VEGF添加量20、25 μg/L组的E2R mRNA表达量显著高于其他3个处理组(P＜0.05),并且这2个组间则差异不显著(P＞0.05),其余3个处理组间亦差异不显著(P＞0.05).     

加味柴胡汤对子宫内膜炎奶牛血液白细胞的影响

本试验选子宫内膜炎患牛10头,随机分为中药治疗组和患病对照组（每组5头）,5头健康奶牛作为正常对照组.中药治疗组子宫灌注复方中药剂加味柴胡汤,连用6d,其他两组给予生理盐水.在用药前1d和用药2d、4d、6d采血,用血液自动分析仪检测.结果：（1）治疗前,中药治疗组和患病对照组白细胞数量显著（P＜0.05）高于正常对照组,治疗6d后,中药治疗组显著（P＜0.05）低于治疗前和患病对照纽.（2）治疗前,中药治疗组和患病对照组中间细胞数量和比率均显著（P＜0.05）高于正常对照组,用药6d后,中药治疗组显著（P＜0.05）或极显著（P＜0.01）低于患病对照组和治疗前.（3）治疗前,中药治疗组和患病对照组嗜中性粒细胞数量比率显著（p＜0.05）高于正常对照组.用药6d后,中药治疗组与正常对照组均显著（P＜0.05）低于患病对照组,低于治疗前.（4）治疗前,中药治疗组和患病对照组淋巴细胞数量显著（P＜0.05）高于正常对照组,比率显著（P＜0.05）低于正常对照组.用药6d后,中药治疗组和正常对照组淋巴细胞数量和比率显著（P＜0.05）高于患病对照纽.结论：加味柴胡汤可降低子宫内膜炎奶牛的炎症反应,提高免疫力.     

猪骨髓间充质干细胞分离培养及生物学特性

采集健康猪股骨骨髓,利用percoll梯度离心法分离单个核细胞,进行体外原代和传代培养,并用不同代数细胞进行了细胞生长能力、膜表面抗原(CD105、CD90、CD45)及诱导分化能力的检测.结果显示分离培养的单个核细胞贴壁生长,形态呈成纤维细胞样及涡旋状克隆团；细胞传代至第17代生长状况仍然良好；用F3代细胞进行膜表面抗原标记显示,CD90和CD105阳性表达率分别为(97.4±1.8)％和(99.6±0.9)％,而CD45阳性表达率仅为(1.8±0.55)％,证实这些细胞具有猪骨髓间充质干细胞(Mesenchymal stem Cells,MSCs)表面抗原；经地塞米松、维生素C和β-磷酸甘油等诱导F3代细胞,21d后出现钙物质沉积细胞群,用茜素红和Von kossa染色呈阳性,证实这些细胞已分化形成成骨细胞；经地塞米松、IBMX、胰岛素及吲哚美辛等诱导F3代细胞,21d后细胞质出现脂滴样结构,用油红O染色呈阳性,证实已分化形成成脂细胞；冷冻-解冻细胞的膜表面抗原及多能分化能力与未冻存细胞的检测结果基本一致.结果证实,从猪骨髓中分离培养及经传代扩增获得的贴壁细胞是纯化的MSCs.     

小鼠小肠上皮细胞的分离培养研究

通过组织块培养法进行胎鼠原代细胞培养,采用刮除法和相差消化及相差贴壁法对细胞进行纯化,用0.05%的胰蛋白酶进行消化传代对小鼠小肠上皮细胞进行了原代分离培养、传代、纯化及鉴定。结果表明:组织块培养法得到了小鼠生长状况良好的细胞,经过纯化,得到了较纯的小肠上皮细胞;形态学观察和免疫组化检测显示得到的细胞为小鼠上皮细胞。用组织块培养可以分离纯化出增殖能力较强的上皮细胞,并能进行传代纯化,通过免疫组化鉴定,证明分离出的小鼠IEC为阳性,即为小肠上皮细胞。     

四氯联苯对鸡胚生殖新月原始生殖细胞分布的影响

为了探索四氯联苯(2,2′,5,5′)对鸡胚生殖新月原始生殖细胞(PGCs)分布的影响,本试验将四氯联苯注入种蛋的胚盘(第X期)处,在38℃,相对湿度60%,每2h45度转蛋的条件下孵化至原条期,通过不同方法检测原条期生殖新月明区、暗区及血液循环中的PGCs,探讨四氯联苯对胚盘生殖新月区PGCs分布的影响。结果表明,四氯联苯明显降低了原条期明区和血液中PGSs的数量(P<0.01),但对暗区PGCs的数量影响不明显。这说明四氯联苯对血液中PGCs的影响是由于降低了生殖新月明区PGCs的缘故,和暗区的PGCs无相关性。     

猪前体脂肪细胞与肌卫星细胞体外联合培养

利用差速贴壁法分别纯化前体脂肪细胞与肌卫星细胞,按1∶10的浓度比接种混合培养。以单独两类细胞培养作对照,采用油红O染色和Desmin免疫组化染色鉴定,通过形态学观察和MTT比色绘制生长曲线研究细胞生长规律。结果表明,Desmin免疫组化染色观察肌细胞阳性率达97%以上,油红O染色计数诱导分化细胞比率大于60%;形态学观察发现,联合培养细胞接触汇合前形态呈梭形,接触后不规则重叠交织生长;充脂细胞呈现较晚,且数量稀少;HE染色可见多核细胞融合成肌管样结构;测定2～9d细胞生长曲线显示,联合细胞组增殖比率大于对照组,经历2～3d潜伏期、3～7d指数生长期后,并未与对照组同步进入平台期,而呈持续缓慢上升趋势。实验初步实现了体外猪前体脂肪细胞与肌卫星细胞的直接联合培养,并显示其与对照组生长特性存在差异。