索拉非尼对敏感人肝癌细胞株MAPK信号通路基因表达的影响

 目的：筛选对索拉非尼敏感的人肝癌细胞株,检测索拉非尼作用于敏感细胞株后MAPK信号通路基因表达谱的变化。方法：索拉非尼作用于人肝癌细胞株Huh7、 MHCC97H、 HepG2 、SMCC7721、 HepG2.2.15和PLC,流式细胞术检测肝癌细胞凋亡,CCK-8测定细胞增殖的抑制率,筛选敏感肝癌细胞株;采用实时定量PCR基因芯片,观察索拉非尼作用于敏感细胞株前后MAPK信号通路基因的表达。结果：索拉非尼作用后,流式细胞术的结果显示,凋亡较明显的细胞株为PLC和HepG2.2.15,相对不明显的细胞株为SMCC7721和MHCC97H;CCK-8测定索拉非尼对PLC、HepG2.2.15、Huh7、HepG2、MHCC97H和SMCC7721细胞的IC50值分别为5.25 μmol/L、5.30 μmol/L、6.80 μmol/L、7.01 μmol/L、11.7 μmol/L和15.0 μmol/L。对索拉非尼相对敏感细胞株和不敏感细胞株分别为PLC和SMCC7721。索拉非尼作用敏感细胞株PLC后,MAPK信号通路表达 >2.0倍的有2个,≤0.5倍的有6个。结论：PLC为对索拉非尼相对敏感的人肝癌细胞株;索拉非尼主要导致MAPK信号通路中与调控细胞周期和活化转录因子相关的基因表达发生变化。     

人胎脑动脉发育相关基因表达谱的研究

目的:比较人胎儿脑动脉与成人脑动脉基因表达谱的差异，筛选与人胎脑动脉发育相关的差异表达基因。 方法:收集3例意外流产的胎儿（胎龄18-20周）及3例成人Willis脑动脉环及主要分支，抽提组织中的总RNA，制备成生物素标记的cRNA探针后分别与Affymetrix U133A基因芯片杂交，扫描杂交信号并用专用软件分析杂交结果，筛选出在两种组织中差异表达的基因。随机选择3个差异表达基因，用荧光定量RT-PCR验证。 结果:在所检测的总共 22 215 个基因中，胎儿脑动脉与成人脑动脉相比，表达水平相差2倍、3倍、4倍、5倍以上的基因分别有935个、302个、177个、110个；在表达水平相差5倍以上的基因中，32个基因在胎儿脑动脉中高表达，78个基因在胎儿脑动脉中低表达。基因功能分类显示：25个基因与细胞信号转导和通讯有关，16个基因与细胞外基质和细胞骨架有关，与细胞防御反应有关的基因有14个，转录因子10个，另外有45个为未分类或功能未知的基因或表达序列标签（EST）片段。 结论:人胎脑动脉的正常发育受到众多基因的调控，研究这些基因的功能可能有助于认清脑动脉瘤等脑动脉发育相关性疾病的发病机制。     

HLA-DR expression in a human colonic carcinoma cell line

A human colonic carcinoma cell line (HCA-7) isolated from a well differentiated mucoid adenocarcinoma of the colon has been maintained in vitro for 3 years. It spontaneously synthesizes HLA-DR which is mainly intracytoplasmic. Stimulation with lymphocyte conditioned medium and recombinant gamma-interferon results in enhanced synthesis of HLA-DR and the appearance of the antigen on the cell surface. A dose response study showed that maximal stimulation of the culture was achieved with 50 units/ml of recombinant gamma-interferon. Staining for HLA-DR was uneven being confined to focal areas of the monolayer, which is similar to the focal expression of HLA-DR seen in sections of adenocarcinoma of the colon. The functional significance of this phenomenon is unclear, but it may explain the presence of lymphoid infiltrates in tumours.     

Polypeptide profile of HBSAg excreted by a human hepatoma cell line.

In this study the polypeptide composition of HB_sAg particles secreted by a human hepatoma cell line (Alexander) was analysed. Five major components were identified with molecular weights of 22,000, 28,000, 33,000, 38,000, and 49,000. The two smaller species (MW 22,000 and 28,000) were found to be the major components. The polypeptide profile is similar to that described for HB_sAg derived from sera of acute hepatitis patients and of chronic carriers. Data are also presented suggesting that the various HB_sAg polypeptide components are aggregates and glycosylated derivatives of the 22,000 MW polypeptide subunit.     

Neuromelanin inhibits CXCL10 expression in human astroglial cells

Increasing evidence indicates neuroinflammation is instrumental in the pathogenesis of Parkinson's disease (PD). In PD, there is selective degeneration of neuromelanin (NM)-containing dopamine neurons. Neuromelanin is predominantly cytoprotective within dopaminergic neurons, whereas, NM released from damaged neurons activates microglia. However, the effects of NM on astroglial cells remain largely unknown. Astroglia are essential to neuronal homeostasis and responsive to injury, in part, through secretion of chemokines, including interferon γ inducible protein-10 (CXCL10). Thus, we used an in vitro approach to identify the effects of NM on TNFα-induced CXCL10 expression in human astroglial cells. TNFα-induced CXCL10 expression was inhibited in NM exposed cells. Additionally, TNFα-induced NF-кB activation was inhibited by NM. Given that CXCL10 expression is NF-кB-dependent in human astroglial cells, these findings suggest that NM may inhibit CXCL10 expression, in part, through an NF-кB-dependent mechanism. While the in vivo consequences of NM mediated effects on astroglial CXCL10 expression remain to be fully elucidated, insights obtained in this study further our understanding of the effects of NM on inflammatory signaling in human astroglial cells.     

柯萨奇病毒B组5型感染的人恶性胚胎横纹肌肉瘤细胞系lncRNA表达谱分析

目的探索柯萨奇病毒B组5型(CVB5)感染人恶性胚胎横纹肌肉瘤细胞系(RD)中差异长链非编码RNA(lncRNA)表达谱,为研究CVB5与宿主之间相互作用分子机制提供参考。方法 CVB5按感染复数(MOI)为1接种RD细胞24 h后提取总RNA。采用转录组测序技术获取细胞中lncRNA差异表达谱;通过聚类分析、GO分析和KEGG通路富集对差异表达转录本进行了生物信息学分析。同时,利用RNAfold软件对lncRNA进行了二级结构预测。结果与对照组相比,CVB5感染RD细胞后,共有1 754个mRNAs和508个lncRNA呈上调表达,3 106个mRNAs和760个lncRNA呈下调表达。差异表达lncRNA的共表达基因主要富集在分子结构活性、蛋白质分子结合和体液免疫反应等生物过程;lncRNA靶基因主要参与嗅觉传导途径、细胞因子受体相互作用和神经活性配体受体相互作用等通路。此外,实时荧光定量PCR验证的7个差异表达lncRNA与测序结果一致。结论 CVB5感染RD细胞后,差异显著性lncRNA主要参与了免疫相关过程,为充分理解lncRNA在CVB5感染中的调控作用奠定了基础。     

Stable expression of a Norwalk virus RNA replicon in a human hepatoma cell line

Norwalk virus (NV) is a prototype strain of the genus Norovirus in the family Caliciviridae. The human noroviruses have emerged as major agents of acute gastroenteritis in all age groups, but there are no vaccines or antiviral agents partly due to the absence of a cell culture system. We report the generation of cells expressing self-replicating NV RNA (NV replicon) following transfection of NV RNA bearing an engineered neomycin resistance gene into cell lines of human (Huh-7) or hamster (BHK21) origin. Expression of replicon RNA was significantly reduced in the presence of interferon (IFN)-alpha in a dose-dependent manner in the NV replicon-bearing cells, suggesting a role for innate immunity in the control of human norovirus replication. This stable NV replicon system should lead to new insights into norovirus replication, virus-host interactions, and approaches for the treatment of norovirus disease.     

人肺癌相关抗原基因ALT04-AG表达抑制对人肺癌细胞株生长及基因表达谱的影响

目的探讨抑制人肺癌相关抗原基因ALT04-AG表达对细胞生长特性及相关基因表达的影响。方法重组反义ALT04-AGRNA真核表达质粒,经脂质体介导转染人肺癌细胞株(L78),以MTT、FCM法分析转染细胞生长特性,以Northern b lot、免疫组化染色及基因芯片技术检测相关基因的表达。结果构建了重组表达反义ALT04-AGRNA的真核表达质粒[pALT04-AG(as)],经该质粒转染及二氟甲基鸟氨酸(d ifluorom ethylorn ith ine,DFMO)处理的L78细胞均引起ALT04-AG表达下调及细胞的增殖抑制,后者还导致细胞凋亡比例增加。结论pALT04-AG(as)转染L78细胞或用DFMO抑制多胺生物合成,可通过对相关基因表达的调控促使L78细胞恶性表型逆转,而后者的作用更为广泛。本结果为探索肿瘤的诊断与治疗提供了有意义的线索。     

Epidermal growth factor receptor expression in a retinoic acid-treated human melanoma cell line

Treatment of a human cell line (HXG-2), established from a metastatic melanoma, with retinoic acid (RA) induced morphologic differentiation and eliminated its cloning capacity in soft agar. With the v-erb B oncogene as a probe, slot blot hybridization of genomic DNA from parental HXG-2 cells did not show epidermal growth factor (EGF) receptor gene amplification as compared with normal diploid fibroblasts. Analysis of RNA as well as EGF receptor determinations from HXG-2 and RA-treated HXG-2 cells showed essentially no differences, indicating that RA treatment does not modulate EGF receptor gene expression. Although enhanced EGF receptor expression is found in some advanced-stage melanomas, RA-induced changes in the transformation phenotype of cell line HXG-2 probably do not result from modulation of the EGF-mediated pathway.     

NP9对鼻咽癌细胞系CNE1基因表达谱的影响

目的： 确定NP9表达对鼻咽癌CNE1细胞基因表达谱的影响。方法: 稳定表达NP9基因和稳定转染空载体的CNE1细胞分别作为基因芯片分析的实验组和对照组，用高通量的基因芯片筛选实验组细胞的差异表达基因，荧光定量逆转录PCR验证部分基因表达差异。结果: 所分析的14 500个基因中，266个检测出有表达差异，其中82个基因呈现表达上调（RA＞1），184个基因显示表达下调（RA＜1）。显著上调和下调的基因分别为34个（RA＞1.5）和75个（RA＜1.5）。结论: NP9基因的表达导致了CNE1细胞中与细胞周期调控、细胞增殖和分化、细胞信号转导、细胞黏附相关基因表达的改变，为NP9的功能及分子机制研究提供了新的线索。     

Induction of FAS ligand expression in a human hepatoblastoma cell line by HCV core protein

Tumour cells and virus infected cells expressing Fas ligand (FasL) can evade immune surveillance by inducing apoptosis in T cells expressing Fas. In order to characterise a possible role of hepatitis C virus (HCV) core protein in similar mechanisms during HCV infection, we investigated Fas ligand expression and activity in a human hepatoblastoma cell line (HepG2) constitutively expressing this protein.Strong FasL induction was detected by immunoblotting and flow cytometry analysis in the core expressing cell lines Hep39. In contrast, vector transfected cells or cell lines expressing HCV E1-E2 proteins did not show FasL expression. Co-cultivation experiments of Hep39 cells with a Fas-sensitive T cell line indicated that FasL induced by the core protein had apoptotic activity toward target cells. Effect of the core protein on induction of FasL promoter was further examined by co-transfection of HepG2 cells with core-bearing plasmid and a vector in which luciferase gene expression is driven by human FasL promoter. Results of the luciferase assay indicated a positive regulation of FasL promoter by the core protein. In conclusion, HCV core protein plays a role in the induction of functional FasL in hepatoblastoma cell line and apoptosis in a target T cell line expressing Fas. Similar mechanisms may contribute, in vivo, to establishment of chronic infection and development of hepatocellular carcinoma (HCC).     

Induction of HLA-DQ antigen expression on a human promyelocytic leukemia cell line HL-60

A variant strain of HL-60, which is positive for HLA-DR antigen, was induced to express HLA-DQ antigen following treatment with phorbol esther. It was preceded by cell cycle arrest in the G1 phase and was accompanied by augmented phagocytosis. This differential expression of HLA class II antigens on this subline may contribute to understanding the functional role of HLA class II antigens in the hematopoietic differentiation of macrophage cells.     

Multilineage differentiation and characterization of the human fetal osteoblastic 1.19 cell line: a possible in vitro model of human mesenchymal progenitors

The in vitro study of human bone marrow mesenchymal stromal cells (BMMSCs) has largely depended on the use of primary cultures. Although these are excellent model systems, their scarcity, heterogeneity, and limited lifespan restrict their usefulness. This has led researchers to look for other sources of MSCs, and recently, such a population of progenitor/stem cells has been found in mesodermal tissues, including bone. We therefore hypothesized that a well-studied and commercially available clonal human osteoprogenitor cell line, the fetal osteoblastic 1.19 cell line (hFOB), may have multilineage differentiation potential. We found that undifferentiated hFOB cells possess similar cell surface markers as BMMSCs and also express the embryonic stem cell-related pluripotency gene, Oct-4, as well as the neural progenitor marker nestin. hFOB cells can also undergo multilineage differentiation into the mesodermal lineages of chondrogenic and adipocytic cell types in addition to its predetermined pathway, the mature osteoblast. Moreover, as with BMMSCs, under neural-inducing conditions, hFOB cells acquire a neural-like phenotype. This human cell line has been a widely used model of normal osteoblast differentiation. Our data suggest that hFOB cells may provide for researchers an easily available, homogeneous, and consistent in vitro model for study of human mesenchymal progenitor cells.