螺旋藻多糖对人宫颈癌Hela细胞体外生长的影响

目的：探讨螺旋藻多糖（PSP）对体外培养的Hela细胞生长的影响。方法：采用MTT法测定 PSP的抗肿瘤活性，并观察其对Hela细胞形态学的影响。结果：随PSP浓度的升高，Hela细胞存活率逐渐降低，抑制率逐渐增加；光镜下观察显示，PSP作用24-48h后，细胞出现明显的形态学改变。结论：PSP能抑制Hela细胞增殖。     

新城疫病毒pIRHN核酸疫苗构建和表达及对肿瘤细胞的影响

目的：研究NDV HN基因抗肿瘤作用及其可能机制。方法：以 pIRES1neo为表达载体构建了 NDV HN基因的pIRHN核酸疫苗，在体外转染HeLa细胞，用间接免疫荧光和Western blot检测pIRHN在真核细胞中表达状况，用荧光显微镜、DNA琼脂糖电泳及TUNEL染色等方法，检测HN基因导致细胞死亡的类型；用3，5-二羟基甲苯法测定HeLa细胞唾液酸含量的变化。结果：pIRHN 转染HeLa细胞后，能够在真核细胞中表达，能促进肿瘤细胞死亡，其死亡方式主要以诱导细胞凋亡为主，pIRHN使 HeLa细胞唾液酸含量减少。结论：用 NDV HN基因所构建的核酸疫苗能够在真核细胞中高效表达，表达的 HN蛋白主要位于胞膜，胞浆中亦有 HN蛋白表达；pIRHN具有抗肿瘤作用，可能通过其表达产物与肿瘤细胞唾液酸受体的相互作用，发挥其抗肿瘤作用。本实验为迸一步阐明NDV抗肿瘤作用机制提供了理论依据。     

榄香烯对人肝癌细胞7402、宫颈癌细胞Hela的凋亡诱导作用及下调Bcl—2蛋白表达

目的 研究中药榄香烯对人肝癌细胞7402、宫颈癌细胞Hela的体外抑瘤效应及其作用机制。方法 应用MTT比色法观察榄香烯对细胞的生长抑制作用;荧光显微镜和透射电镜观察细胞结构的改变;DNA凝胶电泳和流式细胞术等分析细胞凋亡、周期变化及Bcl-2蛋白表达情况。结果 榄香烯对7402和Hela细胞有较强的体外增殖抑制作用。榄香烯能诱导7402和Hela细胞凋亡,同时伴随有Bcl-2蛋白表达下调。结论 榄香烯对7402、Hela细胞有较强的抗瘤效应,其可能与Bcl-2蛋白下调引起的细胞凋亡有关。     

Nucleostemin特异性RNA干扰对HeLa细胞体内外增殖能力的影响

目的：检测肿瘤细胞Nucleostemin（NS）的表达，研究NS特异性RNA干扰对HeLa细胞体内外增殖的影响。方法：提取6种肿瘤细胞总RNA，用 RT-PCR和Northern blot方法检测NS的表达。用NS特异性siRNA表达载体转染HeLa细胞，观察转染的HeLa细胞（简称NS-siRNA-HeLa细胞）体内外增殖的变化。结果：6种肿瘤细胞中NS明显高表达。体外培养的NS-siRNA-HeLa细胞中NS表达显著低于对照组，G0/G1期细胞百分率显著升高。体内致瘤实验显示，NS-siRNA-HeLa细胞在裸鼠体内增殖显著低于对照组。结论： NS在肿瘤细胞中高表达具有普遍性。NS特异性RNA干扰使HeLa细胞进入S期受阻，并可明显降低HeLa细胞体内外的增殖能力。     

Immune responses to the expressed products of the CSP antige n gene of Plasmodium falciparum southern China isolate FCC1/HN in Hela cell

Objective To construct a eukaryotic expression system with pcDNA3-PfCSP/Hela for the Circ umsporozoite protein (CSP) gene of Plasmodium falciparum (P.falciparum), t o observe the immune responses in BALB/c mice induced by the expressed proteins .Methods The recombinant plasmid pcDNA3-PfCSP was transformed into the Hela cell line. The expressed protein was isolated and analyzed by using SDS-PAGE and used for immunization of BALB/c mice by subcutaneous, intravenous, and intraperitone al adminstration.Enzyme-linked immunosorbent assay(ELISA), Dot-ELISA, Wester n blot, T lymphocyte proliferation test, natural killer cell(NKC) activity assay , and CD4(+) and CD8(+) T cell detection were used for observation of humoral an d cellular immune responses.Results Immune sera strongly reacted with the expressed protein, antibody titer was up to 1∶6400 as detected by ELISA.Western blot analysis revealed a specific b and at 38.3 Kda.When the spleen cells of normal and immunized BALB/c mice we re specifically stimulated with expressed protein, the optical densities were 0 .12±0.03 and 0.34±0.04, respectively.The latter were significantly highe r than the former (P<0.01).We used the MTT colorimetric assay to measure NKC activity of mice spleen.The results showed that the NKC activity of immuni zed BALB/c mice was remarkably higher than that of the controls (P<0.05). CD4(+) and CD8(+) T cells were detected by using monoclonal antibody immunofluor escence methods.The results showed that the percentage of CD4(+) and CD8(+) T cells of immunized group were significantly higher than that of control group ( P<0.05).Conclusions The humoral and cell-mediated immune responses and elevated NKC activity to pr oducts made with a eukaryotic expression system could be specifically detected i n BALB/c mice.These findings indicate that the expressed protein could enhance the immune function in mice.     

HSV-1致Hela细胞凋亡的扫描电镜及透射电镜观察

目的 探讨HSV－1感染Hela细胞时,是否发生凋亡。方法 用扫描电镜及透射电镜观察感染72h的Hela细胞的形态变化。结果 正常的Hela细胞表面有丰富的微绒毛。细胞间隔清晰,可幼稚连接,连接不紧密。胞浆内线粒体嵴结构完整,并可大量的粗面内质网及游离核糖体,极少的脂滴。细胞核内多为常染色体,核仁清晰,核浆比约1：1。凋亡细胞中核内异染色质增多,染色质浓缩成块状,边集于核膜。线粒体轻度肿胀,溶酶体代偿性增多。核膜、胞膜、各细胞器膜均完整。坏死细胞的染色质呈不规则的团块状,但不象凋亡细胞那样集中于核周边。细胞膜、核膜和细胞器的结构破坏,线粒体空泡变,细胞崩解,胞浆及其内容物外泄。HSV－1感染的Hela细胞体积明显缩小,核浆比例明显增加。细胞表面的微绒毛断裂、减少,细胞膜球状突起。核膜完整,核致密,呈块状分布,边集于核膜。细胞膜、细胞器结构完整。可见出泡现象和凋亡小体形成现象。在胞核内可见病毒体。结论 ①HSV－1感染Hela细胞时,造成细胞死亡形式是核死亡形式即凋亡;②HSV－1感染Hela细胞时,细胞凋亡占绝对优势,同时伴有细胞坏死。     

流式细胞术检测九节龙皂甙对Hela细胞凋亡的诱导作用

0　引言　细胞凋亡 (Apoptosis)是一种生理性细胞程序化死亡 ,与肿瘤的发生、发展密切相关 ,已成为当前肿瘤研究的新热点 .九节龙皂甙是从川产九节龙全草中提出的单体[1 ] ,我们观察九节龙皂甙诱导 Hela细胞凋亡 ,采用流式细胞术(flow cytometry,FCM)进行分析 ,结果如下 :1　材料和方法1.1　细胞系及细胞培养　人宫颈癌细胞株 (Hela)为上皮样癌细胞 ,本校 40 3研究所提供 .细胞培养方法见参考文献[2 ].1.2　细胞凋亡的诱导和分析　实验组分别以 1.5 6～ 5 0 .0μg· m L- 1九节龙皂甙 (本校药物研究所提供 )作用 Hela细胞(10 6 · m L- …     

含有纯化标签的重组人表皮生长因子的构建及表达

目的:高效表达和制备有活性的人表皮生长因子(hum an ep iderm al growth factor,hEGF)。方法:用DNA重组技术构建EGF基因并插入表达载体pQE-40,与载体上的6×H is标签融合,获得的表达质粒pQE-EGF转化E.coliM15[pREP4],得工程菌M15[pQE-EGF]。MTT法测定活性,培养工程菌M15[pQE-EGF]并诱导目的蛋白表达,镍离子螯合亲和层析纯化重组EGF。结果:所构建的EGF序列正确,重组EGF在大肠杆菌中以包涵体形式存在,表达量占菌体总蛋白的21.2%,经镍离子亲和层析一步纯化后的重组EGF纯度达90%,得率为94%。复性后的重组EGF具有促Hela细胞生长活性。结论:以E.coliM15[pREP4]/pQE-40系统表达EGF具有技术路线简单、目的蛋白得率高、成本低等优点,是一种制备活性hEGF的有效手段。     

The influence of serum for spreading of tumor cells on synthetic glycolipid films

Adhesion and spreading of tumor cells to the films of a galactose-, glucose-, or phosphatidylcholine-bearing lipid was studied. Human adenocarcinoma Hela cells, B16 mouse melanoma cells, and HuH-7 human hepatoma cells selectively adhered and spread on galactose-bearing lipid in serum-containing medium, but not in serum-free medium. The spreading of the tumor cells in serum-containing medium was inhibited in the presence of lactose, but not in the presence of maltose. Cell spreading also occured on the galactose-bearing glycolipid film pre-treated with serum. From quantitative analysis for the adsorption of serum components by a quartz-crystal microbalance, the surfaces of the lipid films were found to be entirely covered with serum components. These results suggested that serum components pre-adsorbed on the galactose-bearing lipid influence the cell spreading.     

17-β雌二醇对宫颈癌Hela细胞环氧合酶-2和增殖细胞核抗原表达的影响

目的:探讨17-β雌二醇(E2)对宫颈癌Hela细胞环氧合酶-2(cyclooxygenase-2,COX-2)和增殖细胞核抗原(proliferatingcell nuclear antigen,PCNA)表达的影响。方法:Hela细胞分别用0.1%乙醇(对照组)、E2(E2组)、E2+依托昔布(E2+依托昔布组)作用24 h后,采用免疫细胞化学、Western blot及RT-PCR方法检测COX-2和PCNA的表达。结果:E2组COX-2和PCNA蛋白及mRNA水平较对照组明显升高(P0.01);而E2+依托昔布组PCNA水平较E2组显著降低(P0.01),但依然高于对照组。结论:E2可促进Hela细胞COX-2表达,并进一步上调PCNA水平。