Significance of cyclin E, pl6ink4a and ki67 Overexpression in Cervical Exfoliated-cell Specimens for Primary Screening of HPV-related Cervical Carcinoma

The aim of this study is to investigate cyclin E, pl6inkda and ki67 as possible diagnostic biomarkers for cervical preneoplasia using cervical exfoliated-cell specimens, and evaluate the significance for screening patients at high risk of developing cervical carcinoma. The expression of cyclin E, pl6inkda and ki67 was examinated in 78 cervical exfoliated epithelial specimens diagnosed as atypical squamous cells of undetermined significance (ASCUS) (12 cases), cervical intraepithelial neoplasia (CIN) of type 1 (17 cases), CIN2_3 (38 cases) and invasive carcinoma (11 cases) using immunohistochemical analysis, and simultaneously, the DNA status of human papillomavims (HPY) type 16/18 was detected by polymerase chain reaction (PCR) using type specific primers, cyclin E, pl6inkda and ki67 were all overexpressed in CINs and invasive carcinoma, compared with little expression in ASCUS ( P <0.005). Overexpression of cyclin E was observed in CIN1(94.1%, X^2 = 21.16, P < 0.01), and p16inkda and ki67 were overexpressed in invasive carcinoma (100% and 90.9% respectively). The degree of pl6inkda and ki67 expression correlated well with that of epithelial lesions ( P < 0.005). HPV16/18 infection was assessed in C1Ns and invasive carcinoma samples, and revealed a significant relationship with the degree of cervical epithelial lession. The expression level of pl6inkda and ki67 seemed more closely associated with HPVI6 infection than that of cyclin E (rs= 1.0 vs rs=0.4). Only 1 case in CINI and d cases in CIN2-3 of HPV18 positive samples were detected. Therefore no statistical significance was found by statistical analysis. Overexpression of cyclin E, pl6inkda and ki67 in CINs and invasive carcinoma cells demonstrates the potential use of cyclin E, pl6inkda and ki67 as diagnostic biomarkers for HPV-related cervical neoplastic lesions. In addition, this technique can be used for screening patients at high risk of developing cervical carcinoma.     

RNAi沉默HMGA1基因对肝癌细胞凋亡和细胞周期的影响

目的:筛选HMGA1基因沉默稳定转染肝癌细胞株,检测沉默后肿瘤细胞凋亡、周期和增殖的变化。方法:靶向HMGA1的RNAi载体转染肝癌细胞,G418筛选得到稳定转染细胞株并鉴定;MTT法和FACS检测细胞增殖、凋亡及细胞周期。结果:成功建立基因沉默HMGA1稳定细胞株;HMGA1基因沉默后,细胞凋亡百分率(29.46±3.04%)明显高于质粒对照组和空白对照组(1.96±0.76%和2.04±0.70%,P<0.01);G0-G1期细胞百分率(75.21±2.35%)明显高于质粒对照组和空白对照组(37.98±3.02%和39.23±3.63%,P<0.01),G2-S期(24.79±2.35%)显著低于质粒对照组和空白对照组(62.03±3.01%和60.77±3.63%,P<0.01);MTT检测显示,HMGA1沉默细胞增殖与质粒对照组和空白对照组相比显著降低(P<0.01或0.05)。结论:基因沉默HMGA1可促进肿瘤细胞凋亡,抑制细胞增殖。     

脂质体转染细胞周期素B1反义脱氧寡核苷酸对HL60细胞增殖调控的影响

目的:探讨脂质体转染细胞周期素B1(cyclinB1)反义脱氧寡核苷酸(ASON)对HL60细胞增殖调控的作用。方法:用针对cyclinB1mRNA5’端编码区起始密码子(ATG/AUG)的ASON,通过脂质体导入HL60细胞共培养后,用流式细胞术(FCM)和RT-PCR分别检测cyclinB1蛋白和mRNA的表达水平,电镜和原位细胞凋亡检测法(POD)、FCM及DNA凝胶电泳法检测细胞凋亡。结果:CyclinB1ASON组与SON及空白对照组相比,ASON能特异地抑制cyclinB1蛋白及mRNA水平的表达,当ASON的浓度达到一定程度时,HL60细胞的增殖及集落形成率均明显受抑制,出现细胞凋亡,并且此作用随ASON浓度的升高而增强。结论:CyclinB1的特异ASON能封闭其蛋白及mRNA的表达水平,可剂量依赖性地抑制白血病细胞增殖,诱导细胞凋亡。     

NF2基因在丙肝病毒非结构蛋白4B调节细胞周期中的作用

目的在HEPG2细胞内观察丙肝病毒(HCV)非结构蛋白4B(NS4B)对神经纤维瘤病Ⅱ型(NF2)基因的影响及NF2在NS4B调节细胞周期中的作用。方法将细胞分为多组,其中3组对照,分别为无处理组、pCDNA3.1空质粒组和si-NF2 control组;3组作用组,分别为NS4B质粒组、NF2质粒组和si-NF2沉默基因组。每组经过转染、培养后收获细胞;RT-qPCR和Western blot检测细胞内相应基因转录及表达;流式细胞仪检测细胞周期。结果 NS4B可降低NF2转录水平(P0.01),同时NS4B的高表达和NF2的低表达对细胞周期具有相似的作用,及促进细胞周期向S期的转变;而NF2的高表达则使细胞周期停滞在G_0/G_1。结论 NS4B对NF2具有一定的调控作用,NF2可能参与NS4B调控细胞周期机制。     

表皮生长因子影响肿瘤患者舌苔变化的分子机制研究

目的 研究表皮生长因子（EGF）对食管癌细胞Eca-109细胞周期及表皮生长因子受体（EGF－R）表达的影响，探讨EGF促进肿瘤患者舌苔增厚的分子机制。方法 应用流式细胞术，检测EGF－R的表达和细胞周期。结果 EGF能明显促进Eca-109细胞膜上EGF－R的表达，使细胞增殖活性大大增强。结论EGF有是通过促进EGF－R的表达影响舌苔形成。     

人参单体皂甙Rh2对前列腺癌细胞株PC-3M细胞移行周期的影响

目的:探讨Rh₂对体外培养的PC-3M细胞移行周期的影响。方法:应用[³H]-TdR掺入法和流式细胞仪测定DNA含量及进行细胞周期的分析。结果:PC-3M细胞的[³H]-TdR掺入量明显少于对照组(P＜0.01);流式细胞仪测定G₁期细胞数百分比明显高于对照组。结论:Rh₂可使PC-3M细胞增殖周期阻滞在G₁期,肿瘤细胞增殖减慢。     

p53介导的凋亡相关基因的研究进展

p5 3是重要的细胞周期调控基因之一 ,在参与维持细胞周期的正常运行中起着重要作用。p5 3基因通过诱导细胞发生 G1 期阻滞修复损伤 DNA和诱导细胞凋亡来抑制肿瘤细胞的增殖。在 p5 3基因介导细胞凋亡的通路中 ,有许多已知和未知的相关基因发生作用。本文就 p5 3基因调控细胞凋亡途径的上、下游已知基因作一综述。     

MTS1／p16抑制基因的克隆及其对宫颈癌细胞系的影响

利用重组ＲＴ－ＰＣＲ方法从人胎盘中扩增多重肿瘤抑制基因（ＭＴＳ１）全长ｃＤＮＡ片断，克隆测序后，亚顾隆入哺乳动物高铲表达质粒ｐＣＥＰ中。将表达质业转染两种遗传背影不同的吕颈癌细胞系，发现外源基因ＭＴＳ１／ｐ１６基因的导入对ＨＰ〖Ｖ阳性的宫颈癌细胞系具有明显生长抑制作用，并出现细胞滞留Ｇ１期的特性。     

应用组织芯片技术研究霍奇金淋巴瘤细胞凋亡和细胞周期相关基因蛋白的表达

目的探讨细胞凋亡和细胞周期相关基因蛋白与经典型霍奇金淋巴瘤临床分期的关系.方法应用免疫组化ABC法检测bcl-2,ABC和TUNEL双标记技术检测细胞凋亡,ABC和SABAP双标记方法检测MIB1、TopoⅡa、Rb和p21在62例经典型霍奇金淋巴瘤组织芯片中的表达.结果62例经典型霍奇金淋巴瘤中35例bcl-2阳性,在62例非肿瘤性背景细胞中均有凋亡小体,而在H-RS细胞仅有10例胞质内见凋亡小体;H-RS细胞bcl-2表达与细胞凋亡呈负相关性(P＜0.05);MIB1及TopoⅡa高表达与临床分期呈正相关性(P＜0.01),反之,Rb及p21高表达与临床分期呈负相关性(P＜0.01).所有抗原的表达与霍奇金淋巴瘤的组织学亚型均无相关性(P＞0.05).结论bcl-2的过度表达可以阻断H-RS细胞的凋亡;CD30阳性H-RS细胞中MIB1及TopoⅡa表达指数随病程的进展而升高,而Rb及p21表达指数随病程的进展而降低,他们可作为霍奇金淋巴瘤预后判断的参考指标.霍奇金淋巴瘤的病程进展可能与细胞凋亡和细胞周期相关基因及蛋白表达的变化有关.