宫颈癌化疗前后肿瘤细胞凋亡及相关基因Bcl-2、Bax表达的对比分析

目的 探讨宫颈癌化疗前后肿瘤细胞Bcl-2、Bax蛋白表达及凋亡的变化及其临床意义.方法 采用免疫组织化学S-P法及TUNEL法,检测55例宫颈癌单一拓扑替康化疗前后的110个肿瘤样本中Bcl-2、Bax蛋白水平及细胞凋亡水平.结果 子宫颈癌单一拓扑替康化疗后有效率为34.5%(19/55);化疗前后凋亡阳性率分别为41.8%、50.9%,无显著性差异(P＞0.05).Bcl-2蛋白在子宫颈癌化疗前后的表达阳性率分别为54.5%和49.1%,无显著性差异(P＞0.05);Bax蛋白在子宫颈癌化疗前后的表达阳性率分别为50.9%和74.5%,有显著性差异(P＜0.05);组织学有效的病例化疗前癌组织中Bcl-2的阳性率为52.6%(9/19),组织学无效病例的阳性率为55.6%(16/36),无显著性差异(P＞0.05);组织学有效的病例化疗前癌组织中Bax阳性率为63.2%(7/19),组织学无效病例的阳性率为44.4%(20/36),无显著性差异(P＞0.05).结论 拓扑替康具有诱导宫颈癌细胞凋亡的作用,其作用与上调Bax的表达相关.化疗前癌组织Bcl-2、Bax蛋白表达量与拓扑替康组织学疗效无相关性.     

口腔癌组织中Bcl-2和Bax基因表达与凋亡关系的研究

目的通过对口腔癌组织及正常组织中Bcl-2和Bax基因表达及凋亡细胞的研究,探讨口腔癌组织及正常组织中凋亡与基因表达的关系.方法利用免疫组化法检测67例口腔癌组织标本及42例癌旁正常口腔组织中Bcl-2和Bax基因表达,利用TUNEL方法检测口腔癌组织标本中凋亡的情况.结果Bcl-2、Bax基因在口腔癌组织中的阳性表达率分别为49.3%和47.8%;在正常口腔组织中的阳性表达率分别为23.8%和81.0%.Bcl-2、Bax基因在口腔癌组织及正常口腔组织中的阳性及阴性表达均具有显著性差异;二者在口腔癌组织中的阳性及阴性表达,其细胞凋亡的指数均有显著性差异.Bcl-2、Bax基因表达呈显著性负相关(γ=-0.4669).结论口腔癌组织中Bcl-2基因阳性表达时,其本底凋亡指数较阴性表达时低;而Bax基因阳性表达时,其本底凋亡指数较阴性表达时高,二者在细胞凋亡过程中存在着拮抗作用的关系.     

苦参碱对大肠癌细胞凋亡及Bax、Bcl-2表达的影响

[目的]探讨苦参碱对人大肠癌Lovo细胞增殖抑制和凋亡诱导作用及其对Bax、Bcl-2表达的影响。[方法]0.05~1.6mg/ml不同浓度苦参碱作用Lovo细胞,采用MTT法检测苦参碱对大肠癌Lovo细胞增殖抑制作用,DNAladder、AnnexinⅤ-PI法及TUNEL染色检测细胞凋亡,WesternBlot法检测凋亡相关蛋白Bax、Bcl-2表达的变化。[结果]0.05～1.6mg/ml苦参碱处理Lovo细胞24h或48h后,细胞增殖均明显受抑制;DNAladder、AnnexinⅤ-PI法及TUNEL染色检测结果显示苦参碱呈时间、剂量依赖性诱导细胞凋亡;促凋亡蛋白Bax随着苦参碱剂量增加表达增加,抗凋亡蛋白Bcl-2随着苦参碱剂量增加表达减少。[结论]苦参碱具有抑制大肠癌细胞增殖,诱导其凋亡的作用。苦参碱诱导大肠癌细胞凋亡的机制可能与促凋亡蛋白Bax表达增加、抗凋亡蛋白Bcl-2表达减少有关。     

TGF-β1诱导HL-60细胞凋亡及TIEG1,Bcl-2/Bax的表达变化

目的:研究TGF-β1诱导HL-60细胞凋亡后TIEG1及Bcl-2/Bax的表达变化。方法:用不同浓度的TGF-β1处理HL-60细胞,采用MTT法检测细胞生长抑制率。用10.4 ng/ml TGF-β1处理HL-60细胞,以流式细胞仪检测细胞凋亡,以RT-PCR方法检测TIEG1、Bcl-2及Bax的表达。结果:TGF-β1对HL-60细胞具有增殖抑制作用,其抑制效应呈时间及剂量依赖性。10.4 ng/ml TGF-β1增殖抑制作用较为明显。在细胞凋亡过程中,TIEG1、Bax表达呈升高趋势,而Bcl-2呈下降趋势。结论:TGF-β1可以抑制HL-60细胞增殖,并诱导其凋亡,且呈时间、剂量依赖性。在TGF-β1诱导HL-60细胞凋亡的过程中,TIEG1表达逐渐加强,与HL-60细胞凋亡呈负相关。Bcl-2/Bax与TGF-β1诱导HL-60细胞的凋亡相关,且与TIEG1的表达有相关性。     

大肠癌中FHIT基因表达及与Bcl-2、Bax表达的相关性研究

目的　探讨Fhit蛋白在大肠癌组织中的表达情况及其与Bcl 2、Bax蛋白之间的关系。方法　采用免疫组化S P法检测 84例大肠癌组织中Fhit、Bcl 2、Bax蛋白的表达。结果　Fhit、Bcl 2、Bax蛋白在大肠癌中表达阳性率分别为 48 81%、5 3 3 7%和 47 62 %。Fhit蛋白低或不表达与肿瘤的组织学类型无关 (P >0 0 5 )而与浸润深度、分化程度、Ducks’分期、转移有关 (P <0 0 5 )。大肠癌中Fhit蛋白表达与Bcl 2、Bax蛋白表达有显著相关性 (P <0 0 5 ) ,Bcl 2、Bax蛋白表达二者之间无相关性 (P >0 0 5 )。结论　①Fhit蛋白表达缺失在大肠癌中是频发事件 ,FHIT基因参与大肠癌的发生发展 ,是一个侯选的抑癌基因。②大肠癌中Fhit蛋白表达失活可能通过上调Bcl 2蛋白和下调Bax蛋白表达导致大肠癌的发生。     

siRNA介导的STAT3基因沉默对宫颈癌HeLa细胞凋亡和化疗敏感性的影响

目的:应用siRNA介导的STAT3基因沉默,研究其对HeLa细胞凋亡和化疗敏感性的影响.方法:体外合成靶向STAT3的siRNA-1和siRNA-2,并用脂质体转染HeLa细胞,RT-PCR检测干扰STAT3 mR-NA敲减的效率,蛋白质印迹法检测敲减后STAT3蛋白的表达;通过记录生长曲线观察干扰后细胞的增殖情况.用AnnexinV/PI双染,结合流式细胞仪检测细胞早期凋亡率.MTT法检测肿瘤细胞对几种化疗药物的敏感性.结果:siRNA使STAT3表达下调,mRNA水平抑制率为74.8%和60.4%;蛋白表达水平抑制达68.0%和59.0%.干扰后HeLa细胞增殖缓慢,早期凋亡率明显增加.对顺铂、长春瑞滨和吉西他滨有增敏作用.结论:siRNA介导的STAT3沉默可以抑制宫颈癌HeLa细胞的增殖,诱导凋亡,增加对化疗药物的敏感性,可望成为一种新的治疗策略.     

Bcl-2基因对丝裂霉素C诱导的膀胱癌细胞凋亡抑制及其机制的研究

目的：探讨Bcl-2基因对丝裂霉素C（MMC）诱导的膀胱癌细胞凋亡的抑制作用及其分子机制。方法：采用DNA重组技术构建真核表达载体pcDNA3．1（＋）／Bel-2;利用脂质体将重组载体pcDNA3．1（＋）／Bcl—2和空载体pcDNA3．1（＋）转入人膀胱癌BIU-87细胞,RT-PCR检测基因转录水平;MMC作用于转染前、后的细胞,应用四甲基偶氮唑蓝（MTT）比色法、吖啶橙（AO）荧光染色法、Western blot技术对细胞生长、凋亡、Caspase-3活性及细胞色素C在细胞内分布的变化进行检测。结果：1）成功构建真核表达载体pcDNA3．1（＋）／Bcl-2,建立Bcl-2基因高表达的膀胱癌细胞株BIU-87／Bcl-2。2）在MMC作用下,与未转染Bcl-2基因的膀胱癌细胞株BIU-87和BIU-87／neo相比,BIU87／Bcl-2细胞株存活率上升,P〈0．01,其IC50是后者的15倍,q＝6．41,Pd0．01;BIU-87／Bcl-2的凋亡率显著降低,q值分别为22．62和20．45,P值均〈0．01,Caspase-3激活和线粒体细胞色素C的释放受到抑制,BIU-87／Bcl-2的A450显著低于BIU-87和BIU87／neo,q值分别为28．20和26．70,P值均〈0．01,表明BIU-87／Bcl-2的Caspase-3激活受限。结论：Bcl-2基因高表达通过抑制线粒体细胞色素C的释放和Caspase-3的激活,使膀胱癌细胞对MMC诱导的凋亡产生抗性,导致耐药性的产生。     

宫颈癌组织中Trail受体DR4与凋亡相关蛋白Bax表达及其临床意义的研究

目的:通过检测宫颈癌组织中肿瘤坏死因子相关凋亡诱导配体(Trail)的受体DR4及凋亡相关蛋白Bax的表达,探讨宫颈癌组织中的细胞凋亡因素,为寻找新的治疗途径提供思路.方法:随机选取41例宫颈鳞癌患者按照分化程度分高/中分化组(A组)23例和低分化组(B组)18例,选取20例慢性宫颈炎患者作为对照组(C组),利用免疫组织化学ABC染色法检测宫颈癌组织中DR4和Bax的表达.结果:DR4、Bax在3组中均有表达,C组中DR4表达最高,为4.72±0.35,与A、B组比较差异有统计学意义,P<0.05;A组DR4表达为3.23±1.94,高于B组的1.35±1.14,差异有统计学意义,P<0.05.Bax在C组中表达最低,为6.05±1.04,在A组中表达最高,3组比较差异有统计学意义,P<0.05,结论:分化程度高的癌组织中DR4和凋亡因子Bax表达高于分化程度低的癌组织,DR4、Bax的表达与宫颈癌的预后有关.     

Bcl-2基因在人膀胱癌细胞中表达对CTL凋亡诱导作用的影响

目的:探讨Bcl-2基因在人膀胱癌细胞中表达对细胞毒性T淋巴细胞(CTL)凋亡诱导作用的影响。方法:采用基因重组技术构建真核表达载体pcDNA3.1(+)/Bcl-2;应用脂质体介导的基因转染技术将pcDNA3.1(+)/Bcl-2导入膀胱癌BIU-87细胞,RT-PCR检测Bcl-2基因表达水平;不同浓度抗Fas单克隆抗体(Anti-Fasmab)模拟CTL的Fas-配体(Fas-L)处理转染与未转染Bcl-2基因的膀胱癌细胞,采用四甲基偶氮唑蓝(MTT)比色法、吖啶橙(AO)荧光染色法和流式细胞检测仪(FCM)分析细胞存活和凋亡情况。结果:酶切和核酸测序证明真核表达载体pcDNA3.1(+)/Bcl-2构建成功。将重组质粒转染膀胱癌细胞后,RT-PCR分析表明,转染Bcl-2基因的BIU-87/Bcl-2细胞的Bcl-2基因表达水平较BIU-87细胞和转染空载体的BIU-87/neo细胞显著增高,P<0.01。An-ti-Fasmab作用后,BIU87/Bcl-2细胞的存活率显著高于BIU-87和BIU-87/neo细胞,P<0.05或P<0.01。3种细胞虽均发生凋亡的形态学改变,但BIU87和BIU87/neo细胞改变更为明显,两者的凋亡率分别为(25.33±3.00)%和(27.05±1.75)%,而BIU-87/Bcl-2细胞的凋亡率为(18.06±1.41)%,显著低于前两者,P<0.01。结论:Bcl-2基因高表达通过阻断CTL杀伤膀胱癌细胞的Fas/Fas-L途径,使膀胱癌细胞能够抵御免疫系统中CTL的攻击。     

Survivin在胃癌组织中的表达及其与Bcl-2、Bax表达的关系

目的: 探讨Survivin在胃癌组织中的表达及其与Bcl2、Bax表达的关系,并讨论它们相关的临床意义。 方法：选择临床及病理资料完整的中国医科大学附属盛京医院2005-2007年手术切除并经病理证实为胃腺癌术前均未行化放疗的蜡块标本54例,另取良性胃黏膜组织15例。应用免疫组化SP法检测54例胃癌组织中Survivin、Bcl2、Bax的表达,并检测15例正常胃黏膜组织中Survivin的表达。 结果：54 例胃癌组织中有39例Survivin 表达阳性,阳性率为72.2%;15例正常胃黏膜组织中无Survivin 阳性表达。 Survivin的表达与胃癌的浸润深度、淋巴结转移和TNM分期密切相关（P<0.05或P<0.01),而与患者的性别、年龄、肿瘤大小、远隔转移及分化程度无相关性。Bcl2阳性表达者中Survivin阳性表达率为81.8%,Bcl2阴性表达者中Survivin阳性表达率为57.1%,Survivin与Bcl2的表达呈正相关（P<0.01);而Survivin与Bax的表达无明显相关性。结论：胃癌组织中Survivin的表达与肿瘤的浸润深度、淋巴结转移和临床分期密切相关;Bcl2的表达呈正相关,而与Bax的表达无明显相关性     

Survivin在胃癌组织中的表达及其与Bcl-2、Bax表达的关系

目的:探讨Survivin在胃癌组织中的表达及其与Bcl-2、Bax表达的关系,并讨论它们相关的临床意义. 方法:选择临床及病理资料完整的中国医科大学附属盛京医院2005-2007年手术切除并经病理证实为胃腺癌术前均未行化放疗的蜡块标本54例,另取良性胃黏膜组织15例.应用免疫组化S-P法检测54例胃癌组织中Survivin、Bcl-2、Bax的表达,并检测15例正常胃黏膜组织中Survivin的表达. 结果:54 例胃癌组织中有39例Survivin 表达阳性,阳性率为72.2%;15例正常胃黏膜组织中无Survivin 阳性表达. Survivin的表达与胃癌的浸润深度、淋巴结转移和TNM分期密切相关(P<0.05或P<0.01),而与患者的性别、年龄、肿瘤大小、远隔转移及分化程度无相关性.Bcl-2阳性表达者中Survivin阳性表达率为81.8%,Bcl-2阴性表达者中Survivin阳性表达率为57.1%,Survivin与Bcl-2的表达呈正相关(P<0.01);而Survivin与Bax的表达无明显相关性. 结论:胃癌组织中Survivin的表达与肿瘤的浸润深度、淋巴结转移和临床分期密切相关;Bcl-2的表达呈正相关,而与Bax的表达无明显相关性.     

抗凋亡基因bcl-2在子宫内膜癌的表达及意义

目的探讨抗凋亡基因ｂｃｌ２与子宫内膜癌的关系。方法应用免疫组化链霉菌－抗生物素过氧化物酶染色法,检测１５例子宫内膜非典型增生及５０例子宫内膜腺癌的石蜡包埋标本中ｂｃｌ２基因的表达。结果子宫内膜非典型增生组织中,ｂｃｌ２蛋白高表达,子宫内膜腺癌中ｂｃｌ２低表达（Ｐ＜０．０５）。在子宫内膜腺癌中,随着临床分期的升高、组织病理分级的降低及肌层浸润的加深,ｂｃｌ２表达阳性率逐渐下降（Ｐ＜０．０５）。结论ｂｃｌ２可能与子宫内膜组织的异常增殖有关,并主要在早期起作用。     

Bax and Bcl-2 protein expression following radiation therapy versus radiation plus thermoradiotherapy in stage IIIB cervical carcinoma

BACKGROUND: The relative amounts of Bcl-2 and Bax proteins determine cell survival or death following an apoptotic stimulus. To clarify the molecular mechanism of cell death after radiotherapy or thermoradiotherapy and its relation to the response of AJCC/UICC Stage IIIB cervical carcinomas, the expression of Bax and Bcl-2 proteins was investigated both before and in the course of treatment given during this study. METHODS: Thirty-seven patients with Stage IIIB carcinoma of the uterine cervix were treated with external beam irradiation to the pelvis combined with iridium-192 high-dose-rate intracavitary brachytherapy. All patients were randomized to one of the following two groups: the radiotherapy (RT) group of 19 patients who were given radiotherapy alone, and the thermoradiotherapy (TRT) group of 18 patients who were given 3 sessions of hyperthermia in addition to RT. Specimens of the cervical tumors were obtained by punch biopsy both before and in the course of the treatment (after a total dose of 10.8 grays ?Gy for the RT group or after 10.8 Gy plus 1 session of hyperthermia for the TRT group). The tumor sections were stained with anti-Bax and anti-Bcl-2 monoclonal antibody. On the basis of the percentage of immunopositive cells, both pretreatment and posttreatment samples were scored. Furthermore, relative changes in protein expression were determined by comparing the pretreatment scores with those in the course of treatment. In addition, treatment response was evaluated. RESULTS: A complete response was achieved in 52.6% (10 of 19) of the RT group versus 83. 3% (15 of 18) of the TRT group (P = 0.049). Better tumor control was accompanied by increased Bax expression, i.e., 10.5% (2 of 19) of the RT group versus 44.4% (8 of 18) of the TRT group (P = 0.02). The respective number of patients who partially responded (PR) or did not respond to treatment (NC) was 26.3% (5 of 19) and 21.1% (4 of 19) of the RT group versus 11.1% (2 of 18) and 5.6% (1 of 18) of the TRT group (P = 0.2 for both the PR and NC subgroups). CONCLUSIONS: TRT was found to result in better treatment responses than RT for patients with Stage IIIB cervical carcinoma. An additive or synergistic antitumor effect of TRT is likely to occur through induction of apoptosis involving one of the bax pathways.     

凋亡相关蛋白 Bcl-2和 Bax 在基底细胞样乳腺癌中的表达及意义

目的探讨 Bcl-2和 Bax 蛋白在基底细胞样乳腺癌（BLBC）中的表达及意义。方法免疫组织化学法检测43例 BLBC、57例非基底细胞样乳腺癌（non-BLBC）、60例正常乳腺组织中 Bcl-2和Bax 蛋白的表达,分析其与患者生理病理的关系。结果Bcl-2在 BLBC 中的表达率为69．77％,高于non-BLBC 的43．86％（χ2＝6．647,P ＝0．010）和正常乳腺组织的21．67％（χ2＝23．831,P ＝0．001）;Bax在 BLBC 中的表达率为20．93％,低于 non-BLBC 的45．61％（χ2＝6．564,P ＝0．010）和正常乳腺组织的76．67％（χ2＝31．270,P ＝0．001）。Bcl-2和 Bax 在 BLBC 中的表达水平与淋巴结转移（χ2＝6．927,P ＝0．008;χ2＝6．203,P ＝0．013）及 pTNM分期（χ2＝6．331,P ＝0．012;χ2＝5．972,P ＝0．015）相关。Bcl-2与 Bax 在 BLBC 中的表达呈负相关（r ＝－0．408,P ＜0．010）。结论Bcl-2和 Bax 分别在 BLBC 中高表达和低表达,两者可能共同作用使细胞增殖与凋亡平衡失调,从而促进了 BLBC 的发生、发展。     

化疗前后宫颈癌中VEGF表达和微血管密度变化的研究

目的:观察宫颈癌化疗前后癌组织中微血管形成及VEGF表达的变化,分析二者与临床病理因素和化疗疗效之间的关系.方法:免疫组织化学S-P法检测54例宫颈癌经单一拓扑替康化疗前后癌组织中的微血管密度(MVD)及癌细胞VEGF的表达水平.结果:化疗前MVD和VEGF的表达与病理类型有关,腺癌明显高于鳞癌,差异有显著性(P＜0.05);VEGF阳性组MVD值比VEGF阴性组高,差异有显著性(P＜0.001),且VEGF与MVD值呈正相关关系(P＜0.05);化疗前、后MVD值分别为76.22±31.25,52.22±32.56,差异有显著性(P＜0.005);化疗前、后VEGF的阳性表达率分别为28(51.9%)、12(22.2%),差异有显著性(P＜0.001);化疗疗效Ⅱ级(有效)和Ⅲ级(显效)的MVD和VEGF表达均高于无效组,差异有显著性(P＜0.05).结论:VEGF能促进宫颈癌新生血管形成.拓扑替康化疗可降低肿瘤细胞VEGF阳性表达率,对抑制肿瘤血管生长有一定的作用.肿瘤中VEGF促进肿瘤微血管生长的同时也有利于化疗药物发挥药效.     

Transient increases of apoptosis and Bax expression occurring during radiotherapy in patients with invasive cervical carcinoma.

BACKGROUND: Apoptosis plays a crucial role in radiation therapy (RT) in various carcinomas. This study was designed to investigate the relation between apoptosis and RT in invasive squamous cell carcinoma (ISCC) of the uterine cervix METHODS: Thirty-five specimens were obtained from 7 patients with ISCC before and during a fractionated RT. The occurrence of apoptosis was examined by end labeling of DNA gel fractionation and in situ 3' end labeling of DNA. The expression of Bax and Bcl-2 proteins was investigated by immunohistochemical staining. RESULTS: Autoradiographic analysis revealed that high molecular weight DNA was predominant in the untreated ISCC specimens. However, a ladder-like pattern, characteristic of the apoptotic breakdown of DNA, was identified at doses of 900 cGy and 1980 cGy. At doses >1980 cGy, DNA laddering disappeared without any extensive smearing. Quantitative analysis of low molecular weight fragments of DNA revealed significant increases at doses of 900 cGy and 1980 cGy compared with those before RT and at doses of >1980 cGy. Labeling of DNA in situ indicated that cells undergoing apoptosis increased dramatically at a dose of 900 cGy. However, apoptotic cells decreased at a dose of 3960 cGy. In addition, a large fraction of tumor cells was immunonegative for Bcl-2 before and during RT. By contrast, immunoreactive Bax was observed intensely in many neoplastic cells at doses of 900 cGy and 1980 cGy. CONCLUSIONS: The current investigation indicates that low doses of RT result in apoptotic cell death in ISCC in association with the increased expression of Bax but not with increased Bcl-2 expression.     

Apoptosis and the expression of Bax and Bcl-2 in squamous cell carcinoma and adenocarcinoma of the uterine cervix

BACKGROUND: Apoptosis plays a crucial role in the suicide and turnover of cells in various tumors. This study was designed to investigate the relation between apoptosis and the histologic types of cell in invasive cervical carcinoma. METHODS: Cervical tissues were obtained from 19 patients with invasive squamous cell carcinoma (ISCC), 9 patients with invasive endocervical adenocarcinoma (IEAC), and 15 patients with myoma uteri (which were used as controls). Each tissue was rapidly frozen and/or fixed in Bouin's solution. The occurrence of apoptosis was examined by end labeling of DNA with (alpha-32P)dideoxyATP and electrophoretic fractionation and by end labeling of DNA in situ with digoxigenin-dideoxyUTP. The expression of Bcl-2 and Bax proteins was examined by immunohistochemical staining with appropriate antibodies. RESULTS: Autoradiographic analysis revealed that high molecular weight DNA was predominant in the normal cervical epithelium (NCE) and in ISCC. However, a ladder-like pattern of DNA fragments, characteristic of the apoptotic breakdown of DNA, was identified in IEAC. Quantitative analysis of low molecular weight fragments of DNA revealed a significant increase in IEAC but not in ISCC compared with NCE. Labeling of DNA in situ indicated that cells undergoing apoptosis were predominant among the neoplastic cells of IEAC. However, no apoptotic cells were noted in ISCC, with the exception of cells in some tumor nests. A large fraction of IEAC and ISCC was immunonegative for Bcl-2. Although the expression of Bax was detected weakly in a small fraction of ISCC, strong expression of Bax was observed in all cases of IEAC. CONCLUSIONS: Apoptosis appears to occur in the cancerous cells of invasive adenocarcinoma of the uterine cervix in association with a high level of expression of Bax but not of bcl-2.