胶质瘤来源exosome的鉴定及其蛋白质组成研究

目的初步分析胶质瘤细胞分泌的exosome蛋白组成,探讨胶质瘤来源exosome的潜在免疫调节功能,从而为进一步利用exosome对胶质瘤进行免疫治疗提供理论依据。方法采用差速离心法从U251胶质瘤细胞培养上清液和Ⅲ级星形胶质瘤囊液中分别提纯exosome,用透射电镜鉴定;利用二维电泳分离、分析exosome内蛋白质,并用质谱技术鉴定了部分蛋白质。结果胶质瘤细胞可以产生exosome,其平均直径约100 nm。二维电泳图显示U251细胞分泌的exosome含有270个蛋白点,与数据库相符的有66个;而来自Ⅲ级星形胶质瘤囊液的exosome含有242个蛋白点,与数据库相符的有60个;两者有130个蛋白点在等电点和表观分子量方面相同,其中包含HSP70、RNA结合蛋白、核酸外切酶、MHCI及MHCII类分子等。部分蛋白质点质谱鉴定结果为hCG、低密度脂蛋白、T细胞受体等。结论胶质瘤细胞可分泌exosome,其一般特性与已报道的exosome一致,其蛋白组成与其他细胞来源的exosome具有共性,体内和体外培养的胶质瘤细胞分泌的exosome的蛋白质组成具有同源性与差异性。胶质瘤细胞源的exosome具有一定的免疫调节功能,可以为胶质瘤免疫治疗提供理论基础。     

胶质瘤细胞来源exosome的鉴定及其蛋白组成研究

目的证实胶质瘤细胞是否可分泌exosome，并初步分析其蛋白组成，探讨胶质瘤来源exosome的潜在免疫调节功能，从而为进一步利用exosome对胶质瘤行免疫治疗提供理论依据。方法差速离心法从胶质瘤细胞培养上清液中提纯exosome，用透射电镜鉴定，利用SDS-PAGE初步分析exosome蛋白组成，并用western-blot验证hsp70的存在。结果胶质瘤细胞可以产生大小均一的囊泡性结构，即exosome，其平均直径约100nm。exosome内所含蛋白质经初步分析，其表观分子量介于270—30kD之间。经exosome-blot证实其内含有hsp70分子。结论胶质瘤细胞也可分泌exosome，其一般特性与已报道的exosome一致，蛋白组成与其他细胞来源的exosome具有共性，且含有hsp70分子，具有一定的免疫调节功能，可以为抗胶质瘤免疫治疗提供理论基础。     

树突状细胞负载的肿瘤抗原肽疫苗抗脑胶质瘤作用的实验研究

目的探讨以树突状细胞（dendritic cells,DC）作为载体,将由脑胶质瘤细胞提取的热休克蛋白．肿瘤肽复合物（heat shock protein-tumor peptidecomplex,HSP-PC）负载到DC上所制备的肿瘤疫苗对脑胶质瘤细胞U251的杀伤作用。方法用脐带血在体外培养诱导DC,用脑胶质瘤患者术后标本提取肿瘤混合抗原并与DC共同培养,检测其对U251细胞的杀伤活性。结果经抗原负载后的DC对U251细胞的杀伤能力明显高于单纯DC对肿瘤细胞的杀伤能力。结论用肿瘤抗原负载的DC经体外实验证实具有明显的抗肿瘤效应,为临床肿瘤免疫治疗的开展提供了有效的数据。     

顺铂联合血管内皮抑制素对体外培养的U251人胶质瘤细胞生长的影响

目的以体外培养的U251人胶质瘤细胞作为模型,观察顺铂联合血管内皮抑制素对肿瘤细胞生长的影响。方法采用不同浓度的顺铂和血管内皮抑制素及顺铂与血管内皮抑制素的组合作用于体外培养的U251人胶质瘤细胞,以MTT比色法检测其对肿瘤细胞生长的影响。结果不同浓度的顺铂与血管内皮抑制素联合应用对体外培养的U251人胶质瘤细胞生长具有非常明显的协同抑制作用,强于单独使用顺铂或血管内皮抑制素组。结论顺铂联合血管内皮抑制素对体外培养的U251人胶质瘤细胞具有协同杀伤作用。     

青蒿素对体外培养人U251细胞凋亡和增殖的作用

目的:探讨青蒿素对体外培养的人U251胶质瘤细胞株是否具有诱导凋亡的作用。方法:(1)体外培养的U251细胞培养液中按10μg/mL终浓度加入青蒿素培养3d后进行实验;(2)倒置显微镜下观察U251细胞形态;(3)用台盼蓝拒染色法进行U251细胞的活细胞计数;(4)用MTT比色法检测青蒿素对U251细胞的抑制率;(5)透射电镜下观察U251细胞的形态。结果:(1)光镜下U251细胞部分出现碎裂崩解;(2)台盼蓝法U251细胞的活细胞计数青蒿素组少于对照组(7.6±1.273vs16.7±1.593,P<0.05);(3)MTT比色法检测青蒿素对U251细胞的抑制率为(50.0±3.2)%;(4)透射电镜下U251细胞的亚细胞结构发生凋亡变化。结论:青蒿素对体外培养的人U251胶质瘤细胞株具有诱导凋亡和抑制增殖的作用。     

特定沉默胶质瘤U251细胞株的PDGF-B基因对细胞凋亡和增殖的影响研究

目的利用RNA干扰基因治疗技术,特定沉默胶质瘤U251细胞株的血小板衍生生长因子B(PDGF-B)基因,观察其对U251细胞株细胞凋亡和增殖的影响。方法用血清培养液(含10%胎牛血清DMEM培养基)体外培养人U251脑胶质瘤细胞系。随机分为3组,第1组为实验组:转染siRNA的细胞,第2组为阴性对照组:转染空白质粒的细胞;第3组为空白对照组:细胞常规培养不予以干预。利用脂质体将siRNA转染进入U251细胞,在细胞内形成双链siRNA,识别并降解PDGF-B mRNA。通过RT-PCR检测U251细胞中PDGF-B基因mRNA的表达水平,Western blot检测PDGF-B的蛋白表达,MTT法检测U251细胞的增殖情况,流式细胞学对比检测3组细胞的凋亡情况。结果利用脂质体将靶向PDGF-B基因siRNA转染进入U251细胞,48 h后利用RT-PCR检测PDGF-B基因mRNA的表达,结果显示与阴性对照组和空白对照组对比,转染组PDGF-B基因mRNA表达水平明显下降;转染72 h后,Western blot检测显示siRNA转染组PDGF-B蛋白表达明显抑制(抑制率60%);MTT结果显示siRNA转染组U25细胞增殖较对照组明显降低;流式细胞学检测显示,与阴性对照组和空对照组相比,转染siRNA的细胞的实验组中,G1期细胞明显增多,S期细胞明显减少,提示降低PDGF-B在胶质瘤细胞的表达,能够使细胞周期阻滞在G1期,从而抑制胶质瘤细胞的增殖,并促进细胞的凋亡。结论构建靶向胶质瘤细胞PDGF-B的RNA干扰质粒并转染人胶质瘤U251细胞株后,通过体外试验观察可明显抑制U251细胞株PDGF的表达,其所产生的RNA干扰效应对人胶质瘤U251细胞株有明显的生长的抑制和促进凋亡作用。     

中国人脑星形胶质细胞瘤中表皮生长因子受体的表达及其靶向治疗

背景越来越多恶性脑胶质瘤存在的基因缺陷被发现,恶性脑胶质瘤的基因治疗逐渐成为热点,胶质瘤的治疗可以靶向于脑胶质瘤的基因缺陷.目的研究中国人脑星形胶质细胞瘤中表皮生长因子受体(epidermalgrowthfactor receptor,EGFR)的表达,并探讨其临床意义.设计以金标准为依据的诊断性试验的病例观察性研究.地点、对象和干预人脑星形胶质细胞瘤新鲜手术标本37例,其中男22例,女15例,年龄11～69岁,平均37岁,均来自于1998/2000长征医院手术病例.样本包含术中所切除肿瘤组织及配对瘤周组织,按照Kernohan病理分级,分为低级别组(Ⅰ～Ⅱ级,12例)和高级别组(Ⅲ～Ⅳ级,25例).人脑胶质瘤细胞株U87MG,U251MG,大鼠脑胶质瘤细胞株C6.应用免疫组化方法(Evision-HRP法)检测人脑星形胶质瘤标本(含配对瘤周组织)和体外培养的多株人和大鼠胶质瘤细胞系的EGFR表达.主要观察指标EGFR在人星形细胞瘤、脑胶质瘤细胞株中的表达;EGFR表达与肿瘤病理分极的相关性.结果人脑星形胶质瘤组织EGFR阳性率为70%(26/37),明显高于瘤周组织(32%,12/37),差异有显著性意义(x2=10.602,P＜0.01).Ⅲ～Ⅳ级肿瘤EGFR阳性率为84%(21/25),Ⅰ～Ⅱ级为42%(5/12),高级别组肿瘤EGFR表达水平显著低于低级别组,差异有显著性意义.(x2=6.955,P＜0.01),体外实验显示不同来源细胞系间的EGFR表达率差异存在显著性意义.结论中国人脑星形胶质瘤存在EGFR的过度表达,提示EGFR可作为恶性脑胶质瘤细胞的标志物.为基因治疗提供了适宜的靶向,为研究EGFR介导的脑胶质瘤靶向性基因治疗建立基础.     

蒿甲醚对人恶性胶质瘤细胞的抑制作用

目的 探讨蒿甲醚对U251人胶质瘤细胞增殖活性、凋亡的影响及其机制.方法 将浓度为0、50、100、200、400、600及800 μmol/L的蒿甲醚孵育U251胶质瘤细胞24、48 h.利用MTT法测定细胞增殖活性.然后将利用浓度为0、200、400及800μmol/L的蒿甲醚培养U251人胶质瘤细胞48 h,利用Annexin V-FITC/PI法检测细胞凋亡.最后利用Western blot法检测胶质瘤细胞的磷酸化AKt的表达水平.结果 400 μmol/L及更高浓度的蒿甲醚培养48 h,U251细胞的增殖活性受到显著抑制,同时随着浓度升高抑制作用增强.与对照组相比,蒿甲醚显著促进U251细胞的凋亡.400 μmol/L和800 μmol/L浓度的蒿甲醚显著抑制了磷酸化Akt的表达水平.结论 蒿甲醚能够以剂量依赖的方式显著抑制U251细胞的增殖活性并促进其凋亡,该抑制作用可能与其抑制Akt信号通路有关.     

中国人脑星形胶质细胞瘤中表皮生长因子受体的表达及其靶向治疗

背景：越来越多恶性脑胶质瘤存在的基因缺陷被发现。恶性脑胶质瘤的基因治疗逐渐成为热点，胶质瘤的治疗可以靶向于脑胶质瘤的基因缺陷。目的：研究中国人脑星形胶质细胞瘤中表皮生长因子受体(epidermal growth factor receptor，EGFR)的表达，并探讨其临床意义。设计：以金标准为依据的诊断性试验的病例观察性研究。地点、对象和干预：人脑星形胶质细胞瘤新鲜手术标本37例，其中男22例。女15例，年龄11～69岁，平均37岁，均来自于：1998／2000长征医院手术病例。样本包含术中所切除肿瘤组织及配对瘤周组织，按照Kernohan病理分级，分为低级别组(Ⅰ～Ⅱ级，12例)和高级别组(Ⅲ～Ⅳ级。25例)。人脑胶质瘤细胞株U87MG，U251MG，大鼠脑胶质瘤细胞株C6。应用免疫组化方法(Evision-HRP法)检测人脑星形胶质瘤标本(含配对瘤周组织)和体外培养的多株人和大鼠胶质瘤细胞系的EGFR表达。主要观察指标：EGFR在人星形细胞瘤、脑胶质瘤细胞株中的表达；EGFR表达与肿瘤病理分极的相关性。结果：人脑星形胶质瘤组织EGFR阳性率为70％(26／37)，明显高于瘤周组织(32％，12／37)，差异有显著性意义(χ^2=10．602，P＜0．01)。Ⅲ～Ⅳ级肿瘤：EGFR阳性率为84％(21／25)，Ⅰ～Ⅱ级为42％(5／12)，高级别组肿瘤：EGFR表达水平显著低于低级别组，差异有显著性意义。(χ^2=6．955，P＜0．01)，体外实验显示不同来源细胞系间的EGFR表达率差异存在显著性意义。结论：中国人脑星形胶质瘤存在EGFR的过度表达，提示EGFR可作为恶性脑胶质瘤细胞的标志物。为基因治疗提供了适宜的靶向，为研究EGFR介导的脑胶质瘤靶向性基因治疗建立基础。     

MiR-195对人脑胶质瘤细胞 U251和 SHG-44增殖的抑制作用

目的探讨 miR-195过表达对体外培养的人脑胶质瘤细胞U251和 SHG-44增殖的影响。方法通过脂质体将miR-195 mimics转染至U251和SHG-44细胞,同时设立空白对照组和阴性对照组,实时荧光定量PCR检测转染前后miR-195的表达水平;用CCK-8法评价细胞的增殖能力;流式细胞仪分析细胞周期。结果胶质瘤细胞转染miR-195 mimics后,发现U251细胞和SHG-44细胞中miR-195的表达分别为5.93＆#177;0.43和6.43＆#177;0.48,较空白对照组和阴性对照组均增高约6倍左右。与空白对照组和阴性对照组相比,转染miR-195 mimics后的胶质瘤细胞的增殖能力明显受到抑制,细胞周期阻滞于G0/G1期,S期细胞减少。结论过表达miR-195可以阻滞胶质瘤细胞周期进展,从而抑制其增殖,为胶质瘤的基因治疗提供新的策略。     

Expression of CYP4A1 in U251 human glioma cell induces hyperproliferative phenotype in vitro and rapidly growing tumors in vivo

Exogenous 20-hydroxyeicosatetraenoic acid (20-HETE) increases the growth of human glioma cells in vitro. However, glioma cells in culture show negligible 20-HETE synthesis. We examined whether inducing the expression of a 20-HETE synthase in a human glioma U251 cell line would increase proliferation. U251 cells transfected with CYP4A1 cDNA (termed U251 O) increased the formation of 20-HETE from less than 1 to over 60 pmol/min/mg proteins and increased their proliferation rate by 2-fold (p < 0.01). Compared with control U251, U251 O cells were rounded, smaller, showed a disorganized cytoskeleton, exhibited reduced vinculin staining, and were easily detached from the growing surface. They showed a marked increase in dihydroethidium staining, suggesting increased oxidative stress. The expression of phosphorylated extracellular signal-regulated kinase 1/2, cyclin D1/2, and vascular endothelial growth factor was markedly elevated in U251 O. The hyperproliferative and signaling effects seen in U251 O cells are abolished by selective CYP4A inhibition of 20-HETE formation with HET0016 [N-hydroxy-N'-(4-butyl-2-methylphenyl)-formamidine], by small interfering RNA against the enzyme, and by the putative 20-HETE antagonist, 20-hydroxyeicosa-5(Z),14(Z)-dienoic acid. In vivo, implantation of U251O cells in the brain of nude rats resulted in a approximately 10-fold larger tumor volume (10 days postimplantation) compared with animals receiving mock-transfected U251 cells. These data show that elevations in 20-HETE synthesis in U251 cells lead to an increased growth both in vitro and in vivo. This suggests that 20-HETE may have proto-oncogenic properties in U251 human gliomas. Further studies are needed to determine whether 20-HETE plays a role promoting growth of some human gliomas.     

Inhibition of Akt by the alkylphospholipid perifosine does not enhance the radiosensitivity of human glioma cells

Akt has been implicated as a molecular determinant of cellular radiosensitivity. Because it is often constitutively activated or overexpressed in malignant gliomas, it has been suggested as a target for brain tumor radiosensitization. To evaluate the role of Akt in glioma radioresponse, we have determined the effects of perifosine, a clinically relevant alkylphospholipid that inhibits Akt activation, on the radiosensitivity of three human glioma cell lines (U87, U251, and LN229). Each of the glioma cell lines expressed clearly detectable levels of phosphorylated Akt indicative of constitutive Akt activity. Exposure to a perifosine concentration that reduced survival by approximately 50% significantly reduced the level of phosphorylated Akt as well as Akt activity. Cell survival analysis using a clonogenic assay, however, revealed that this Akt-inhibiting perifosine treatment did not enhance the radiosensitivity of the glioma cell lines. This evaluation was then extended to an in vivo model using U251 xenografts. Perifosine delivered to mice bearing U251 xenografts substantially reduced tumor phosphorylated Akt levels and inhibited tumor growth rate. However, the combination of perifosine and radiation resulted in a less than additive increase in tumor growth delay. Thus, in vitro and in vivo data indicate that the perifosine-mediated decrease in Akt activity does not enhance the radiosensitivity of three genetically disparate glioma cell lines. These results suggest that, although Akt may influence the radiosensitivity of other tumor types, it does not seem to be a target for glioma cell radiosensitization.     

氯化甲基汞-L-半胱氨酸共轭物诱导恶性脑胶质瘤凋亡的体外研究

目的探讨氯化甲基汞(methylmercury chloride,MMC)-L-半胱氨酸(L-cys)共轭物体外诱导恶性脑胶质瘤U251细胞凋亡并抑制其增殖的作用。方法采用四甲基偶氮唑蓝(MTT)比色法测定不同剂量的MMC-L-cys共轭物分别作用24、48、72h后对U251胶质瘤细胞增殖的影响,并与三氧化二砷(ATO)做平行对照研究。流式细胞仪用异硫氰酸荧光素标记的膜联蛋白V(Annexin V-FITC)和碘化丙啶(PI)双染检测凋亡情况。结果 MMC-L-cys共轭物抑制U251胶质瘤细胞的生长和增殖,与正常对照组相比差异显著(P<0.05),其抑制作用强于ATO组(P<0.05),抑制作用呈剂量、时间依赖性;U251胶质瘤经不同浓度的MMC-L-cys共轭物作用后,24h其凋亡率显著高于对照组(P<0.05),诱导凋亡作用具有剂量依赖关系。结论 MMC-L-cys共轭物能诱导胶质瘤细胞凋亡、抑制其增殖,呈时间剂量依赖性,为胶质瘤的治疗提供实验依据。     

NG2 expression regulates vascular morphology and function in human brain tumours

Tumour angiogenesis is a tightly regulated process involving cross-talk between tumour cells and the host tissue. The underlying mechanisms that regulate such interactions remain largely unknown. NG2 is a transmembrane proteoglycan whose presence on transformed cells has been demonstrated to increase proliferation in vitro and angiogenesis in vivo. To study the effects of NG2 during tumour growth and progression, we engineered an NG2 positive human glioma cell line (U251-NG2) from parental NG2 negative cells (U251-WT) and implanted both cell types stereotactically into immunodeficient nude rat brains. The tumours were longitudinally monitored in vivo using multispectral MRI employing two differently sized contrast agents (Gd-DTPA-BMA and Gadomer) to assess vascular leakiness, vasogenic oedema, tumour volumes and necrosis. Comparisons of Gd-DTPA-BMA and Gadomer revealed differences in their spatial distribution in the U251-NG2 and U251-WT tumours. The U251-NG2 tumours exhibited a higher leakiness of the larger molecular weight Gadomer and displayed a stronger vasogenic oedema (69.9 +/- 15.2, P = 0.018, compared to the controls (10.7 +/- 7.7). Moreover, immunohistochemistry and electron microscopy revealed that the U251-NG2 tumours had a higher microvascular density (11.81 +/- 0.54; P = 0.0010) compared to controls (5.76 +/- 0.87), with vessels that displayed larger gaps between the endothelial cells. Thus, tumour cells can regulate both the function and structure of the host-derived tumour vasculature through NG2 expression, suggesting a role for NG2 in the cross-talk between tumour-host compartments.     

Enhanced apoptosis following treatment with TRA-8 anti-human DR5 monoclonal antibody and overexpression of exogenous Bax in human glioma cells

Specific activation of apoptosis in tumor cells offers a promising approach for cancer therapy. Induction of apoptosis leads to activation of specific proteases. Two major pathways for caspase activation in mammalian cells have been described. One apoptotic pathway involves members of the tumor necrosis factor family of cytokine receptors (eg death receptor 5 (DR5)). The other pathway is controlled by the Bcl-2 family of proteins. The purpose of this study was to investigate whether increased apoptosis occurs in human glioma cells following infection with a recombinant adenoviral vector encoding the human Bax gene under the control of human vascular endothelial growth factor (VEGF) promoter element (AdVEGFBax) in combination with an anti-human DR5 monoclonal antibody (TRA-8). Specific overexpression of exogenous Bax protein induced apoptosis and cell death in glioma cell lines, through activation of both caspase-8 and -9, leading to activation of downstream caspase-3. The relative sensitivity to AdVEGFBax for the glioma cell lines was U251MG>U373MG>U87MG>D54MG. The recently characterized TRA-8 monoclonal antibody induces apoptosis of most TRAIL-sensitive tumor cells by specific binding to DR5 receptors on the cellular membrane. TRA-8 induced rapid apoptosis and cell death in glioma cells, but did not demonstrate detectable cytotoxicity of primary normal human astrocytes. The efficiency of TRA-8-induced apoptosis was variable in different glioma cell lines. The relative sensitivity to TRA-8 was U373MG>U87MG>U251MG>D54MG. The combination of TRA-8 treatment and overexpression of Bax overcame TRA-8 resistance of glioma cells in vitro. Cell viability of U251MG cells was 71.1% for TRA-8 (100 ng/ml) alone, 75.9% for AdVEGFBax (5 MOI) alone and 41.1% for their combination as measured by MTS assay. Similar enhanced apoptosis results were obtained for the other glioma cell lines. In vivo studies demonstrated that the combined treatment significantly (P<0.05) suppressed the growth of U251MG xenografts and produced 60% complete tumor regressions without recurrence. These data suggest that the combination of TRA-8 treatment with specific overexpression of Bax using AdVEGFBax may be an effective approach for the treatment of human malignant gliomas.     

儿茶素对脑胶质瘤细胞U251增殖的抑制作用及机制初探

目的:通过体外实验初步研究儿茶素对脑胶质瘤细胞U251增殖的抑制作用及其诱导凋亡的机制.方法:选取8个不同梯度浓度的儿茶素进行甲基噻唑蓝(MTT)实验,观察儿茶素的半致死浓度;倒置显微镜下观察细胞形态,台盼蓝拒染法及Hoechst33342荧光染色检测细胞凋亡,用流式细胞术检测儿茶素对细胞周期的影响.结果:儿茶素对U251细胞的半抑制浓度为62.25 μg/mL,儿茶素作用后U251细胞出现凋亡小体,阻滞于S期,Hoechst33342检测表明细胞凋亡明显.结论:儿茶素明显抑制U251细胞增殖,阻滞细胞周期于S期.儿茶素具有开发为治疗脑胶质瘤药物的可能性.     

NOB1影响胶质瘤细胞增殖、凋亡的实验研究

目的 利用RNA干扰（RNAi）在人胶质瘤U251和U87-MG细胞中沉默NOB1基因的表达,探讨NOB1对恶性胶质瘤细胞增殖以及凋亡的影响.方法 构建NOB1基因的短发卡（shRNA）慢病毒表达载体,包装成病毒颗粒并感染人胶质瘤U251和U87-MG细胞,采用实时定量聚合酶链反应检测NOB1在恶性胶质瘤细胞系中的表达.采用甲基噻唑基四唑（MTT）比色法和克隆形成实验检测细胞增殖和克隆形成能力情况;同时利用PI染色流式细胞仪检测细胞凋亡情况,观察NOB1基因对U251和U87-MG细胞增殖、凋亡的影响.结果 NOB1基因在人胶质瘤U251、U87-MG细胞系中均明显高表达.NOB1-shRNA慢病毒能有效感染胶质瘤细胞,实验结果显示感染后U251和U87-MG细胞的增殖能力明显下降;U251克隆形成能力明显受到抑制;细胞周期在G0/G1停滞,而且细胞出现显著性凋亡;上述差异均具有统计学意义（P＜0.05）.结论 NOB1在人胶质瘤U251和U87-MG细胞中高表达;降低NOB1基因的表达,可以显著降低胶质瘤细胞的增殖,并促进胶质瘤细胞凋亡;NOB1基因在体外对胶质瘤细胞的发生发展可能具有癌基因的调节作用.     

Adenovirus-mediated transfer of caspase-8 augments cell death in gliomas: implication for gene therapy

Caspase-8 is a member of the family of caspases, which are involved in the execution of apoptosis. To investigate whether caspase-8 can be used for gene therapy of gliomas, we transduced A-172 and U251 glioma cells with the caspase-8 gene via an adenoviral vector (Adv) controlled by the chicken beta-actin (CA) promoter (Advcaspase-8), and found that a similar level of caspase-8 protein induced A-172 cells to undergo necrotic cell death and U251 cells to undergo apoptotic cell death. Neither Bcl-XL nor Bcl-2, which play important roles in antiapoptotic mechanisms in gliomas, protected glioma cells from apoptosis induced by overexpression of caspase-8. Injection of Adv-caspase-8 suppressed the in vivo growth of U251 xenografts, in which apoptotic cell death remarkably increased as revealed by TUNEL analysis. Finally, we assessed whether gene therapy with a tissue-specific promoter, the myelin basic protein (MBP) promoter, is applicable to gliomas. Adv for caspase-8 controlled by the MBP promoter induced drastic apoptosis in U251 and U-373MG glioma cells, whereas it did not induce apoptosis in human endothelial cells, fibroblasts, and nerve growth factor-treated PC12 cells. These results indicate that Adv for caspase-8 effectively induced cell death in gliomas, and that this approach may be a useful modality for gene therapy of gliomas.