Caspase-8和DR5在TRAIL诱导神经母细胞瘤细胞凋亡中的作用

目的：肿瘤坏死因子相关凋亡诱导配体(TRAIL)能诱导多种肿瘤细胞凋亡而对正常细胞无诱导凋亡作用。大多数神经母细胞瘤细胞株对TRAIL的诱导凋亡作用耐受与其Caspase8表达缺失有关,也与细胞表面TRAIL受体的表达和分布有关。该文主要探讨Caspase8及TRAIL受体DR5的表达在TRAIL诱导神经母细胞瘤细胞株SKNDZ凋亡中的作用及其发生机制。方法：应用RTPCR方法检测IFNγ作用前后SKNDZ细胞Caspase8的表达;应用WesternBlot方法检测化疗药作用前后SKNDZ细胞DR5的表达;应用四甲基偶氮唑蓝(MTT)比色法及流式细胞仪(FCM)检测TRAIL、IFNγ+TRAIL、化疗药+TRAIL及化疗药+IFNγ+TRAIL对SKNDZ细胞生长及凋亡的影响。结果：SKNDZ细胞不表达Caspase8,IFNγ作用后的SKNDZ细胞Caspase8表达明显增加。对照组未检测到DR5蛋白表达,而阿霉素和依托泊苷处理后检测到DR5蛋白表达。表达Caspase8的SKNDZ细胞对TRAIL的诱导凋亡作用仍不敏感,而同时表达Caspase8和DR5的SKNDZ细胞对TRAIL的诱导凋亡作用敏感。阿霉素/依托泊苷+IFNγ+TRAIL组早期凋亡率为:(17.9±3.6)%、(14.8±3.3)%,与IFNγ+TRAIL组(3.9±1.2)%比较,差异有显著性(F=26.233,P<0.01)。结论：同时表达Caspase8和DR5的SKNDZ细胞恢复了对TRAIL的敏感性,Caspase8和DR5在TRAIL诱导SKNDZ细胞凋亡中起着十分关键的作用。     

MYCN基因表达抑制促TRAIL诱导神经母细胞瘤细胞凋亡的研究

目的神经母细胞瘤(NB)为小儿常见恶性肿瘤。MYCN基因高扩增NB细胞对TRAIL诱导凋亡多有抵抗。我们以MYCN基因高扩增NB细胞株为研究对象,抑制其MYCN基因的表达,探讨其对TRAIL诱导凋亡有无增强作用。方法构建MYCN siRNA,转染NB细胞株SK-N-BE(2),抑制MYCN基因表达后,Western-blot法检测SK-N-BE(2)细胞Bcl-2、Bcl-xL、Bid、DR4、DR5表达的变化,ELISA法检测细胞凋亡。结果转染MYCN siRNA的SK-N-BE(2)细胞中MY-CN基因表达显著降低(P<0.05)。MYCN基因表达抑制后,SK-N-BE(2)细胞DR5、Bid表达增高(P<0.05),Bcl-2、Bcl-xL表达降低(P<0.05),DR4表达无变化(P>0.05),TRAIL诱导细胞凋亡显著增强(P<0.05)。结论 MYCN基因表达抑制后,可调控Bcl家族中相关蛋白、DR5的表达,促进TRAIL诱导NB细胞凋亡。     

颅咽管瘤中肿瘤坏死因子相关凋亡诱导配体受体的表达及意义

目的　研究肿瘤坏死因子相关凋亡诱导配体受体 (tumornecorisisfactorrelatedapoptosisinducedligandreceptor,TRAILR)在颅咽管瘤细胞中的表达 ,探讨其临床意义。方法　采用免疫组化方法检测颅咽管瘤细胞及正常脑组织中TRAILR蛋白的表达。同时 ,采用原位杂交方法检测颅咽管瘤中诱骗受体 (Decoyreceptor,DcR)在mRNA水平的表达。 结果　免疫组化染色显示 ,2 4例颅咽管瘤均大量表达死亡受体 (Deathreceptor,DR)DR4和DR5 ,7例 (2 9.2 % )表达DcR1 ,5例 (2 0 .8% )表达DcR2 ,而 1 6例正常脑组织普遍表达DcR ,7例 (43 .8% )表达DR4 ,5例 (31 .3 % )表达DR5。颅咽管瘤组织中DR的高表达以及DcR的低表达不同于正常脑组织中DR的低表达及DcR的高表达 ,两者间有显著差异 (P <0 .0 1 )。原位杂交显示 2 4例颅咽管瘤组织分别有 1 9例 (79.2 % )DcR1和 1 5例 (62 .5 % )DcR2在mRNA水平呈强阳性表达 ,DcR在转录水平的表达明显高于翻译水平 (P <0 .0 1 )。结论　颅咽管瘤细胞中普遍存在DR的高表达和DcR的低表达 ,DcR在颅咽管瘤中限制性表达的调控位于转录后水平。这可能为颅咽管瘤的凋亡诱导治疗提供新的靶点和新的策略。     

Resistance to TRAIL-induced apoptosis in neuroblastoma cells correlates with a loss of caspase-8 expression

BACKGROUND: Disruption of apoptotic pathways may be involved in tumor formation, regression, and treatment resistance of neuroblastoma (NB). TNF-related apoptosis-inducing ligand (TRAIL) is a potent inducer of apoptosis in cancer cell lines. PROCEDURE: In this study we analyzed the expression and function of TRAIL, its agonistic and antagonistic receptors, and important intracellular signaling elements in 18 NB cell lines. RESULTS: Semiquantitative RT-PCR revealed that TRAIL-R2 and TRAIL-R3 are the main TRAIL-receptors used by NB cells. Sensitivity to TRAIL-induced apoptosis did not correlate with mRNA expression of TRAIL receptors or cFLIP. Surprisingly, caspase-8 and caspase-10 mRNA was detected in only 5 of 18 NB cell lines. Interestingly, only these five NB cell lines were susceptible to TRAIL-induced apoptosis in a time- and dose-dependent manner. CONCLUSIONS: Treatment with 5-aza-2'-deoxycytidine restored mRNA expression of caspase-8 and -10 and TRAIL sensitivity of resistant cell lines, suggesting that gene methylation is involved in caspase inactivation. Since many cytotoxic drugs induce caspase-dependent apoptosis, failure to express caspase-8 and/or caspase-10 might be an important mechanism of resistance to chemotherapy in NB.     

阿霉素诱导Jurkat白血病细胞凋亡及上调Fas配体和Fas相关死亡域

目的研究阿霉素诱导白血病细胞凋亡的剂量及时间关系,探索其相关的分子机制。方法分别以0.1、0.2、0.5、1.0 mg/L的阿霉素处理人类Jurkat白血病细胞61、22、44、8 h。其中一份样本在加入0.2 mg/L阿霉素前用zVAD-fmk(苄氧羰-缬氨酰-丙氨酰-天冬氨酰-氟甲基酮)预处理。应用AnV/PI双染细胞,在流式细胞仪上分析AnV/PI双阳性的凋亡细胞。采用Western Blot技术检测FasL和FADD(Fas相关死亡域)的表达。结果6 h时所有剂量的阿霉素诱导的凋亡细胞无显著差异(P>0.05),在12 h,只有1.0 mg/L诱导细胞明显凋亡。当细胞与0.2和0.5 mg/L的阿霉素共同培养24 h或36 h,观察到调亡细胞显著增加(P<0.001)。在zVAD-fmk存在的情况下,当细胞与阿霉素一同培养时,由阿霉素诱导的细胞凋亡完全受到抑制(P<0.001)。随着阿霉素作用时间增加,FasL和FADD表达水平相应增加。结论在阿霉素诱导的白血病细胞凋亡中,阿霉素以剂量和时间依赖方式诱导细胞凋亡;上调FasL可能启动FasL信号通路的激话,而caspase是最终的执行者。     

阿霉素诱导Jurkat白血病细胞凋亡及上调Fas协配体和Fas相关死亡域

目的研究阿霉素诱导门血病细胞凋亡的剂量及时间关系，探索其相关的分子机制。方法分别以0．1、0．2、0．5、10mg／L的阿霉素处理人类Jurkat白血病细胞6、12、24、48h。其中一份样本在加入0．2mg／L阿霉素前用zVAD-fmk（苄氧羰-撷氨酰丙氨酰天冬氨酰氟甲基酮）颅处理。应用AnV／PI双染细胞，在流式细胞仪上分析AnV／PI双阳性的凋亡细胞。采用WGstem Blot技术检测FasL和FADD（Fas相关死亡域）的表达。结果6h时所有剂量的阿霉素诱导的凋亡细胞无显著差异（P〉0．05）．在12h，只有10mg／L诱导细胞明显凋亡。当细胞与0.2和0.5mg／L的阿霉素共同培养24h或36h，观察到调亡细胞显著增加（P〈0．001）。在zVAD-fmk存住的情况下。当细胞与阿霉素一同培养时，由阿霉素诱导的细胞凋亡完全受到抑制（P〈0.001）。随着阿霉索作用时间增加，FasL和FADD表达水平相应增加。结论在阿霉素诱导的白血病细胞凋亡中，阿霉素以剂量和时间依赖方式诱导细胞凋亡；上渊FasL可能启动FasL信号通路的激话．而caspase是最终的执行者。     

Loss of caspase-8 expression in neuroblastoma is related to malignancy and resistance to TRAIL-induced apoptosis

Background and Procedures NB-derived cell lines were tested for their sensitivity to apoptosis induced by the tumor-selective apoptotic ligand TRAIL. Noninvasive S-type cell lines are highly sensitive to TRAIL, whereas invasive N-type cell lines are resistant. RESULTS: Although both S- and N-type cell lines express TRAIL-R2, FADD, and caspases-3 and -10, only S-type cells express caspase-8. Reduced levels of caspase-8 protein were also observed in a stage IV NB tumor when compared to a ganglioneuroma. The caspase-8 gene is not deleted in either N-type NB cell lines or high-stage tumors, and expression can be induced by demethylation. CONCLUSIONS: Therefore, caspase-8 expression is silenced in malignant NB, which correlates to tumor severity and resistance to TRAIL-induced apoptosis.     

Interferon-gamma sensitizes osteosarcoma cells to Fas-induced apoptosis by up-regulating Fas receptors and caspase-8

BACKGROUND: Osteosarcoma is the third most frequent neoplasm in adolescents. Although chemotherapy, frequently used in pre- and post-operative settings, has resulted in significant improvement in disease-free survival, some patients show little sensitivity to chemotherapy and alternative therapeutic strategies are needed. Because the Fas ligand/Fas receptor (CD95, APO-1) apoptosis pathway is a potential therapeutic target in osteosarcomas, we examined the effect of IFN-gamma on Fas-induced apoptosis in four osteosarcoma cell lines. PROCEDURE AND RESULTS: As measured by flow cytometry, all cell lines expressed cell surface IFN-gamma receptors, and when cultured for 2 days in the presence of IFN-gamma, all cell lines exhibited a significant increase in expression of Fas receptors. By flow cytometric detection of intracellular fragmented DNA as a marker of apoptosis, all cell lines cultured with either IFN-gamma or anti-Fas antibody (clone CH-11) alone showed only moderate apoptosis, whereas significantly high levels of apoptosis occurred in cells cultured with both IFN-gamma and CH-11. Western blotting analysis also revealed that IFN-gamma caused up-regulation of caspase-8 in all cell lines, but no change in Fas-associated death domain protein (FADD/MORT1) or caspase-3. Both caspase-8 and caspase-3 were activated when apoptosis was induced with both IFN-gamma and CH-11. Addition to cultures of z-IETD-fmk, an inhibitor of caspase-8, significantly blocked this apoptosis. CONCLUSIONS: IFN-gamma sensitizes osteosarcoma cells to Fas-induced apoptosis through up-regulation of Fas receptor and caspase-8. Combined immunotherapy with IFN-gamma and either anti-Fas monoclonal antibody or cytotoxic T cells that bear Fas ligand might be a useful adjunctive therapy for patients with osteosarcoma.     

ROLE OF CASPASE 8 AS A DETERMINANT IN TRAIL SENSITIVITY OF NEUROBLASTOMA CELL LINES

Objective: Resistance of neuroblastoma (NB) cells to tumor necrosis factor-related apoptosis inducing ligand (TRAIL)-mediated apoptosis is thought to be caused by loss of caspase 8 expression. The aim of this study is to investigate if induction of caspase 8 by γ-interferon (IFNγ) renders NB cells sensitive to TRAIL. Methods: Caspase 8 expression was monitored by RT-PCR and Western blot analysis. The effects of IFNγ, TRAIL, IFNγ + TRAIL, and caspase 8 inhibitor + TRAIL on the growth and apoptosis of NB cells were detected with the methods of reduction rate of Alamar blue assay and flow cytometry. The relative caspase 8 activity was measured with colorimetric assay. Results: An increased expression of caspase 8 mRNA and protein was found after treatment with IFNγ. IFNγ in combination with TRAIL decreased proliferation of NB cell line SH-SY5Y cells. The killing effect of TRAIL on NB cells expressing caspase 8 was depressed by caspase 8 inhibitor. The relative caspase 8 activity of NB cells expressing caspase 8 increased with the prolongation of TRAIL action time. Conclusions: TRAIL induced apoptosis in NB cells expressing caspase 8.     

TRAIL及其受体DR5在病毒性心肌炎小鼠心肌组织中的表达和意义

目的 研究肿瘤坏死因子相关的凋亡诱导配体(TRAIL)及其死亡受体5(DR5)在病毒性心肌炎小鼠心肌组织中病变不同时期的表达及与细胞凋亡的关系.方法 建立病毒性心肌炎小鼠模型,于注射病毒后第7、10、14、21、28天取心肌组织,HE染色做心肌病理积分,TUNEL法检测凋亡细胞百分率,免疫组织化学法和逆转录-聚合酶链反应(RT-PCR)检测心肌组织中TRAIL和DR5蛋白及mRNA的表达.结果 病毒感染组小鼠和正常对照组小鼠心肌组织中均存在TRAIL和DR5蛋白和mRNA的表达.病毒感染后第14天,病毒感染组小鼠心肌组织中TRAIL蛋白表达高于同期正常对照组(P<0.05)和第7天病毒感染组(P<0.05),组间比较差异有统计学意义(F=9.17,P<0.01).病毒感染后第10、14、21天,病毒感染组小鼠DR5蛋白表达高于同期正常对照组(P<0.01)和第7天病毒感染组(P<0.01),组间比较差异有统计学意义(F=13.32,P<0.01).且TRAIL和DR5蛋白的表达与心肌病理积分、心肌细胞凋亡率密切相关(P<0.05).病毒感染组第10天TRAIL mRNA表达高于同期正常对照组(P<0.01)和第7天病毒感染组(P<0.01),组间比较差异有统计学意义(F=10.86,P<0.01).病毒感染组第10、14天DR5 mRNA表达高于同期正常对照组(P<0.01)和第7天病毒感染组(P<0.01),组间比较差异有统计学意义(F=22.75,P<0.01).结论 病毒性心肌炎小鼠发病中期,TRAIL及其受体DR5在心肌组织中出现高表达,且与病理积分、心肌细胞凋亡率成正相关,TRAIL及其受体DR5可能通过调控心肌细胞凋亡参与病毒性心肌炎的病理过程.     

TRAIL蛋白联合顺铂抑制裸鼠横纹肌肉瘤生长的实验研究

目的 探讨肿瘤坏死因子相关凋亡诱导配体(TRAIL)蛋白及联合顺铂对RD人横纹肌肉瘤裸鼠移植瘤生长的影响及其可能的作用机制.方法 将RD人横纹肌肉瘤细胞制成细胞悬液后接种于裸鼠皮下,成瘤后将裸鼠随机分4组:生理盐水组、TRAIL组、顺铂组及两药联合组.检测裸鼠的重量和体积;测定裸鼠肝功能、尿素氮及肌酐;另取肝肾组织进行切片,HE染色观察;用FCM方法检测肿瘤组织Fas表达;用RT-PCR检测DR4和DR5mRNA的表达.结果 各组裸鼠实验前后的体重变化无明显差异;DDP组和TRAIL+DDP组裸鼠的血清丙氨酸转氨酶高于生理盐水组,DDP组与TRAIL+DDP组裸鼠尿素氮和肌酐值比较,无明显差异;TRAIL组、DDP组和TRAIL+DDP组移植瘤的重量均明显降低;各组肝肾组织切片观察未见明显改变;TRAIL+DDP组Fas表达水平高于DDP组和TRAIL组;DDP组和TRAIL+DDP组裸鼠移植瘤细胞的DR5mRNA、DR4mRNA表达水平明显升高,而TRAIL组没有明显改变.结论 TRAIL和顺铂对裸鼠移植瘤的生长有抑制作用,联用有协同作用.     

TRAIL及其在白血病治疗中的研究进展

肿瘤坏死因子相关凋亡诱导配体(TRAIL)是近几年研究较热的抗肿瘤新型生物制剂,通过其死亡受体诱导肿瘤细胞凋亡,而对正常细胞无毒性,且与化疗药物具有协同性,但也存在抵抗机制.该文就TRAIL的生物学特点及其受体、TRAIL诱导肿瘤细胞凋亡的机制以及TRAIL在白血病治疗中的进展作一综述.     

MycN sensitizes neuroblastoma cells for drug-triggered apoptosis

BACKGROUND: Amplification of the MYCN gene is found in a large proportion of neuroblastomas and is associated with a poor prognosis. PROCEDURE: To investigate the effect of ectopic MycN expression on the susceptibility of neuroblastoma cells to cytotoxic drugs, we used a human neuroblastoma cell line with tetracycline-controlled expression of MycN. RESULTS: Neither conditional expression of MycN alone nor low drug concentrations induced apoptosis. However, MycN and cytotoxic drugs cooperated to induce cell death. Apoptosis triggered by MycN and doxorubicin was mediated by cleavage of caspases and involved activation of the CD95 system. MycN overexpression and cytotoxic drugs also synergized to induce p53 and Bax protein expression and to trigger mitochondrial permeability transition and cytochrome c release. CONCLUSION: In that amplification of MYCN is considered an adverse prognostic factor, these findings suggest that dysfunctions in apoptosis pathways may be a mechanism by which MycN-induced apoptosis of neuroblastoma cells is inhibited.     

紫铆因抗急性淋巴细胞白血病细胞的体外作用及机制研究

目的探讨紫铆因对急性淋巴细胞白血病(ALL)细胞增殖及细胞凋亡的体外作用及其机制。方法采用Ficoll-Paque分离液分离4例初发ALL患儿骨髓原代细胞及4例正常儿童骨髓单个核细胞并购买HK-2、Nalm-6、Jurkat、sup-B15及MOLT-3 ALL细胞株;用MTS技术检测紫铆因对细胞增殖的影响;采用流式细胞技术检测紫铆因对细胞凋亡的影响,并用蛋白印迹技术(Western Blot)检测cleaved caspase-3、Bim、Puma及cleaved PARP的表达。结果紫铆因对正常单个核细胞无明显抑制增殖作用,对人ALL原代细胞及ALL细胞株均具有明显的抑制增殖作用,并呈现出剂量效应和时间效应关系;紫铆因能逆转氟美松耐药细胞株对氟美松的敏感性;紫铆因能明显促进人ALL细胞株的细胞凋亡,并且随着紫铆因剂量的增加,细胞凋亡越明显;随着紫铆因浓度的增加,凋亡相关蛋白cleaved caspase-3、Bim、Puma及cleaved PARP表达量也逐渐增多。结论紫铆因对ALL细胞包括氟美松耐药细胞株具有抑制细胞增殖和促进细胞凋亡的作用。     

2-甲氧基雌二醇诱导K562细胞凋亡的机制研究

目的探讨2-甲氧基雌二醇(2-ME)对K562细胞的凋亡诱导效应及其作用机制。方法使用不同浓度2-ME(0,5,10,20,40μmol/L)作用于K562细胞,48 h后用流式细胞术检测2-ME作用后K562细胞的凋亡率的变化;用western blot方法检测2-ME作用48 h后K562细胞Bcl-2、Bax和Caspase-3蛋白的表达。结果 (1)2-ME在5~40μmol/L浓度范围内作用48 h对K562细胞有凋亡诱导作用,与对照组比较差异有显著性(P<0.05),这种作用呈剂量依赖关系;(2)2-ME作用48 h后,K562细胞Bcl-2蛋白的表达较对照组减少,差异有显著性(P<0.05),而Bax和Caspase-3表达较对照组增高,差异有显著性(P<0.05)。结论 2-ME可能通过线粒体途径诱导K562白血病细胞凋亡。     

PI3K/AKT通路在二烯丙基二硫诱导K562细胞凋亡中的作用机制研究

目的 研究PI3K/AKT 信号通路在二烯丙基二硫（DADS）诱导K562 细胞凋亡中的作用。方法 用 10、20、40、80 mg/L DADS 处理K562 细胞48 h,采用吖啶橙/溴化乙啶（AO/EB）染色法倒置显微镜观察细胞形态学变化;AnnexinV-FITC/PI 双染法检测细胞凋亡率;Western blot 技术检测AKT、p-AKT、Caspases-3 蛋白的表达,并设置空白对照组和溶媒对照组。结果 DADS 作用K562 细胞48 h 后,细胞出现体积缩小、形态不规则、胞膜突出等凋亡特征。AO/EB 染色显示：随DADS 浓度增加,胞体缩小、胞核呈固缩状或圆珠状,染色质凝集,核染色呈橘红色的凋亡细胞数增多,以40 mg/L DADS 组最明显。10、20、40、80 mg/L DADS 组的凋亡率均高于对照组（P P > 0.05）;随DADS 浓度增加,p-AKT 蛋白逐渐下降、Caspases 3 蛋白逐渐升高,差异有统计学意义（P 结论 DADS 诱导K562 细胞发生凋亡,其机制可能与DADS 抑制PI3K/AKT 信号通路的表达相关。     

5-氮杂胞苷增强阿霉素对神经母细胞瘤细胞的抗瘤活性

目的：许多神经母细胞瘤细胞系及肿瘤组织标本中caspase-8表达缺失, caspase-8基因沉寂的主要原因是其启动子区域高度甲基化。该文探讨去甲基化药物5-氮杂胞苷对caspase-8表达及对化疗药阿霉素抗人神经母细胞瘤细胞作用的影响及其影响机制。方法：应用RT-PCR方法检测5-氮杂胞苷作用前后SH-SY5Y细胞中caspase-8 mRNＡ水平变化。MTT分析研究5-氮杂胞苷与化疗药阿霉素联合应用前后人神经母细胞瘤细胞SH-SY5Y的存活率,以及加入caspase-8活性抑制剂后存活率的变化。结果：RT-PCR方法发现SH-SY5Y细胞株不表达caspase-8 mRNA, 5-氮杂胞苷作用3 d后可检测到 caspase-8 mRNA表达,5 d后表达较第3天增加; 5-氮杂胞苷与不同浓度阿霉素(0.05, 0.1, 0.25, 0.5 μg/mL)联合应用后SH-SY5Y细胞存活率为(77.61±7.30)%,（57.35±6.64）%,（46.25±4.46）%,（35.59±5.12）%,同一浓度单独应用阿霉素组细胞存活率为（94.89±4.15）%,（80.60±8.50）%,（64.48±4.92）%,（52.32±6.71）%,5-氮杂胞苷与阿霉素联用组细胞存活率明显低于单用阿霉素组,差异均有显著性。Caspase-8抑制剂组与同一浓度的5-氮杂胞苷+阿霉素组比较,细胞存活率明显增加,依次为（92.95±3.48）%,（78.39±4.28）%,（62.31±6.50）%,（49.92±5.77）%,差异均有显著性。结论：阿霉素对神经母细胞瘤细胞SH-SY5Y具有增殖抑制作用,5-氮杂胞苷可以增强阿霉素的抗瘤活性。其发生机制可能是通过上调caspase-8 mRNA的表达而实现的。［中国当代儿科杂志,2007,9（6）：577-579］     

Caspase-8在TRAIL诱导神经母细胞瘤细胞凋亡中的作用

目的：应用干扰素(IFNγ)诱导神经母细胞瘤（neuroblastoma, NB）细胞caspase 8的表达并观察其是否可以恢复NB细胞对肿瘤坏死因子相关凋亡诱导配体（TRAIL）的敏感性。方法：应用RT PCR方法检测IFNγ作用前后NB细胞caspase-8 mRNA的表达;应用Alamar Blue法及流式细胞术检测IFNγ、TRAIL、IFNγ+TRAIL、IFNγ+caspase-8抑制剂zIETD-FMK+TRAIL对NB细胞生长及凋亡的影响;应用比色法测定NB细胞caspase-8相对活性。结果：对TRAIL敏感的CHP212细胞表达caspase-8,且经IFNγ处理后caspase-8 表达水平逐步增加;对TRAIL不敏感的SY5Y细胞不表达caspase-8,IFNγ作用后其caspase-8 mRNA表达明显增加。IFNγ与TRAIL联用对SY5Y细胞有明显诱导凋亡作用。CHP212细胞caspase-8相对活性随TRAIL作用时间的延长逐步升高;IFNγ与TRAIL联合作用的SY5Y细胞caspase-8相对活性明显高于未加药物处理的对照组、IFNγ组、TRAIL组及抑制剂组。结论：表达caspase-8的NB细胞对TRAIL的诱导凋亡作用敏感,TRAIL诱导NB细胞凋亡过程中伴随caspase-8活性的增加。［中国当代儿科杂志,2010,12(11)：902-907］     

Serabelisib抗人肝母细胞瘤HepG2细胞的作用及其机制探讨

目的探讨小分子抑制剂Serabelisib抑制PI3K-Akt信号通路活性及对人肝母细胞瘤细胞系增殖、凋亡的效应作用及其相关机制。方法采用不同浓度的小分子抑制剂Serabelisib(0μmol/L、2μmol/L、4μmol/L、8μmol/L)处理人肝母细胞瘤细胞Hep G2细胞,通过四氮唑盐比色法检测Serabelisib对细胞增殖的抑制作用,采用流式细胞仪检测细胞凋亡变化情况,采用蛋白质免疫印迹检测Serabelisib处理后Hep G2细胞中PI3K信号通路活性及凋亡相关蛋白表达变化情况。结果当Serabelisib浓度≥4μmol/L时,对人肝母细胞瘤细胞系Hep G2细胞增殖具有显著抑制作用,随着Serabelisib浓度增加,其对Hep G2细胞增殖的抑制率逐渐增大,具有剂量和时间依赖性。随着Serabelisib浓度增加,Hep G2细胞凋亡率明显升高,同样具有剂量依赖性。蛋白质免疫印迹检测发现Serabelisib处理后的Hep G2细胞中PI3K信号通路中关键激酶Akt磷酸化水平降低,促凋亡蛋白PARP 89KD裂解片段表达增加。结论Serabelisib通过抑制PI3K/Akt信号转导通路,抑制人肝母细胞瘤Hep G2细胞增殖,并诱导凋亡。     

Arsenic trioxide inhibits cell growth in SH-SY5Y and SK-N-AS neuroblastoma cell lines by a different mechanism

Neuroblastoma, characterized by heterogeneous cell population, is a common solid tumor in childhood and some malignant neuroblastomas are refractory to conventional chemotherapy. Recently, treatment with arsenic trioxide (As₂O₃) was found effective in the treatment of acute promyelocytic leukemia as well as neuroblastoma cells by inducing apoptosis. To define the mechanism contributing to cell death in those heterogenous cell populations, the authors used two different types of neuroblastoma cells, SH-SY5Y and SK-N-AS, to compare the pathways that mediate death response to arsenic trioxide. With arsenic trioxide exposure, both cell lines were arrested at the S-G2/M phase with the increase of cyclin B expression and CDK1 activity. Although caspase 3 was activated in both cell lines, the NF-κB activity and the expression of cyclin D1, cyclin E, and p27 were different. Therefore, arsenic trioxide could be an effective cytotoxic drug for the treatment of heterogeneous cellular population of neuroblastoma.