c-FLIP在骨肉瘤中的表达及其临床意义

目的:探讨c-FLIP(细胞型Fas相关死亡域样白介素-1β转换酶抑制蛋白)在骨肉瘤组织中的表达情况,分析其表达与临床病理特征和预后的关系。方法:收集47例临床资料完整石蜡包埋的骨肉瘤肿瘤标本,25例骨软骨瘤标本,8例正常的骨组织标本,用免疫组化SP法和蛋白质印记法测定c-FLIP的表达水平,分析其表达与肿瘤组织学性状和预后的关系。结果:骨肉瘤标本中c-FLIP表达的阳性率是83%(39/47),而在骨软骨瘤标本中表达的阳性率是16%(4/25),两者比较P<0.01;结合临床资料分析,c-FLIP蛋白表达与患者年龄、发病次数、肿瘤体积、病理类型及有无转移无关(P>0.05),而与临床Ennecking分期有关(P<0.05)。结论:c-FLIP蛋白的过量表达在骨肉瘤的发生发展中可能起重要的作用。     

c-FLIP在骨肉瘤中的表达及其临床意义

目的：探讨c-FLIP（细胞型Fas相关死亡域样白介素-1β转换酶抑制蛋白）在骨肉瘤组织中的表达情况,分析其表达与临床病理特征和预后的关系。方法：收集47例临床资料完整石蜡包埋的骨肉瘤肿瘤标本,25例骨软骨瘤标本,8例正常的骨组织标本,用免疫组化SP法和蛋白质印记法测定c-FLIP的表达水平,分析其表达与肿瘤组织学性状和预后的关系。结果：骨肉瘤标本中c-FLIP表达的阳性率是83%（39/47）,而在骨软骨瘤标本中表达的阳性率是16%（4/25）,两者比较P〈0.01;结合临床资料分析,c-FLIP蛋白表达与患者年龄、发病次数、肿瘤体积、病理类型及有无转移无关（P〉0.05）,而与临床Ennecking分期有关（P〈0.05）。结论：c-FLIP蛋白的过量表达在骨肉瘤的发生发展中可能起重要的作用。     

c-FLIP和NF-κB p65在乳腺癌组织中的表达及临床意义

目的:探讨凋亡相关蛋白c-FLIP和NF-κB p65在人乳腺癌的表达及相关性,分析其与乳腺癌转移复发的关系。方法:应用免疫组织化学法检测80例乳腺癌组织和相应的癌旁组织中c-FLIP和NF-κBp65的表达情况。结果:80例乳腺癌组织中c-FLIP及NF-κB p65表达的阳性率显著高于相应的癌旁组织(P<0.05),c-FLIP的表达与TNM分期、组织学分级、腋窝淋巴结转移、术后复发及C-erbB-2表达密切相关(P<0.05);NF-κB p65的表达组织学分级、肿瘤大小及腋窝淋巴结转移密切相关(P<0.05)。c-FLIP和NF-κB p65的表达有显著正相关性(γ=0.271,P=0.015)。结论:c-FLIP和NF-κB p65的高表达可能在乳腺癌的发生及进展中起着重要作用。     

表皮肿瘤组织超氧化物歧化酶的表达及意义

目的 :探讨铜锌超氧化物歧化酶(CuZnSOD)和锰超氧化物歧化酶 (MnSOD)在表皮肿瘤中的表达及作用。方法 :通过半定量逆转录聚合酶链式反应 (RT PCR)和蛋白免疫印记 (WesternBlot)方法 ,研究了 2 6例基底细胞癌(Basalcellcarcinoma)、2 3例鳞状细胞癌 (Squa mascellcarcinoma)、18例日光性角化 (ActinicKeratosis)以及 8例曝光部正常皮肤、8例非曝光部正常皮肤中CuZnSOD和MnSOD的mRNA和蛋白表达水平。结果 :正常皮肤曝光部和非曝光部 2种SOD的mRNA和蛋白表达无差别 ;表皮肿瘤中CuZnSODmRNA表达与正常皮肤一致 ,MnSODmRNA表达升高 ;表皮肿瘤中 2种SOD蛋白的表达较正常皮肤低。结论 :表皮肿瘤中SOD的mRNA与蛋白表达水平存在差异 ,低蛋白表达可能与表皮肿瘤发生有关。     

c-FLIP及Caspase-8表达在乳腺癌组织中的表达及临床意义

目的探讨凋亡相关蛋白c-FLIP和Caspase-8在人乳腺癌组织中的表达及相关性,分析其与乳腺癌转移复发的关系。方法应用免疫组织化学法检测80例乳腺癌组织和相应的癌旁组织中c-FLIP和Caspase-8的表达情况。结果 80例乳腺癌组织中c-FLIP表达的阳性率显著高于相应的癌旁组织(P<0.05),c-FLIP的表达与TNM分期、组织学分级、腋窝淋巴结转移、术后复发及C-erbB-2表达密切相关(P<0.05);Caspase-8表达的阳性率显著低于相应的癌旁组织(P<0.05),Caspase-8的表达与组织学分级、肿瘤大小密切相关(P<0.05)。c-FLIP表达和Caspase-8表达有显著负相关性(γ=-0.380,P=0.001)。结论 c-FLIP的高表达及Caspase-8表达缺失可能在乳腺癌发生发展中起重要作用。     

c-FLIP和RECK在喉鳞状细胞癌及癌旁黏膜组织中的表达及其对于喉癌手术切除的意义

目的探讨癌基因c-FLIP和抑癌基因RECK在喉癌组织中的表达与喉癌临床特征的关系,了解喉癌的生物学行为,并从细胞和基因水平探讨喉癌手术安全切缘范围。方法分别采用PV9000免疫组化两步法及RT-PCR方法检测38例喉鳞癌组织及距癌组织边缘2 mm、5 mm、10mm组织中c-FLIP和RECK蛋白的表达及mRNA转录情况,同时取距离癌组织边缘15 mm的正常黏膜作为对照。结果(1)c-FLIP和RECK在喉鳞癌组织中的表达水平与患者临床分期和肿瘤病理学分级分别成正相关和负相关。(2)c-FLIP在伴颈淋巴结转移组中的表达水平明显高于非转移组,而RECK则正好相反。(3)c-FLIP表达情况依癌组织→癌旁2 mm→癌旁5 mm→癌旁10 mm→癌旁15 mm黏膜顺序递减,而RECK则递增。c-FLIP和RECK在癌组织与癌旁5 mm、10 mm及正常黏膜组织之间的表达水平有显著性差异(P〈0.05),而与癌旁2 mm处组织之间的差异无统计学意义。结论(1)c-FLIP和RECK两因子在喉鳞癌组织中的表达水平与喉鳞癌患者的临床病理参数密切相关。(2)c-FLlP和RECK可能都参与了喉鳞癌的发生、发展,并与喉鳞癌的浸润及转移有密切关系。(3)c-FLIP和RECK在喉鳞癌组织中及癌旁不同距离组织中的表达情况进一步表明喉鳞状细胞癌手术切除的范围应该包括距离肿瘤边缘5 mm之内的癌旁黏膜组织。     

Livin蛋白在表皮肿瘤中的表达及其意义

[目的]观察Livin蛋白在皮肤基底细胞癌、脂溢性角化病等表皮肿瘤中的表达情况。[方法]采用免疫组化和流式细胞术检测Livin蛋白在30例脂溢性角化病和15例皮肤基底细胞癌皮损中的表达情况,以20例正常皮肤组织作为对照。并采用流式细胞术对标本进行Livin蛋白定量分析。[结果]Livin蛋白在基底细胞癌中表达（86．67％）高于在脂溢性角化病组织（53．33％）和正常皮肤组织（5％）的表达（P〈0．05）,在30例脂溢性角化病皮损中,Livin蛋白阳性表达量高于在正常20例皮肤组织中的表达（P〈0．05）。[结论]脂溢性角化病和皮肤基底细胞癌的发生发展可能与Livin蛋白的异常表达有关。     

MMP-2在上皮性卵巢癌中的表达及临床意义

目的:探讨MMP-2在上皮性卵巢癌中的表达及临床意义.方法:15例正常卵巢组织、15例良性上皮性卵巢肿瘤组织, 89例上皮性卵巢癌组织标本,用免疫组化法检测MMP-2蛋白表达.结果:正常卵巢组织及良性卵巢肿瘤组织中,MMP-2蛋白不表达或低表达.上皮性卵巢癌中,MMP-2表达水平明显升高,高表达率为50.6%.MMP-2高表达与肿瘤晚期、瘤细胞的低分化、肿瘤转移有显著的相关性(P<0.05).结论:上皮性卵巢癌组织中,MMP-2蛋白表达上调.MMP-2蛋白在上皮性卵巢癌的生长和转移过程中发挥重要作用,有可能作为判断卵巢癌预后的指标.     

人肺癌组织端粒酶活性与p16基因二者相关性探讨

目的:探讨非小细胞肺癌(NSCLC)组织中端粒酶活性与p16基因的相关性及二者与肺癌发生发展的关系.方法:肺癌标本48例,12例癌旁组织,7例非肿瘤病例的正常肺组织.以Telomerase PCR ELISA检测标本端粒酶活性,以RT-PCR的方法检测p16基因mRNA转录情况,以免疫组化方法检测组织标本p16蛋白表达.结果:1)肺癌组织标本36/48(75.00%)和1例癌旁组织检出端粒酶活性.2)48例NSCLC组织中32例进行了p16 mRNA及蛋白表达检测.16/32(50.00%)检测到p16mRNA转录,7例正常组织中均检测到p16 mRNA转录.NSCLC组织标本中17/32(53.13%)未检测到p16蛋白表达,7例正常肺组织中均检测到p16蛋白表达.3)24例端粒酶阳性NSCLC组织中15例p16 mRNA表达缺失,8例端粒酶阴性的NSCLC组织中仅1例p16mRNA表达缺失.相关系数为-0.433,P=0.013,具有显著负相关.结论:端粒酶、p16基因在NSCLC的发生发展中起重要作用,端粒酶活性有望成为预测肿瘤发生、肿瘤诊断的良好指标.端粒、端粒酶与p16基因可能成为抗肿瘤治疗的新靶点.     

人骨肉瘤中nm23基因表达的初步研究

在人类肿瘤中ｎｍ２３基因的研究是肿瘤研究热之一，但该基因在骨肉瘤中的研究报道较少。作者应用免疫组化和原位杂交方法检测了人骨肉瘤和正常骨组织中ｎｍ２３基因的表达。材料和方法经病理检查证实的新鲜骨肉瘤标本３０例和非肿瘤病人术中切除的正常骨组织标本１５例作...     

Activation of Wnt/beta-catenin/Tcf signaling in mouse skin carcinogenesis

Although Wnt/beta-catenin/Tcf signaling pathway has been shown to be an important factor in the development of many malignancies including colorectal, ovarian, prostate, and many other cancers, little is known about its role in non-melanoma skin cancers. Here, we report the first evidence that beta-catenin/Tcf signaling pathway is constitutively activated in non-melanocytic skin tumors induced by two stage chemical carcinogenesis protocol. Mouse skin tumors showed cytoplasmic and nuclear accumulation of beta-catenin, and upregulation of beta-catenin/Tcf target genes (c-myc and c-jun). We found high levels of skin-expressed Wnt proteins (Wnt 3, 4, and 10b) in different parts of the tumors, likely representing key upstream events in beta-catenin/Tcf activation during mouse skin carcinogenesis. Inhibition of beta-catenin/Tcf signaling by ectopic expression of dominant negative Tcf4 resulted in significant inhibition of growth in squamous cell carcinoma cells. A role of the constitutive activation of beta-catenin/Tcf signaling in skin carcinogenesis is discussed.     

PPARgamma ligands enhance TRAIL-induced apoptosis through DR5 upregulation and c-FLIP downregulation in human lung cancer cells

Peroxisome proliferator-activated receptor gamma (PPARgamma) ligands are potential chemo-preventive agents. Many studies have shown that PPARy ligands induce apoptosis in various types of cancer cells including lung cancer cells. Some PPAR gamma ligands have been shown to downregulate c-FLIP expression and thus enhance tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis in some cancer cell lines. In the current study, we further show that PPARy ligands induced the expression of death receptor 5 (DR5) and increased DR5 distribution at the cell surface in addition to reducing c-FLIP levels in human lung cancer cells. These agents cooperated with TRAIL to enhance induction of apoptosis in human lung cancer cells. Both overexpression of c-FLIP and knockdown of DR5 abrogated PPARgamma ligand's ability to enhance TRAIL-induced apoptosis. Thus, it appears that not only c-FLIP downregulation but also DR5 upregulation contribute to PPARy ligand-mediated enhancement of TRAIL-induced apoptosis in human lung cancer cells. Both the PPARgamma antagonist GW9662 and silencing PPARgamma expression failed to diminish PPARgamma ligand-induced DR5 upregulation or c-FLIP downregulation, indicating that PPARy ligands modulate the expression of DR5 and c-FLIP through a PPARy-independent mechanism. Collectively, we conclude that PPARy ligands exert PPARy-independent effects on inducing DR5 expression and downregulating c-FLIP levels, leading to enhancement of TRAIL-induced apoptosis.     

Flavopiridol induces cellular FLICE-inhibitory protein degradation by the proteasome and promotes TRAIL-induced early signaling and apoptosis in breast tumor cells

The cyclin-dependent kinase inhibitor flavopiridol is undergoing clinical trials as an antitumor drug. We show here that pretreatment of different human breast cancer cell lines with flavopiridol facilitates tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis. In breast tumor cells, apoptosis induction by TRAIL is blocked at the level of apical caspase-8 activation. Flavopiridol treatment enhances TRAIL-induced formation of death-inducing signaling complex and early processing of procaspase-8. Subsequently, a TRAIL-induced, mitochondria-operated pathway of apoptosis is activated in cells treated with flavopiridol. Down-regulation of cellular FLICE-inhibitory proteins (c-FLIP; c-FLIP(L) and c-FLIP(S)) is observed on flavopiridol treatment. c-FLIP loss and apoptosis sensitization by flavopiridol are both prevented in cells treated with an inhibitor of the ubiquitin-proteasome system. Furthermore, targeting c-FLIP directly with small interfering RNA oligonucleotides also sensitizes various human breast tumor cell lines to TRAIL-induced apoptosis. Our results indicate that flavopiridol sensitizes breast cancer cells to TRAIL-induced apoptosis by facilitating early events in the apoptotic pathway, and this combination treatment could be regarded as a potential therapeutic tool against breast tumors.     

Immunohistopathological Study of c-FLIP Protein in Mycosis Fungoides

Background: Mycosis fungoides (MF) is the commonest variant of primary cutaneous T cell lymphoma with several clinicopathologic variants. Defective apoptotic mechanism may be important in the pathogenesis and progression of MF. c-FLIP protein is an important anti-apoptotic marker and chemotherapeutic resistant factor. This study aimed to evaluate the c-FLIP expression in MF and its role in the pathogenesis of MF. Methods: Twenty patients of MF and ten normal persons were included in this study. Skin biopsies were obtained from both patients and controls. They were studied and examined immunohistochemically for the expression of CD4 and c-FLIP. Results: c-FLIP expression was significantly increased in patients when compared to controls in both epidermis and dermis. There were positive correlations between c-FLIP expression and CD4+ expression in both epidermal and dermal lesions of patients group. There were statistically significant positive correlations between c-FLIP expression (in both dermal and epidermal lesions) and the age of patients. c-FLIP expression increased with the tumor progression but with no statistical significance. Conclusion: Defective regulation of apoptosis has been considered as a main cause for accumulation of clonal T cells, and it was related to an increased expression of c-FLIP which may have a role in the pathogenesis of MF. Also, c-FLIP may have prognostic information in MF as its level increased with both age of the patients and tumor progression.     

Rocaglamide and a XIAP inhibitor cooperatively sensitize TRAIL-mediated apoptosis in Hodgkin's lymphomas

Although most of the patients with Hodgkin's lymphoma (HL) can be cured by the current regimen of high-dose multiagent chemotherapy, the treatment causes high risks of later toxicities including secondary malignancies. Therefore, new rational strategies are needed for HL treatment. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising anticancer agent due to its tumor selectivity and its lack of toxicity for normal cells. Unfortunately, many cancers remain resistant to TRAIL including HL. HL is characterized by enhanced expression of cellular caspase-8 (FLICE)-inhibitory protein (c-FLIP) and X-linked inhibitor of apoptosis (XIAP), which block receptor-mediated apoptosis by inhibiting caspase-8 and caspase-3, respectively. We have recently discovered the herbal compound Rocaglamide, which breaks TRAIL-resistance in acute T cell leukemia through inhibition of c-FLIP expression. We have also shown that small molecule XIAP inhibitors can sensitize TRAIL-mediated apoptosis in several resistant tumors. However, whether targeting XIAP or c-FLIP is also a suitable strategy to prime HL cells for TRAIL-induced apoptosis has not yet been investigated. In our study, we show that Rocaglamide suppresses c-FLIP expression in HL cells in a dose- and time-dependent manner. However, downregulation of c-FLIP alone was not sufficient to sensitize TRAIL-induced apoptosis in HL cells. Similarly, treatment of HL cells with a small molecule XIAP inhibitor resulted in a moderate induction of apoptosis. However, inhibition of XIAP alone was also not sufficient to enhance TRAIL-induced cell death. Synergistic increase in TRAIL-mediated killing of HL cells was only obtained by combination of Rocaglamide and XIAP inhibitors. Our study demonstrates that targeting both c-FLIP and XIAP are necessary for an efficient treatment of HL.     

Resistance of pancreatic cancer to gemcitabine treatment is dependent on mitochondria-mediated apoptosis

Palliative chemotherapy with gemcitabine, a common mode of treatment of pancreatic cancer, has little influence on patients' survival. We investigated the impact of anti-apoptotic Bcl-xL protein and its antagonist Bax on gemcitabine-induced apoptosis in human pancreatic carcinoma cells in vitro and in vivo. The level of Bcl-xL and Bax expression was determined in 3 established pancreatic cancer cell lines that differ in their sensitivity to gemcitabine-mediated apoptosis. Bcl-xL and Bax genes were transduced into Colo357 cells by retroviral infection. In addition, cells were transfected with c-FLIP to assess involvement of CD95 and caspase-8. The impact of Bax/Bcl-xL expression on gemcitabine-sensitivity in vivo was evaluated in orthotopic Colo357 tumors in SCID mice. The apoptotic index revealed a strong inverse correlation between Bcl-xL expression and gemcitabine-induced apoptosis in the pancreatic carcinoma cell lines tested. Caspase-8 and Bid were cleaved in Colo357 cells exposed to gemcitabine, and there was no correlation with either Bcl-xL or with Bax expression. In contrast, the lack of mitochondrial transmembrane potential transition, release of cytochrome-c and absence of caspase-9- and PARP-cleavage showed a strong correlation with Bcl-xL expression. Expression of c-FLIP significantly increased the resistance towards gemcitabine. Orthotopically growing Colo357-bcl-xl tumors in SCID mice were refractory to gemcitabine treatment, and in contrast to the in vitro data, Colo357-bax tumors exhibited a 12-fold greater tumor regression than Colo357-wild-type tumors in the control group. Gemcitabine-induced apoptosis involves the mitochondria-mediated signaling pathway. A functional restoration of this pathway appears to be essential to overcome the resistance mechanisms of pancreatic tumor cells and to improve the response to therapy as demonstrated by Bax overexpression in a clinically relevant tumor model.