Six1对甲状腺癌细胞侵袭迁移的影响

目的探讨Six1对甲状腺癌细胞侵袭迁移的影响。方法甲状腺癌BCPAP细胞转染Six1小干扰RNA(Six1 siRNA)和小干扰RNA阴性对照(siRNA control),采用细胞划痕实验检测细胞迁移,Transwell小室检测细胞侵袭,Western印迹检测Six1、神经钙黏素(N-cadherin)、上皮钙黏附素(E-cadherin)蛋白表达。结果转染siRNA control后的BCPAP细胞迁移率、侵袭细胞数目及细胞中Six1、N-cadherin、E-cadherin蛋白表达水平与没有转染的BCPAP细胞相比无变化。转染Six1 siRNA后的BCPAP细胞迁移率、侵袭细胞数目及细胞中Six1、N-cadherin蛋白表达水平与没有转染的BCPAP细胞相比明显降低,而E-cadherin蛋白水平明显升高。结论敲低Six1可能通过影响E-cadherin、N-cadherin表达抑制甲状腺癌细胞侵袭迁移。     

沉默GRP78基因对人卵巢癌SKOV3细胞侵袭力的抑制作用

目的 探讨RNA干涉技术(RNAi)沉默葡萄糖调节蛋白78(GRP78)基因对人卵巢癌SKOV3细胞侵袭力的影响,阐明GRP78基因沉默对SKOV3细胞侵袭力抑制作用的生物学机制.方法 设计并构建pSilencerTM3.0-H1-GRP78 siRNA重组质粒,脂质体介导转染至SKOV3细胞.RT-PCR和Western印迹法检测GRP78基因表达;RT-PCR检测MMP-2、MMP-9 mRNA表达;Transwell小室侵袭实验检测细胞侵袭力;Transwell小室迁移实验检测细胞迁移率.结果 RT-PCR及Western印迹检测转染GRP78 siRNA重组质粒SKOV3细胞GRP78基因表达抑制;RT-PCR结果显示与对照组相比转染GRP78 siRNA重组质粒SKOV3细胞MMP-2、MMP-9 mRNA表达降低;Transwell小室侵袭实验和迁移实验结果显示转染GRP78 siRNA重组质粒SKOV3细胞穿透细胞数(P＜0.05)和迁移率均明显降低(P＜0.05).结论 ①转染GRP78 siRNA重组质粒有效抑制SKOV3细胞GRP78基因表达;②抑制GRP78基因表达能降低SKOV3细胞的侵袭力和迁移力,有望为抑制卵巢癌转移基因治疗提供新的靶点.     

Kiss-1在胃腺癌组织中的表达及对胃腺癌MKN-45细胞增殖、迁移能力的影响

目的探讨肿瘤抑制因子(Kiss)-1在胃腺癌组织中的表达及对胃腺癌细胞增殖、迁移能力的影响。方法 qRT-PCR检测50例胃腺癌组织和对应的癌旁组织中Kiss-1水平。分别将Kiss-1小干扰RNA(siRNA Kiss-1)和对照小RNA(siRNA control)、Kiss-1过表达载体(p EGFP-N1-Kiss-1)和对照空载体(p EGFP-N1)转染至胃腺癌细胞株(MKN-45)中,培养48 h后,Western印迹检测细胞中Kiss-1、基质金属蛋白酶(MMP)-9、MMP-2、β-连环蛋白(β-catenin)、C-myc蛋白水平,噻唑蓝(MTT)检测细胞增殖能力,细胞划痕实验检测细胞迁移能力。用Wnt/β-catenin信号通路抑制剂FH535作用胃腺癌细胞后,检测细胞增殖和凋亡情况。结果 Kiss-1在胃腺癌组织中的表达水平显著低于癌旁组织(P<0.01)。p EGFP-N1-Kiss-1组细胞存活率和迁移率显著低于p EGFP-N1组,siRNA Kiss-1组显著高于siRNA control组(P<0.01)。p EGFP-N1-Kiss-1组细胞中MMP-9、MMP-2、β-catenin、C-myc水平显著低于p EGFP-N1组,siRNA Kiss-1组显著高于siRNA control组(P<0.01)。抑制剂作用后的胃腺癌细胞增殖迁移趋势与p EGFP-N1-Kiss-1组一致。结论 Kiss-1在胃腺癌组织中低表达,Kiss-1通过Wnt/β-catenin信号通路抑制胃腺癌细胞增殖迁移能力。     

支架蛋白RACK1小干扰RNA抑制卵巢癌细胞CAOV3的增殖、迁移和侵袭

目的观察支架蛋白RACK1小干扰RNA(siRNA)对卵巢癌CAOV3细胞增殖、迁移和侵袭的影响和基质金属蛋白酶(MMP)-2和MMP-9的表达变化。方法体外培养CAOV3细胞,实验分scramble siRNA组和RACK1 siRNA组;Lipofectamine 2000转染CAOV3细胞,Western印迹检测RACK1的干涉效能;MTT法测定CAOV3细胞的增殖率;划痕实验和transwell迁移和侵袭实验研究RACK1对细胞体外增殖、迁移和侵袭运动能力的影响;Western印迹检测CAOV3细胞中MMP-2和MMP-9蛋白表达。结果与scramble siRNA组比较,RACK1 siRNA组RACK1的蛋白表达水平降低,明显抑制CAOV3细胞的增殖、迁移和侵袭,降低MMP-2和MMP-9蛋白表达。结论下调RACK1表达可抑制卵巢癌细胞CAOV3的增殖、迁移和侵袭,其机制可能与改变MMP-2和MMP-9蛋白表达相关。     

miRNA-21调控胃癌细胞增殖及侵袭的机制

目的探讨靶向沉默miRNA-21表达对胃癌细胞增殖及侵袭的影响和机制。方法将人胃癌SGC-7901细胞分为正常对照组、转染siRNA Control的阴性对照组和转染siRNA miRNA-21基因的沉默组,按照Invitrogen公司的脂质体Lipofectamine~(TM)2000方法进行转染,48 h后,CCK8实验检测细胞增殖;Transwell小室检测细胞侵袭能力;Western印迹检测基质金属蛋白酶(MMP)-2、MMP-9、Notch1、Hes1蛋白表达。结果转染siRNA miRNA-21后能显著降低miRNA-21的表达(P<0.01);与正常对照组及转染阴性组比较,沉默组细胞存活率、细胞侵袭数显著降低,MMP-2、MMP-9、Notch1、Hes1蛋白表达显著下调(P<0.01)。结论靶向沉默miRNA-21表达能显著降低胃癌细胞的增殖及侵袭能力,其机制与抑制Notch1信号通路有关。     

干扰ECT2基因表达对结直肠癌SW620细胞侵袭和迁移的影响及机制研究

目的探讨转染小干扰RNA(siRNA)干扰上皮细胞转化序列2(ECT2)基因的表达对人结直肠癌SW620细胞侵袭和迁移的影响及机制。方法采用脂质体Lipofectamine 2000转染试剂将ECT2特异性siRNA或非特异性siRNA转染至SW620细胞,分别记为si-ECT2组和si-NC组,将未行转染的SW620细胞记为Control组。采用实时荧光定量PCR法(real-time qPCR)和蛋白质印迹法(Western blotting)分别检测各组SW620细胞中ECT2的mRNA和蛋白表达水平,采用Transwell侵袭实验检测各组SW620细胞的侵袭能力,采用划痕愈合实验检测各组SW620细胞的迁移能力,采用Western blotting法检测各组SW620细胞中基质金属蛋白酶-2(MMP-2)和MMP-9蛋白表达水平。结果与si-NC组相比,si-ECT2组SW620细胞中ECT2的mRNA和蛋白表达水平均明显降低,穿膜细胞数量明显减少,划痕愈合率明显降低,MMP-2和MMP-9蛋白表达水平均明显降低,差异均有统计学意义(P均0.05)。Control组的上述各项指标与si-NC组之间的差异均无统计学意义(P均0.05)。结论干扰ECT2基因的表达能够抑制结直肠癌SW620细胞的侵袭和迁移能力,其作用机制可能与下调MMP-2和MMP-9蛋白表达有关。     

HDAC1对结直肠癌细胞凋亡及侵袭能力的影响

目的研究组蛋白去乙酰化酶1(HDAC1)对结直肠癌细胞凋亡及侵袭能力的影响。方法结直肠癌细胞Caco2中转染HDAC1 siRNA和siRNA control记为HDAC1 siRNA和siRNA-NC,以不做转染的细胞为Control。qRT-PCR、Western blotting检测细胞HDAC1 mRNA、HDAC1蛋白水平,流式细胞术测定细胞凋亡,Transwell小室测定细胞侵袭和迁移,Western blotting测定基质金属蛋白酶-2(MMP-2)、基质金属蛋白酶-9(MMP-9)、Bcl-2相关X蛋白(Bax)、活化的含半胱氨酸的天冬氨酸蛋白水解酶3(Cleaved caspase-3)蛋白水平。结果 siRNA-NC细胞HDAC1 mRNA、HDAC1蛋白、细胞凋亡率、细胞侵袭、迁移数目和MMP-2、MMP-9、Bax、Cleaved caspase-3蛋白水平与Control相比无明显变化(P0.05)。HDAC1 siRNA细胞HDAC1 mRNA、HDAC1蛋白明显降低,细胞凋亡率明显升高,细胞侵袭和迁移数目明显减少,细胞MMP-2、MMP-9蛋白水平表达下降,Bax、Cleaved caspase-3蛋白水平升高,与Control相比,差异有统计学意义(P0.05)。结论 HDAC1表达下调诱导结直肠癌细胞凋亡,抑制结直肠癌细胞侵袭及迁移,其作用机制与下调MMP-2、MMP-9和促进Bax、Cleaved caspase-3表达有关。     

NEDD9表达下调对结肠癌生物学行为的影响

背景:NEDD9在细胞迁移、趋化、凋亡、细胞周期中发挥重要作用,NEDD9 siRNA可有效降低基因表达,并从不同水平上促进细胞凋亡、阻断肿瘤细胞迁移和侵袭。目的:探讨NEDD9在结肠癌组织中的表达以及siRNA干扰NEDD9表达对结肠癌Caco-2细胞增殖、凋亡、迁移和侵袭的影响。方法:收集2017年1月—2018年1月漯河市郾城区人民医院确诊的结肠癌组织和相应癌旁组织。常规培养结肠癌Caco-2细胞,采用LipofectamineTM2000转染细胞,并将细胞分为对照组、阴性对照组和siRNA干扰组。采用qRT-PCR法检测结肠癌组织和细胞中NEDD9 mRNA表达,CCK-8法检测细胞增殖,流式细胞术检测细胞凋亡,Transwell实验检测细胞迁移和侵袭,蛋白质印迹法检测NEDD9、MMP-9、Bcl-2、Bax、TIMP1蛋白表达。结果:结肠癌组织NEDD9 mRNA表达显著高于癌旁组织(P 0. 05)。与对照组相比,siRNA干扰组NEDD9 mRNA表达明显降低(P 0. 05),细胞增殖、迁移和侵袭能力受到明显抑制(P 0. 05),细胞凋亡增强(P 0. 05),NEDD9、MMP-9、Bcl-2蛋白表达显著降低(P 0. 05),Bax、TIMP1蛋白表达显著增加(P 0. 05)。结论:NEDD9基因可通过凋亡相关基因Bcl-2和Bax以及侵袭相关基因MMP-9和TIMP1来调控结肠癌Caco-2细胞增殖、凋亡、迁移和侵袭能力。     

血管紧张素原对动脉粥样硬化的作用及机制

目的探讨血管紧张素原(AGT)对动脉粥样硬化(AS)的作用机制。方法构建AS小鼠模型,RT-PCR检测AS小鼠斑块组织和正常小鼠主动脉血管组织中AGT的表达水平。以人巨噬细胞RAW264.7和人主动脉平滑肌细胞(HA-VSMC)为研究对象,转染siRNA AGT、siRNA control,实时荧光定量PCR(RT-PCR)检测转染后的AGT水平。MTT检测细胞转染后细胞的增殖情况,流式细胞仪检测转染后的细胞凋亡情况。Western印迹检测转染后细胞中Caspase-3、Bcl-2、Bax蛋白表达水平。结果 AS小鼠斑块组织中AGT的表达水平高于正常小鼠主动脉组织(P<0.01)。siRNA AGT可以有效抑制巨噬细胞RAW264.7和人主动脉平滑肌细胞HA-VSMC中AGT的转录表达。转染siRNA AGT后的巨噬细胞RAW264.7存活率与siRNA control组比较差异显著(P<0.01),转染siRNA AGT后的主动脉平滑肌细胞HA-VSMC存活率与siRNA control组差异显著(P<0.01),抑制AGT的表达可以抑制巨噬细胞和平滑肌细胞的增殖。转染siRNA AGT后的巨噬细胞RAW264.7凋亡率显著高于siRNA control组(P<0.01);转染siRNA AGT后的主动脉平滑肌细胞HA-VSMC凋亡率显著高于siRNA control组(P<0.01)。转染siRNA AGT后的巨噬细胞RAW264.7和主动脉平滑肌细胞HA-VSMC中Caspase-3、Bax蛋白表达量显著高于siRNA control组,Bcl-2蛋白表达量显著低于siRNA control组(均P<0.01)。结论AGT在鼠AS斑块组织中过表达,抑制AGT可以抑制AS相关细胞增殖,促进细胞凋亡,作用机制与凋亡相关蛋白Caspase-3、Bcl-2、Bax有关。     

Twist对膀胱癌细胞迁移、侵袭及MMP-2、MMP-9表达的影响

目的探讨Twist对膀胱癌细胞迁移、侵袭及基质金属蛋白酶(MMP)-2、MMP-9表达的影响。方法用qRT-PCR和Western印迹法分别检测膀胱癌5637、T24、BIU-87细胞和正常膀胱SVHUC-1细胞中Twist mRNA和蛋白水平,筛选表达水平最高的T24细胞,在T24细胞中转染shRNA Twist、shRNA对照命名为sh-Twist组和sh-NC组,并以不做处理的T24细胞为Con组,用qRT-PCR和Western印迹法测定各组细胞中Twist mRNA和蛋白水平,Transwell小室测定细胞侵袭和迁移,Western印迹测定各组细胞中上皮间质转化(EMT)相关蛋白波形蛋白(Vimentin)、上皮钙黏附素(E-cadherin)和侵袭迁移相关蛋白MMP-2、MMP-9表达水平。结果与正常膀胱SVHUC-1细胞比较,Twist在膀胱癌细胞中转录和表达水平均明显升高(P0.05)。sh-Twist组Twist mRNA和蛋白水平均明显低于Con组(P0.05),sh-NC组Twist mRNA和蛋白水平与Con组相比差异无统计学意义(P0.05)。sh-Twist组细胞迁移数目、侵袭数目及MMP-2、MMP-9蛋白水平均明显低于Con组(P0.05),sh-NC组细胞迁移数目、侵袭数目及MMP-2、MMP-9蛋白水平与Con组相比差异无统计学意义(P0.05)。sh-Twist组E-cadherin蛋白水平与Con组相比明显升高(P0.05),Vimentin蛋白水平与Con组相比明显下降(P0.05),而sh-NC组Vimentin、E-cadherin水平与Con组相比差异无统计学意义(P0.05)。结论 Twist敲低能够降低膀胱癌细胞侵袭和迁移能力,抑制膀胱癌细胞EMT。     

Hereditary ovarian cancer

At least 10% of ovarian tumors are hereditary and associated with highly penetrant, autosomal, dominant genetic predisposition. Three clinical manifestations of hereditary ovarian cancer have been identified: site-specific ovarian cancer, hereditary breast and/or ovarian cancer (HBOC) and hereditary non-polyposis colorectal cancer (HNPCC) syndromes. BRCA germline mutations account for more than 90% of all hereditary epithelial ovarian tumors whereas most of the remaining 10% are caused by MLH1 and MSH2 mutations, which are susceptibility genes of HNPCC. Genetic testing is available for each of the three hereditary syndromes above mentioned. The recommendations for OC surveillance in high-risk women having a strong family history or BRCA mutation carriers include transvaginal pelvic ultrasound with color Doppler and serum CA125 every 6 months. Bilateral salpingo-oophorectomy appears to be effective to reduce the risk of ovarian cancer in BRCA mutation carriers. Hysterosalpingo-oophorectomy should be considered in HNPCC women who undergo surgery for colorectal carcinoma.     

Gonadotropins and ovarian cancer

Ovarian epithelial cancer (OEC) accounts for 90% of all ovarian cancers and is the leading cause of death from gynecological cancers in North America and Europe. Despite its clinical significance, the factors that regulate the development and progression of ovarian cancer are among the least understood of all major human malignancies. The two gonadotropins, FSH and LH, are key regulators of ovarian cell functions, and the potential role of gonadotropins in the pathogenesis of ovarian cancer is suggested. Ovarian carcinomas have been found to express specific receptors for gonadotropins. The presence of gonadotropins in ovarian tumor fluid suggests the importance of these factors in the transformation and progression of ovarian cancers as well as being prognostic indicators. Functionally, there is evidence showing a direct action of gonadotropins on ovarian tumor cell growth. This review summarizes the key findings and recent advances in our understanding of these peptide hormones in ovarian cancer development and progression and their role in potential future cancer therapy. We will first discuss the supporting evidence and controversies in the "gonadotropin theory" and the use of animal models for exploring the involvement of gonadotropins in the etiology of ovarian cancer. The role of gonadotropins in regulating the proliferation, survival, and metastasis of OEC is next summarized. Relevant data from ovarian surface epithelium, which is widely believed to be the precursor of OEC, are also described. Finally, we will discuss the clinical applications of gonadotropins in ovarian cancer and the recent progress in drug development.     

Chemotherapy in advanced ovarian cancer

Summary The chemotherapy of advanced ovarian cancer is reviewed. Treatment with single agents results in low remission rates and few complete remissions. The results have been improved with modern combination chemotherapy, which includes cisplatin, although a longer follow up is needed for definite conclusions to be made concerning survival. Toxicity and drug resistance remain important problems. The future prospects of treatment with emphasis on intraperitoneal chemotherapy are discussed.     

Biochemical markers in ovarian cancer

Fucose and sialic acid were measured in 41 patients with malignant ovarian tumors and in 25 patients with benign tumors. Supplementary, serial determinations done in 20 patients, with advanced ovarian cancer subjected to primary or secondary surgery (optimal tumor debulking) and subsequent chemotherapy enables to suggest the possibility to use these parameters as markers in monitoring ovarian cancer evolution and treatment.     

Diagnosis and staging of ovarian cancer

Summary The diagnostic process in ovarian carcinoma is divided into the pre-and intraoperative procedures, examinations of tumor tissue, and follow-up. For preoperative diagnosis, the probability of a palpable adnexal mass being a malignant tumor should first be ascertained by sonography. This should be followed by an appropriate general examination, a search for tumor outside the abdominal cavity and in the liver parenchyma as well as by determination of markers. Intraoperative diagnosis determines the tumor stage and must be carried out all the more comprehensively when the ovarian carcinoma is more limited. Histologic subtype and degree of differentiation are in direct relation to the tumor stage, whereas the size of the primary tumor is often indirectly proportional to its extent. Besides the morphological analysis, the determination of possible chemoresistance and chemosensitivity, as well as further investigations on fresh tumor tissue are included in the tissue examination. Follow-up after a curative operation consists of gynecologic examination and Douglas lavages if tumor is still present in CT Scans and sonographs. To verify a relapse, laparoscopy can be used, but to ascertain a complete remission, a laparotomy is necessary.Paper presented at the Annual Meeting of the Swiss Society for Oncology, Basel, March 1983     

Dissemination of intraperitoneal ovarian cancer: Discussion of mechanisms and demonstration of lymphatic spreading in ovarian cancer model

Ovarian cancer is often accompanied by severe ascites. This complication aggravates the disease per se and the chances for its successful treatment. The etiology of ascites is not well understood nor are efficient therapies for ascites available. These empirical observations support the view that ascites might be caused by blocking of lymphatic vessels. Furthermore, it suggests that cancer cells might be the blocking agent and could use lymphatic vessels for metastatic spreading. To test this hypothesis, we used labeled cancer cells in an immuno-competent animal model of ovarian cancer and followed their dissemination. These NuTu-19 cells are ovarian cancer cells derived from normal rat ovarian epithelial cells, the origin of the most frequent ovarian cancer. Thus studying NuTu-19 cell behavior in an animal model is likely to reflect the progression of the human disease. To unambiguously document the migration of NuTu-19 cells from the peritoneum to remote organs, we generated EGFP expressing NuTu-19 cells by transduction with EGFP-lentiviral vectors. The EGFP positive NuTu-19 cells were injected intraperitoneally into immuno-competent FISHER 344 rats, and the metastatic spreading was monitored. Metastases were observed on the peritoneum, the omentum and in the parathymus. This clearly demonstrates that systemic spreading of NuTu-19 ovarian cancer cells is conducted by lymphatic ways. Animals die 7 weeks after injection, with severe ascites, which suggests that blockage of lymphatic drainage by the cancer cell growth is an important complication of the disease.