HOXB7基因沉默对人胃癌细胞活力、迁移和侵袭的影响

目的:探讨靶向沉默HOXB7基因对人胃癌SGC-7901细胞活力、迁移和侵袭作用的影响及潜在机制。方法:设计靶向HOXB7的siRNA,然后瞬时转染胃癌SGC-7901细胞,实时荧光定量PCR和Western blot法检测siRNA靶向沉默的效果。MTT法检测细胞的活力,Western blot法检测细胞周期相关蛋白CDK4和cyclin D1蛋白的表达水平,Transwell小室侵袭实验检测SGC-7901细胞的侵袭能力,实时荧光定量PCR和Western blot法检测胃癌SGC-7901细胞PTEN和VEGF的表达水平以及p-AKT的蛋白水平。结果:胃癌组织中HOXB7基因在mRNA和蛋白水平的表达均显著高于对照组织。siRNA可有效靶向沉默SGC-7901细胞中HOXB7的表达,且沉默HOXB7表达后SGC-7901细胞活力明显下降,同时细胞周期相关蛋白CDK4和cyclin D1的表达亦下调。靶向沉默HOXB7可明显抑制SGC-7901细胞中AKT的磷酸化水平,抑制胃癌细胞的侵袭转移,同时下调VEGF的表达水平,抑制胃癌组织中的肿瘤血管生成。结论:HOXB7作为一个癌基因,可上调细胞周期相关蛋白CDK4、cyclin D1和侵袭相关分子VEGF的表达,上调AKT的磷酸化水平,进而抑制PTEN的功能,促进胃癌细胞SGC-7901的生长、迁移和侵袭能力。     

靶向沉默Ku80增强顺铂诱导的人肺腺癌细胞凋亡

目的:观察沉默Ku80能否增强顺铂诱导的人肺腺癌耐药细胞的细胞凋亡。方法:首先采用RT-PCR、Westernblot方法比较Ku80在人肺腺癌亲代(A549)与耐药细胞系(A549/DDP)的表达水平;将Ku80siRNA及si-Scramble转染至A549/DDP细胞后,加药组转染后48小时加顺铂作用24小时;应用MTT法检测顺铂对各组细胞的抑制率,采用流式细胞仪检测凋亡率,Caspase-3活性检测试剂盒检测Caspase-3活化程度。结果:Ku80 mRNA、蛋白在A549/DDP表达水平显著高于其亲代A549;si-Ku80组A549/DDP细胞的Ku80的mRNA、蛋白表达水平下调;顺铂对si-Ku80组细胞的抑制率明显高于si-Scram-ble组与Control组;在6μg/ml顺铂作用24小时后,si-Ku80组细胞凋亡率及Caspase-3活化程度高于si-Scramble组和Control组(P<0.05)。结论:靶向Ku80siRNA导入人肺腺癌耐药细胞特异性下调Ku80的表达,增强顺铂诱导的人肺腺癌耐药细胞凋亡作用。     

Annexin A3抑制顺铂诱导人卵巢上皮癌细胞凋亡

目的 进一步探讨annexin A3能否通过凋亡途径调节卵巢癌细胞对顺铂敏感性.方法 运用反转录病毒载体构建及目的细胞感染技术,获得annexin A3稳定上调和稳定下调卵巢癌细胞系,运用annexin V/碘化丙啶分析方法和免疫印迹法检测细胞凋亡率、细胞内caspase蛋白裂解水平和annexin A3表达水平.结果 顺铂敏感细胞annexin A3表达水平上调后,60μg/mL顺铂诱导细胞凋亡率由68.72%±1.01%下调至29.13%±2.61%(P<0.05);高浓度顺铂诱导caspase和PARP蛋白裂解.相反,顺铂耐药细胞内mumxin A3水平下调后,凋亡率由35.05%±3.06%上升至76.73%±6.42%(P<0.05),低浓度顺铂诱导caspase和PARP蛋白裂解.此外,annexin A3还能够显著抑制卵巢癌细胞内P38 MAPK磷酸化.结论 Annexin A3能通过抑制细胞凋亡调节卵巢癌细胞对顺铂的敏感性.     

顺铂同步放化疗方案治疗中晚期宫颈癌的疗效

目的 探讨顺铂同步放化疗方案治疗中晚期宫颈癌的临床疗效。方法 回顾分析2016年10月~2019年5月于我院就诊的50例中晚期宫颈癌患者的临床资料,随机分为对照组和实验组,各25例。实验组行顺铂同步放化疗方案,对照组行单纯放疗,比较两组近期效果、不良反应及肿瘤标志物含量。结果 实验组肿瘤控制率为84.00%,高于对照组的52.00%,差异有统计学意义（P＜0.05）;实验组糖类抗原50（CA50）、癌胚抗原（CEA）、鳞状上皮细胞癌相关抗原（SCC）水平均低于对照组,差异有统计学意义（P＜0.05）;实验组不良反应发生率为40.00%,高于对照组的32.00%,但差异无统计学意义（P＞0.05）。结论 顺铂同步放化疗方案治疗中晚期宫颈癌近期疗效较好,有助于降低肿瘤恶性程度,且不增加不良反应,安全性高。     

敲除EZH2抑制人宫颈癌细胞系迁移和侵袭

目的 研究zeste基因同源蛋白2(EZH2)对人宫颈癌细胞系迁移和侵袭的影响,并初步探讨其作用机制.方法 利用CRISPR/Cas9敲除人宫颈癌HeLa和人子宫颈鳞癌SiHa细胞系中EZH2的表达.划痕实验检测细胞迁移;Transwell小室法检测细胞体外迁移和侵袭;实时荧光定量PCR检测STAT3 mRNA水平;W...     

miR-451a表达抑制宫颈癌细胞侵袭和顺铂耐药的影响及作用机制

目的 探讨miR-451a对宫颈癌细胞侵袭和顺铂(DDP)耐药的影响及作用机制研究。方法 qRT-PCR检测miR-451a在宫颈癌组织中的表达水平,分析miR-451a的表达水平与患者临床病理参数的关系。宫颈癌细胞HeLa分为对照组(NC HeLa组)和miR-451a模拟物组(miR-451a mimic HeLa组),分别进行各组mimic转染,Boyden实验检测各组细胞侵袭能力。采用药物递增诱导法建立DDP耐药细胞株HeLa(DDP-HeLa),qRT-PCR检测HeLa和DDP-HeLa细胞中miR-451a的表达。DDP-HeLa细胞分为NC DDP-HeLa组和miR-451a mimic DDP-HeLa组,分别进行各组mimic转染,MTS实验检测NC HeLa组、miR-451a mimic HeLa组、NC DDP-HeLa组和miR-451a mimic DDP-HeLa组对DDP敏感性的影响。Western blotting检测各组细胞中EMT相关蛋白E-cadherin、N-cadherin和Vimentin的表达。结果 miR-451a宫颈癌组织中的表达...     

膀胱癌T24顺铂耐药细胞系的建立及生物学特性

目的 采用浓度梯度法诱导建立人膀胱癌顺铂耐药细胞系(T24/DDP),并观察其生物学特性.方法 利用人膀胱癌细胞株T24,采用顺铂(DDP)作为诱导剂在体外建立人膀胱癌顺铂耐药细胞模型T24/DDP(0.6μg/mL).通过倒置相差显微镜下观察T24、T24/DDP细胞形态,MTT法检测细胞多药耐药性,生长曲线检测细胞的增殖能力,流式细胞术测细胞的周期,qRT-PCR检测耐药基因MRP(muhidrug resistance-associated protein)的表达.结果 经过顺铂14个月的诱导,成功建立了T24/DDP.倒置相差显微镜下观察与T24细胞相比较,T24/DDP排列不规则,形态出现多形性,出现巨细胞;与T24相比较,T24/DDP对顺铂、阿霉素、长春新碱、甲氨蝶呤均产生不同程度的交叉耐药,比T24细胞倍增时间明显延长,细胞周期中G1期细胞增多,G2、S期细胞减少.qRT-PCR结果显示,耐药细胞MRP基因的表达明显高于亲本细胞.结论 人膀胱癌耐顺铂细胞株T24/DDP具有多药耐药性,可以作为研究膀胱癌细胞多药耐药的良好实验模型.     

Sorcin的表达影响人胶质瘤细胞对顺铂的敏感性

 目的： 研究可溶性耐药相关钙结合蛋白(sorcin)在人胶质瘤细胞对顺铂敏感性中的作用。方法：构建pSilencerTM 3.1-H1-sorcin siRNA表达质粒;质粒转染人胶质瘤U251细胞;RT-PCR和Western blotting法检测sorcin mRNA和蛋白表达的变化;MTT法检测U251细胞的生存率;Western blotting检测耐药相关蛋白的表达变化。结果：经酶切和测序鉴定,成功构建pSilencerTM3.1-H1-sorcin siRNA表达载体;将该表达载体转染U251细胞,RT-PCR和Western blotting结果显示sorcin mRNA和蛋白表达量降低（P<0.05）;MTT结果显示,抑制sorcin表达可增强U251细胞对顺铂的敏感性（P< 0.05）;同时发现抑制U251细胞的sorcin表达能降低耐药相关蛋白P-糖蛋白(P-gp)和多药耐药相关蛋白1(MRP1)的蛋白水平（P< 0.05）。结论：抑制sorcin表达增强U251细胞对顺铂的敏感性,其作用机制可能与降低耐药相关蛋白P-gp和MRP1的表达有关,提示sorcin可能与胶质瘤细胞顺铂耐药有关。     

HOXB7 induces STAT3-mediated transformation and lung metastasis in immortalized mammary gland NMuMG cells

The homeobox family genes are often dysregulated in various cancer types. Particularly HOXB7 amplification and overexpression correlate with poor prognosis in various cancer such as gastric, pancreatic, and lung cancers. Moreover, HOXB7 is known to contribute to cancer progression by promoting epithelial to mesenchymal transition, anticancer drug resistance, and angiogenesis. In this study, we show that HOXB7 is coamplified with ERBB2 in a subset of breast cancer patients and HOXB7 expression correlates with poor prognosis in HER2-positive breast cancer patients. This clinical observation is supported by the following results—HOXB7 overexpression in an immortalized murine mammary gland epithelial cell line NMuMG induces cellular transformation in vitro, tumorigenesis, and lung metastasis through the activation of JAK-STAT signaling.     

MicroRNA-196a通过HOXB7调控人骨髓间充质干细胞的增殖功能

 目的:研究年龄相关microRNA-196a (miR-196a) 对人骨髓间充质干细胞（hMSCs）增殖功能的调控作用。方法:通过MTT研究年龄对hMSCs增殖能力的影响。通过microRNA芯片和qRT-PCR检测年龄对miR-196a表达的影响。通过转染miR-196a模拟物或抑制物,研究其对hMSCs增殖能力的影响。通过萤光素酶报告基因系统证实HOXB7为miR-196a的靶基因。通过siRNA研究HOXB7对碱性成纤维细胞生f长因子(bFGF)表达及hMSCs增殖功能的影响和研究bFGF对hMSCs增殖功能的影响。结果:随年龄增加,hMSCs的增殖能力下降,miR-196a的表达增加。miR-196a可抑制hMSCs的增殖。抑制miR-196a的表达可促进hMSCs的增殖。同时抑制miR-196a和HOXB7的表达,使miR-196a失去对hMSCs增殖能力的调控作用。抑制HOXB7的表达可使bFGF的表达下调。直接抑制HOXB7或bFGF的表达可抑制hMSCs的增殖。结论:miR-196a通过抑制HOXB7及bFGF的表达导致hMSCs增殖能力下降。     

The significance of HOXB7 and IL17RB serum levels in prognosis of hormonally dependent breast cancer: A pilot study

PurposeImproved prognostication of a patient's outcome could allow for personalized treatment decisions in breast cancer. Homeobox B7 (HOXB7) and interleukin 17 receptor B (IL17RB) are proteins reportedly involved in the development of hormonal therapy resistance. Their prognostic value was previously investigated in tumor tissue but recent mass spectrometric detection of HOXB7 and IL17RB proteins in serum has prompted us to perform the first prognostic evaluation of their serum levels.Patients and methodsThe study included 81 premenopausal breast cancer patients that received adjuvant hormonal therapy. The median follow-up period was 61 months. HOXB7 and IL17RB serum protein levels were measured by quantitative sandwich ELISA and prognostically evaluated by Cox proportional hazards regression analysis.ResultsHOXB7 protein was detected in 96.3% and IL17RB in 33.3% of serum samples. Higher levels of serum HOXB7 significantly associated with favorable disease outcome by prognosticating distant (by HR ​= ​0.04; P ​= ​0.001) and local recurrence (by HR ​= ​0.03, P ​= ​0.001). The recurrence rates in the HOXB7^high and HOXB7^low subgroups of patients (cut-off 81.5 ​pg/mL) were 0% and 17%, respectively. Serum IL17RB levels did not significantly associate with either local or distant events. The multivariate analysis highlighted estrogen receptor, histological grade, nodal status and HOXB7 as independent prognostic parameters.ConclusionsOur findings validate the previous mass-spectrometry data by showing that HOXB7 and IL17RB cellular proteins are detectable in serum by a standard ELISA assay. Furthermore, we show that HOXB7 serum levels are the relevant prognosticator of response to hormonal therapy.     

MST1过表达抑制宫颈癌细胞系SiHa的增殖、迁移和侵袭

目的探讨哺乳动物不育系20样激酶1(MST1)对子宫颈癌SiHa细胞增殖、迁移和侵袭能力的影响。方法Western blot检测正常宫颈上皮细胞H8与宫颈癌细胞SiHa中MST1的表达;构建p J3H-HA-MST1质粒并转染SiHa细胞系,Western blot检测MST1、Ki-67和MMP9的蛋白表达;MTS、划痕实验和Transwell分别检测细胞增殖、迁移和侵袭能力。结果 SiHa细胞的MST1蛋白表达水平明显低于H8细胞(P0.05);SiHa细胞转染MST1质粒后,MST1蛋白表达明显升高,而Ki-67和MMP9蛋白表达水平显著下降(P0.05),SiHa细胞的增殖被明显抑制(P0.05),同时细胞的迁移和侵袭能力也显著降低(P0.01)。结论 MST1过表达可以抑制宫颈癌细胞SiHa的增殖、迁移和侵袭能力。     

Silencing of CCR7 inhibits the growth,invasion and migration of prostate cancer cells induced by VEGFC

Early in prostate cancer development, tumor cells express vascular endothelial growth factor C (VEGF-C), a secreted molecule that is important in angiogenesis progression. CC-chemokine receptor 7 (CCR7), another protein involved in angiogenesis, is strongly expressed in most human cancers, where it activated promotes tumor growth as well as favoring tumor cell invasion and migration. The present study aimed to investigate the effect of down-regulating CCR7 expression on the growth of human prostate cancer cells stimulated by VEGFC. The CCR7-specific small interfering RNA (siRNA) plasmid vector was constructed and then transfected into prostate cancer cells. The expression of CCR7 mRNA and protein was detected by quantitative polymerase chain reaction and western blot analysis, respectively. Cell proliferation, apoptosis, cell cycle distribution and cell migration were assessed following knockdown of CCR7 by RNA interference (RNAi). Western blot analysis was used to identify differentially expressed angiogenesis- and cell cycle-associated proteins in cells with silenced CCR7. The expression levels of CCR7 in prostate cancer cells transfected with siRNA were decreased, leading to a significant inhibition of prostate cancer cell proliferation, migration and invasion induced by VEGFC. Western blot analysis revealed that silencing of CCR7 may inhibit vascular endothelial growth factor, matrix metalloproteinase (MMP)-2 and MMP-9 protein expression. In conclusion, the present study demonstrated that RNAi can effectively silence CCR7 gene expression and inhibit the growth of prostate cancer cells, which indicates that there is a potential of targeting CCR7 as a novel gene therapy approach for the treatment of prostate cancer.     

脂质体紫杉醇联合顺铂对晚期宫颈癌术前化疗的疗效观察

目的探讨脂质体紫杉醇联合顺铂用于治疗局部晚期宫颈癌患者的临床疗效和不良反应。方法选择2005年5月至2009年5月我院收治的局部晚期宫颈癌患者78例,按就诊顺序将患者分为顺铂联合5-氟尿嘧啶组(PF组,39例)和脂质体紫杉醇联合顺铂组(Taoxl组,39例)。PF组患者采用顺铂联合5-氟尿嘧啶化疗方案,Taxol组患者采用脂质体紫杉醇联合顺铂化疗方案。2组患者化疗后均全麻下行广泛子宫切除术和盆腔淋巴结清扫术,评价并比较2组患者的临床疗效、不良反应和术后3年生存率。结果 PF组患者不良反应主要为消化道反应、口腔黏膜炎和骨髓抑制,Taxol组患者的不良反应主要有骨髓抑制、口腔黏膜炎和肾脏毒性。Taxol组患者的化疗后完全缓解率30.77%,高于PF组的10.26%(χ2=5.032,P=0.025)。Taxol组和PF组患者平均生存时间分别是33.5个月和34.7个月,3年生存率分别为59.6%和70.2%,差异无统计学意义(χ2=1.859,P=0.173)。结论 2种化疗方案患者不良反应差别较大,脂质体紫杉醇联合顺铂化疗方案临床疗效优于顺铂联合5-Fu化疗方案。     

Decreased expression of lncRNA GAS5 predicts a poor prognosis in cervical cancer

Introduction: Cervical cancer is the second leading cause of cancer morbidity and mortality for women around the world. Long non-coding RNAs (lncRNAs) have been investigated as a new class of regulators of cellular processes, such as cell growth, apoptosis, and carcinogenesis. Although downregulation of lncRNA GAS5 in several cancers has been studied, its role in cervical cancer remains unknown. The aim of this study is to investigate the expression, clinical significance and biological role in cervical cancer. Methods: Expression of GAS5 was analyzed in cervical cancer tissues by quantitative Real-time PCR (qRT-PCR). And its association with overall survival of patients was analyzed by statistical analysis. Small interfering RNA (siRNA) was used to suppress GAS5 expression in cervical cancer cells. In vitro assays were performed to further explore the biological functions of GAS5 in cervical cancer. Results: We found that GAS5 expression was markedly downregulated in cervical cancer tissues than in corresponding adjacent normal tissues. Decreased GAS5 expression was significantly correlated with FIGO stage, vascular invasion and lymph node metastasis. Moreover, cervical cancer patients with GAS5 lower expression have shown significantly poorer overall survival than those with higher GAS5 expression. And GAS5 expression was an independent prognostic marker of overall survival in a multivariate analysis. In vitro assays our data indicated that knockdown of GAS5 promoted cell proliferation, migration, and invasion. Conclusions: Our study presents that lncRNA GAS5 is a novel molecule involved in cervical cancer progression, which provide a potential prognostic biomarker and therapeutic target.     

粘附分子JAM-A与人宫颈癌的侵袭性

目的观察粘附分子JAM-A在宫颈癌不同肌层侵袭时的表达情况,探讨其与癌细胞侵袭性的关系。方法 35例宫颈癌患者标本,其中癌细胞侵袭浅肌层15例,侵袭深肌层20例,采用免疫组化法观察JAM-A在宫颈癌浅、深两组侵袭深度以及在癌团癌细胞和孤立癌细胞时的表达情况。结果 JAM-A在宫颈癌癌细胞、血管管腔内皮细胞间连接、腺腔内皮细胞连接及血细胞表达,JAM-A在浅肌层癌细胞侵袭组的表达强于深肌层侵袭组(P0.001);JAM-A在癌团的癌细胞表达显著强于孤立癌细胞(P0.0001)。结论宫颈癌JAM-A的表达与癌细胞的侵袭程度呈负相关。