RNA激活技术介导的E—cadherin基因上调表达抑制5637膀胱癌细胞侵袭与迁移

目的探讨通过RNA激活（RNA activation,RNAa）技术上调E-cadherin基因的表达而影响人膀胱癌5637细胞的侵袭、迁移能力。方法将与E-cadherin基因启动子DNA序列互补的双链RNA分子（dsEcad）转染入5637细胞中,采用RT-PCR法及Westernblot检测E-cadherin的表达;用Transwell小室法及划痕法检测RNAa后细胞侵袭、迁移能力的改变;用细胞免疫荧光法及Western blot检测β-catenin蛋白在细胞内的重新分布结果。结果dsEcad转染5637细胞72h后E-cadherin表达显著上调,RNAa有效;5637细胞对细胞外基质的侵袭能力及迁移能力也明显下降。β-catenin在胞核及细胞质的水平明显下降并重新再分布至细胞膜上。结论RNAa技术激活E-cadherin基因表达并抑制细胞侵袭、转移等恶性行为,可作为膀胱癌或其他恶性肿瘤基因治疗的一种有效手段。     

High level of ezrin expression in colorectal cancer tissues is closely related to tumor malignancy

AIM： To investigate the ezrin expression in normal colorectal mucosa and colorectal cancer tissues, and study the correlation between ezrin expression in colorectal cancer tissues and tumor invasion and metastasis. METHODS： Eighty paraffin-embedded cancer tissue samples were selected from primary colorectal adenocarcinoma. Twenty-eight patients had well- differentiated, 22 had moderately differentiated and 30 had poorly differentiated adenocarcinoma. Forty-five patients and 35 patients had lymph node metastasis. Forty-five patients were of Dukes A to B stage, and 35 were of C to D stage. Another 22 paraffin-embedded tissue blocks of normal colorectal epithelium （〉 5 cm away from the edge of the tumor） were selected as the control group. All patients with colorectal cancer were treated surgically and diagnosed histologically, without preoperative chemotherapy or radiotherapy. The immunohistochemistry was used to detect the ezrin expression in paraffin-embedded normal colorectal mucosa tissues and colorectal cancer tissue samples. RESULTS： Ezrin expression in colorectal cancer was significantly higher than in normal colorectal mucosa （75.00% vs 9.09%, P 〈 0.01）, and there was a close relationship between ezrin expression and the degree of tumor differentiation, lymph node metastasis and Dukes stage （88.46% vs 50.00%, P 〈 0.01; 94.28% vs 51.11%, P 〈 0.01; 94.28% vs 51.11%, P 〈 0.01）. CONCLUSION： Ezrin expression is obviously higher in colorectal cancer tissues than in normal colorectal mucosa tissues, and the high level of ezrin expression is closely related to the colorectal cancer invasion and metastasis process.     

阻断FAK表达对涎腺腺样囊性癌细胞SACC—LM的侵袭行为的影响

目的：研究探讨阻断黏着斑激酶（focal adhesion kinase,FAK）表达对涎腺腺样囊性癌肺转移亚系细胞（salivary adenoid cystic carcinoma-Lung metastasis,SACC—LM）侵袭行为的影响及相关机制。方法：应用酪氨酸蛋白激酶抑制剂Genistein处理体外培养的SACC-LM细胞,应用transwell小室观察Genistein对SACC—LM细胞侵袭的影响,并应用RT-PCR法检测FAK和细胞外信号调节激酶（extracellular signal—regulated kinase,ERK）mRNA的表达。所得实验数据采用SPSS10．0统计软件t检验进行分组分析。结果：Genistein使SACC-LM细胞侵袭能力减弱;检测到随着Genistein抑制剂浓度的增高和作用时间的延长,FAK和ERK mRNA的表达水平均降低。结论：阻断FAK表达导致SACC-LM细胞侵袭能力减弱,原因可能是FAK和ERK表达水平的改变。     

GAS5对骨肉瘤U2OS细胞增殖、迁移和侵袭的影响

目的:观察生长停滞特异性转录因子5(GAS5)对骨肉瘤U2OS细胞增殖、迁移和侵袭的影响,探讨其机制。方法:在人骨肉瘤U2OS细胞和人正常成骨hFOB 1.19细胞中,采用实时定量PCR(qRT-PCR)检测GAS5和微小RNA (miR)-221的表达,Western blot检测基质金属蛋白酶抑制物-2 (TIMP2)蛋白的表达。转染pcDNA-GAS5重组质粒后,采用噻唑蓝(MTT)法和Trasnwell小室法检测GAS5对U2OS细胞增殖、迁移和侵袭的影响,qRT-PCR检测GAS5对miR-221表达的影响。利用Starbase软件预测和双荧光素酶报告基因实验验证GAS5与miR-221以及miR-221与TIMP2的靶向关系。转染miR-221模拟物和miR-221抑制剂后,观察miR-221过表达对GAS5调控的U2OS细胞增殖、迁移和侵袭的影响以及miR-221对TIMP2蛋白表达的影响。结果:与正常hFOB 1.19细胞相比,U2OS细胞中GAS5和TIMP2蛋白的表达水平明显降低,而miR-221表达水平显著升高(GAS5:P=0.0001;TIMP2:P=0.0003;miR-221:P=0.0004)。GAS5过表达可抑制U2OS细胞增殖、迁移和侵袭(增殖48h:P=0.0005;增殖72h:P=0.0002;迁移:P=0.002;侵袭:P=0.001),而miR-221过表达可明显逆转GAS5对U2OS细胞增殖、迁移和侵袭的抑制作用(增殖48h:P=0.0002;增殖72h:P=0.0003;迁移:P=0.0001;侵袭:P=0.0001)。Starbase软件预测到miR-221存在能够与GAS5互补结合的位点,还存在能够与TIMP2 3′UTR互补结合的位点;双荧光素酶结果显示miR-221过表达后野生型GAS5(GAS5-WT)和野生型TIMP2 (TIMP2-WT)细胞的荧光素酶活性明显降低(P均为0.0003);同时,GAS5过表达后,U2OS细胞中miR-221表达水平明显降低(P=0.0001),反之miR-221明显升高(P=0.0001);miR-221过表达后,U2OS细胞中TIMP2蛋白的表达水平明显降低(P=0.0003),反之TIMP2蛋白的表达水平明显升高(P=0.0009)。结论:GAS5可通过靶向抑制miR-221促进TIMP2表达,进而抑制U2OS细胞的增殖、迁移和侵袭。     

JAZF1对胶质母细胞瘤侵袭和迁徙的影响及分子机制研究

目的探讨JAZF1对脑胶质母细胞瘤（GBM）细胞侵袭、迁徙的作用及相关作用机制。 方法通过挖掘TCGA数据库中不同级别胶质瘤患者JAZF1的转录水平差异以及患者生存曲线差异,然后通过科内收集的胶质瘤样本提取总蛋白进行Western blot实验研究JAZF1蛋白水平在胶质瘤不同级别中的差异,通过Trans-well实验、细胞划痕实验研究JAZF1对胶质瘤细胞侵袭和迁移的影响,通过Western blot实验研究JAZF1对胶质瘤细胞侵袭和迁徙能力的关键分子的影响。 结果（1）Ⅳ级胶质瘤中JAZF1的mRNA、蛋白水平的表达上相较于Ⅱ级和Ⅲ级胶质瘤明显更高,差异具有统计学意义（P<0.05）;（2）JAZF1高表达组患者的中位生存期为13.0个月,而低表达组中位生存期为30.2个月,差异具有统计学意义（P<0.05）;（3）JAZF1表达下降后,U87M细胞的侵袭和迁徙能力均显著下降,差异具有统计学意义（P<0.05）;（4）干扰JAZF1的表达后,p-Akt、MMP9、MMP2的表达水平显著下调,差异均具有统计学意义（P<0.05）,而Akt的表达变化不明显,差异无统计学意义（P>0.05）。 结论JAZF1在GBM中高表达且显著缩短了患者的中位生存期,通过调控Akt的磷酸化水平进一步调控相应信号通路下游的MMP9、MMP2的表达影响GBM的侵袭和迁徙能力。     

MicroRNA-377 Targets Zinc Finger E-box-Binding Homeobox 2 to Inhibit Cell Proliferation and Invasion of Cervical Cancer

A large number of microRNAs (miRNAs) are aberrantly expressed in cervical cancer and play crucial roles in the onset and progression of cervical cancer by acting as either an oncogene or a tumor suppressor. Therefore, investigation of the expression, biological roles, and underlying mechanisms of miRNAs in cervical cancer might provide valuable therapeutic targets in the treatment for patients with this disease. In this study, miRNA- 377 (miR-377) was downregulated in cervical cancer tissues and cell lines. Decreased miR-377 expression was strongly correlated with the International Federation of Gynecology and Obstetrics (FIGO) stage, lymph node metastasis, and distant metastasis in patients with cervical cancer. Enhanced expression of miR-377 prohibited cell proliferation and invasion in cervical cancer. Bioinformatics analysis predicted that zinc finger E-boxbinding homeobox 2 (ZEB2) was a potential target of miR-377. Subsequent experiments confirmed that ZEB2 is a direct target gene of miR-377 in cervical cancer. In addition, ZEB2 was overexpressed in cervical cancer tissues and was inversely related with miR-377 levels. Furthermore, the suppressive effects of miR-377 on cervical cancer proliferation and invasion were rescued by restored ZEB2 expression. Overall, our findings indicated that miR-377 decreases proliferation and invasion of cervical cancer cells by directly targeting ZEB2 and provides novel evidence of miR-377 as a novel therapeutic strategy for the therapy of patients with this malignancy.     

棕榈酸在体外通过促进凋亡作用抑制宫颈癌Hela细胞增殖的研究

目的 探讨体外棕榈酸(Palmitic acid,PA)抑制人宫颈癌HeLa细胞增殖、迁移、侵袭和诱导凋亡作用及其部分机制。方法 体外培养的宫颈癌HeLa细胞分为对照组(不加药)、PA处理组(分别加入PA 25、50、100μg/mL处理),CCK-8法检测各组HeLa细胞增殖情况,Transwell试验检测细胞的迁移和侵袭能力,qRT-PCR检测不同浓度PA处理后细胞凋亡相关基因Bcl-2、Bax及Cleaved-caspase-3的表达,PA处理后的HaLa细胞用膜联蛋白V/碘化丙啶双标,随后用流式细胞仪检测其凋亡情况。结果 与对照组比较,PA处理HeLa细胞后增殖抑制率逐渐增高(P＜0.05);PA处理后,HaLa细胞的迁移与侵袭能力显著下降(P＜0.05);同时,PA处理HeLa细胞后,随着PA浓度增高,Bcl-2基因表达逐渐降低,而Bax和Cleaved-caspase-3基因表达逐渐增强与流式细胞仪检测结果相符合,HeLa细胞的凋亡率随着PA浓度增高逐渐增高(P＜0.05)。结论 PA体外可抑制HeLa细胞增殖,减少其迁移与侵袭水平,并诱导其凋亡,其机制可能与调节HeLa细胞Bcl-2家族有关。     

老年人胃癌的浸润深度和术后化疗对预后的影响

目的探讨接受手术治疗的老年人胃癌临床病理因素与预后的关系。方法回顾性研究本院2002年1月至2005年12月胃癌手术患者,记录患者的年龄、性别、肿瘤位置、肿瘤大小、共存病（高血压及心脏病）、手术根治程度、胃切除范围、病理类型(WHO)、TNM分期、术后化疗情况,建立数据库。结果老年组中位生存时间为29月（平均44月）,明显低于中年组（78月,P=0.001）和青年组（72月,P=0.009）;老年患者原发肿瘤以T3/T4为主;较少的患者接受联合脏器切除;更多合并高血压及心脏病,而较少患者接受术后化疗。结论老年人胃癌手术患者原发灶的侵犯深度、术后较少接受化疗与预后差有关。