Transactivation of the TIEG1 confers growth inhibition of transforming growth factor-β-susceptible hepatocellular carcinoma cells

AIM:To investigate the role of transforming growth factor(TGF)-β-inducible early gene 1(TIEG1) in TGF-β-induced growth inhibition in hepatocellular carcinoma(HCC) cells.METHODS:Human hepatocyte and HCC cell lines with varied susceptibilities to TGF-β1 were tested by methylthiazoletetrazolium(MTT) assay.The expression changes of Smad2,Smad3,Smad4,Smad7,TIEG1 and TIEG2 gene following treatment with TGF-β1 in a TGF-β-sensitive hepatocyte cell line(MIHA),a TGF-β-sensitive hepatoma cell line(Hep3B) and two TGF-β-insensitive hepatoma cell lines(HepG2 and Bel7404) were examined.SiRNA targeting TIEG1 was transfected into Hep3B cells and the sensitivity of cells to TGF-β1 was examined.Overexpression of TIEG1 was induced by lentiviral-mediated transduction in TGF-β1-resistant hepatoma cell lines(Bel7404 and HepG2).MTT assay and 4',6-Diamidino-2-phenylindole staining were used to identify cell viability and apoptosis,respectively.The expression level of stathmin was measured by reverse transcriptase polymerase chain reaction and Western-blotting analysis,and stathmin promoter activity by TIEG1 was monitored by a luciferase reporter gene system.RESULTS:TIEG1 was significantly upregulated by TGF-β1 in the TGF-β1-sensitive HCC cell line,Hep3B,but not in the resistant cell lines.The suppression of TIEG1 by siRNAs decreased the sensitivity of Hep3B cells to TGF-β1,whereas the overexpression of TIEG1 mediated growth inhibition and apoptosis in TGF-β1-resistant HCC cell lines,which resembled those of TGF-β1-sensitive HCC cells treated with TGF-β1.Our data further suggested that stathmin was a direct target of TIEG1,as stathmin was signif icantly downregulated by TIEG1 overexpression,and stathmin promoter activity was inhibited by TIEG1 in a dose-dependent manner.CONCLUSION:Our data suggest that transactivation of TIEG1 conferred growth inhibition of TGF-β-susceptible human HCC cells.

Refined mapping of loss of heterozygosity on 1q31.1-32.1 in sporadic colorectal carcinoma

AIM: To explore precise deleted regions and screen the candidate tumor suppressor genes related to sporadic colorectal carcinoma. METHODS: Six markers on 1q31.1-32.1 were chosen. These polymorphic microsatellite markers in 83 colorectal cancer patients tumor and normal DNA were analyzed via PCR. PCR products were electrophoresed on an ABI 377 DNA sequencer. Genescan 3.1 and Genotype 2.1 software were used for Loss of heterozygosity (LOH) scanning and analysis. Comparison between LOH frequency and clinicopathological factors was performed by χ2 test. RESULTS: 1q31.1-32.1 exhibited higher LOH frequency in colorectal carcinoma. The average LOH frequency of 1q31.1-32.1 was 23.0%, with the highest frequency of 36.7% (18/49) at D1S2622, and the lowest of 16.4% (11/67) at D1S412, respectively. A minimal region of frequent deletion was located within a 2 cM genomic segment at D1S413-D1S2622 (1q31.3-32.1). There was no significant association between LOH of each marker on 1q31.1-32.1 and the clinicopathological data (patient sex, age, tumor size, growth pattern or Dukes stage), which indicated that on 1q31.1-32.1, LOH was a common phenomenon in all kinds of sporadic colorectal carcinoma. CONCLUSION: Through our refined deletion mapping,the critical and precise deleted region was located within 2 cM chromosomal segment encompassing 2 loci (D1S413, D1S2622). No significant association was found between LOH and clinicopathologic features in 1q31.1-32.1.

Genetic variation of TGF-BR2 as a protective genotype for the development of colorectal cancer in men

BACKGROUND The role of transforming growth factor beta(TGF-β)signaling,including both the cytokine and their receptors,in the etiology of colorectal cancer(CRC)has been of particular interest lately.AIM To investigate the association between promoter polymorphism in TGF-β receptor 2 TGF-BR2G^([-875])A with a CRC risk in a cohort of Bulgarian patients using a casecontrol gene association study approach,as well as the protein levels of TGF-β1 in the peripheral blood.METHODS A cohort of 184 CRC patients and 307 sex and age-matched healthy subjects were recruited in the study.A genotyping of the TGF-BR2G^([-875])A(rs3087465)polymorphism was performed by primer-introduced restriction analysespolymerase chain reaction approaches.RESULTS The frequency of TGF-BR2G^([-875])A genotype was decreased in male patients with CRC than in healthy men(31.3%vs 44.8%;P=0.058).Among males,the TGF-BR2G[-509]G genotype was related to a significantly increased risk of CRC development(OR=1.820,95%CI:0.985-3.362,P=0.055)than the GA+AA genotype.Also,TGF-BR2^([-875])*A-allele itself was rarer in men with CRC than healthy men(19.1%vs 26.9%,P=0.086)and was associated with a protective effect(OR=0.644;95%CI:0.389-1.066;P=0.086).Regarding the genotypes,we found that TGF-β1 serum levels were higher in GG genotype in healthy persons above 50 years than the CRC patients[36.3 ng/mL interquartile range(IQR)19.9-56.5 vs 22.4 ng/mL IQR 14.8-29.7,P=0.014].We found significant differences between higher levels of TGF-β1 serum levels in healthy controls above 50 years(GG genotype)and CRC patients(GG genotype)at the early stage(36.3 ng/mL IQR 19.9-56.5 vs 22.8 ng/mL IQR 14.6-28.6,P=0.037)and advanced CRC(36.3 ng/mL IQR 19.9-56.5 vs 21.6 ng/mL IQR 15.9-33.9,P=0.039).CONCLUSION In summary,our results demonstrated that TGF-BR2 AG and AA genotypes were associated with a reduced risk of CRC,as well as circulating levels of TGF-βcould prevent CRC development in a gender-specific manner.Notably,male carriers of TGF-BR2-875A allele genotypes had a lower risk of CRC development and progression,suggesting that TGF-BR2-875A/G polymorphism significantly affects the protective biological factors that also impact the risk of colon and rectal carcinogenesis.

Detailed Deletion Mapping of Chromosome 9p21-22 in Nasopharyngeal Carcinoma

Objective: To further refine the extent of deletion on chromosome 9p21-22 in nasopharyngeal carcinoma (NPC) and provide evidence for discovering new tumor suppressor gene. Methods: Loss of heterozygosity (LOH) on chromosome 9p21-22 was analyzed in 25 paired blood and tumor samples by using 11 high-density microsatellite polymorphic markers. Results: 17 of 25 cases (68.0%) showed LOH at one or more loci. Higher frequencies of LOH were found at four loci: D9S161 (35.0%), D9S1678 (31.5%), D9S263 (33.3%) and D9S1853 (33.3%), where 6 cases had a contiguous stretch of allelic loss. Conclusion: The minimal common region of deletion might be defined between D9S161 and D9S1853 (estimated about 2.7 cM in extent) at 9p21.1, suggesting that inactivation of one or more tumor suppressor genes located in this region may be an important step in NPC.

共载吲哚菁绿和Nrf2-siRNA的多功能纳米粒子对抗口腔鳞状细胞癌的体外作用研究

目的:构建共载吲哚菁绿(indo cyanine green,ICG)和核因子E2相关因子2(nuclear factor erythroid 2-related factor 2,Nrf2)-短干扰RNA(small interfering RNA,siRNA)的多功能纳米载药粒子,并探究其联合光热疗法与抗氧化抑制作用增效的光动力疗法协同对抗口腔鳞状细胞癌(oral squamous cell carcinoma,OSCC)的体外作用与机制。方法:利用超声乳化法和静电力吸附作用制备基于聚氨基酯/聚乳酸-羟基乙酸共聚物且共载ICG和Nrf2-siRNA的纳米粒子PPI-siRNA。表征PPI-siRNA纳米粒子的形貌、粒径大小和分布、表面电位及其载药情况;评价PPI-siRNA纳米粒子对ICG和Nrf2-siRNA的胞内递送以及Nrf2-siRNA逃逸溶酶体吞噬的性能;通过检测舌鳞状细胞癌细胞SCC-25随激光照射的升温效应、热休克蛋白60表达和活性氧生成水平考察PPI-siRNA纳米粒子的光热和光动力性能;考察细胞内Nrf2及其调控的下游基因谷氨酸-半胱氨酸连接酶修饰亚基和谷氨酸-半胱氨酸连接酶的表达水平,从而探究Nrf2-siRNA助力光动力效应的作用机制;运用噻唑蓝比色法考察PPI-siRNA联合激光照射对SCC-25的细胞毒性作用。结果:PPI-siRNA纳米粒子呈形貌规则的球状结构,粒径约为180 nm,并成功共载ICG与Nrf2-siRNA;PPI-siRNA纳米粒子可以高效递送ICG和Nrf2-siRNA入胞,且可以确保Nrf2-siRNA逃逸溶酶体吞噬,从而发挥基因沉默作用;PPI-siRNA纳米粒子具备良好的光热和光动力性能,Nrf2-siRNA可以通过下调抗氧化蛋白和基因的表达显著提升光学疗法的抗肿瘤效率。结论:PPI-siRNA纳米粒子可以联合光热疗法与基因沉默作用增效的光动力疗法并有效抑制SCC-25的体外生长,在OSCC的临床治疗方面具有广阔的应用前景。

白芍总苷对肝癌耐药细胞BEL-7402/ADM耐药性的逆转作用及机制探讨

目的观察白芍总苷(TPG)对肝癌耐药细胞BEL-7402/ADM耐药性的逆转作用,并探讨其作用机制。方法体外培养人肝癌ADM敏感细胞BEL-7402(亲本组)和ADM耐药细胞BEL-7402/ADM(耐药组),将BEL-7402/ADM细胞转染HDAC3基因siRNA干扰质粒(干扰组)及空载质粒(空载组)24 h,用白芍总苷孵育BEL-7402/ADM细胞48 h(用药组)。取亲本组、耐药组、用药组细胞,分别以不同浓度ADM作用48 h;MTT法测算增殖抑制率,计算各组ADM的半数抑制浓度(IC_(50))。取亲本组、耐药组、空载组、干扰组及用药组细胞,分别采用RT-q PCR法和Western blot法检测细胞多药耐药基因MDR1和去乙酰化酶(HDAC3)mRNA和蛋白。结果亲本组、耐药组和用药组细胞ADM的IC_(50)分别为0.139、8.807、4.890 nmo L/L;用药组细胞对ADM耐药的逆转倍数为1.80倍,逆转率为45.23%。MDR1 mRNA及蛋白相对表达量耐药组、空载组>用药组>干扰组>亲本组(P均<0.05或0.01),HDAC3 mRNA及蛋白相对表达量耐药组、空载组>干扰组>亲本组、用药组(P均<0.05或0.01)。结论白芍总苷通过抑制HDAC3的表达,引起MDR1表达下调,从而逆转BEL-7402/ADM细胞的耐药性。

视黄酸对胃癌细胞生长和p53及c-myc基因表达的影响

目的：探讨视黄酸对胃癌细胞生长和基因表达的影响。方法：以ＭＴＴ法测定细胞生长，软琼脂集落形成实验测定癌细胞恶性程度，Ｎｏｒｔｈｅｒｎｂｌｏｔ测定ｐ５３及ｃｍｙｃ基因表达水平。结果：１０－６ｍｏｌ／Ｌ全反式视黄酸（ＡＴＲＡ）能够有效地抑制ＭＧｃ８０３和ＢＧＣ８２３细胞生长，但不能抑制ＭＫＮ４５细胞生长；ＡＴＲＡ还能够降低３株胃癌细胞形成集落的能力。ＭＧｃ８０３和ＢＧＣ８２３细胞经ＡＴＲＡ诱导后，ｐ５３基因的表达水平提高，ｃｍｙｃ基因的表达水平下降，但ＭＫＮ４５细胞中ｐ５３基因和ｃｍｙｃ基因表达与ＡＴＲＡ诱导无关。视黄酸受体ＲＡＲα基因在３株细胞中呈低水平表达，经ＡＴＲＡ诱导后，在ＭＧｃ８０３和ＢＧＣ８２３细胞中表达水平提高，但在ＭＫＮ４５细胞中表达水平不变。

下调缺氧诱导因子-1表达对肝癌细胞增殖和凋亡的影响

目的:观察肝癌缺氧诱导因子-1(hypoxia inducible factor-1alpha,HIF-1)表达及靶向其基因转录对肝癌细胞增殖的影响.方法:以自身配对法收集术后肝癌及癌周组织,免疫组织化学染色法分析HIF-1胞内表达及分布;按照HIF-1基因序列合成miRNA,筛选干扰效率最佳者转染肝癌(HepG2)细胞,以荧光定量-PCR和Western blot分别检测HIF-1转录和蛋白表达;并以miRNA转染联合阿霉素后,流式细胞仪检测沉默细胞中HIF-1对HepG2细胞增殖的影响.结果:肝癌及癌周组织HIF-1表达呈棕黄色颗粒状,位于胞浆和部分胞核,呈均匀表达,中央静脉周围明显;其阳性率癌组织为80%,低于癌旁100%表达(2=22.35,P<0.001);与瘤体大小和分化程度相关;HepG2细胞经miRNA干扰后,HIF-1在RNA和蛋白水平分别下降87%和56%,发生细胞凋亡(22.46%±0.61%)和G1期阻滞(61.49%±1.12%,P<0.01);加入阿霉素后细胞凋亡率和G1期细胞分别增至36.99%±0.88%和65.68%±0.91%.结论:HIF-1过表达与肝癌密切相关,以特异性miRNA靶向HIF-1,可调控细胞周期,通过加速凋亡的机制抑制癌细胞增殖.

口腔疣状癌中survivin与BCL-2、Skp2与P27两组蛋白表达特点及相关性研究

目的:研究口腔疣状癌(OVC)组织中survivin(生存蛋白)与BCL-2(B淋巴瘤/白血病-2基因)、Skp2(S期激酶蛋白-2)与P27(激酶抑制蛋白)两组蛋白的表达特点及其相关性。方法:以本院2012-01-2015-01期间收治的34例口腔疣状癌患者作为本次研究资料,全部患者均在本院接受手术治疗。将术中取得的口腔疣状癌组织、癌旁组织,以及2015年1月间在本院因良性病变行病理检查患者的正常口腔黏膜组织作为研究标本,测定3种口腔黏膜组织中survivin、BCL-2、Skp2、P27的蛋白表达水平并进行对比分析,分析不同类型、不同分期的口腔疣状癌组织中4项蛋白表达的差异性,研究survivin与BCL-2、Skp2与P27之间的相关性,并据此研究4项蛋白表达水平对于口腔疣状癌诊断治疗的意义。结果:口腔疣状癌组织中Survivin、BCL-2、Skp2、P27表达阳性率均显著高于癌旁组织与正常组织,P<0.01;经进一步两两对比分析,癌旁组织中4项蛋白表达的阳性率与正常组织无明显差异,P>0.05。Survivin在口腔疣状癌组织中的阳性表达与BCL-2的阳性表达呈显著正相关性,P<0.01,0<r<1;Skp2在口腔疣状癌组织中的阳性表达与P27的阳性表达呈显著负相关性,P<0.01,-1<r<0。不同肿瘤类型、不同分期口腔疣状癌组织中,Survivin、BCL-2、Skp2、P27阳性表达率差异具有统计学差异性,P<0.05。结论:survivin、BCL-2、Skp2、P27在口腔疣状癌中均可见特异性表达,Survivin与BCL-2在口腔疣状癌组织中的阳性表达以及Skp2与P27在OVC癌组织中的阳性表达均具有明确相关性。

nm23-H1基因对口腔癌细胞化疗敏感性影响的体外实验

目的 通过 nm2 3- H1的转化及导入人舌粘膜鳞癌细胞 (Tca8113细胞株 ) ,观察 nm2 3- H1对 Tca8113细胞株化疗敏感性的影响。 方法 完成细胞培养和基因转染后 ,MTT法观察 nm2 3- H1对 Tca8113细胞株化疗敏感性影响。 结果 转染前 Tca8113细胞株对阿霉素 (ADM)、5 -氟脲嘧啶 (5 - FU)、氨甲喋呤 (MTX)的化疗敏感性无显著差异。但转染后的 Tca8113细胞株对顺铂 (DDP)明显增敏。 结论 nm2 3- H1可能通过特异性地影响细胞内的能量交换过程来达到对

Overexpression of p53 Gene in Esophageal and Cervical Cancer and the Relationship with Radiotherapy Effects

Objective :To investigate the relationship between p53 -protein overexpression in esophageal and cervical squamous cell cancer and their clinical radiosensitivity. Methods: The immuno-histochemical assays were done for 52 cases with esophageal and cervical squamous cell cancer. The relationship between the assay results and short-term radiotherapy was investigated. Results: p53 overexpression was 52. 38% and 35. 48% respectively, in esophageal cancer and cervical cancer; p53 over-expression in high differentiated squamous cell cancer was lower than those in moderate and poor differentiated cases(P<0. 05). There was no relationship between p53 overexpression and stages(P> 0. 05). In the cases of cervical cancer, p53 overexpression had the less short-term effect(P< 0. 05), and In esophageal cancers, there was no relationship with radiotherapy effect(P>0. 05). Conclusion:This study suggests that y53 gene lias the certain relationship with tumor radiosensitivity.

外源性Egr-1基因表达对食管癌细胞株Eca109细胞生长的影响

目的 检测Egr 1蛋白在食管癌细胞株Eca10 9中的表达。方法 采用免疫细胞化学法检测Egr 1蛋白在Eca10 9细胞中的表达 ,用脂质体基因转染法将真核表达载体PCMV Egr 1质粒导入Eca10 9细胞 ,经Westernblot和免疫细胞化学检测 ,Egr 1蛋白在转染的Eca10 9细胞中呈高表达。 结果 Eca10 9细胞的Egr 1蛋白表达阴性 ,当导入外源性Egr 1基因后 ,Egr 1蛋白表达阳性 ,表达Egr 1蛋白的Eca10 9细胞在形态上发生明显的变化 ,表现为细胞变扁 ,呈散在生长。 结论 Egr 1蛋白的表达在一定程度上能抑制Eca10

Study of the SH3-domain GRB2-1ike 2 gene expression in laryngeal carcinoma

Background Laryngeal carcinoma is a common malignant tumor of the upper respiratory tract, and in 95% of cases the tumor is laryngeal squamous cell carcinoma （LSCC）. The abnormity of SH3-domain GRB2-1ike 2 （SH3GL2） gene was found in LSCC. In order to clarify the relationship between SH3GL2 gene and LSCC, we evaluated the expression of the SH3GL2 gene in LSCC. Method Real-time PCR, immunohistochemistry and Western blotting were used to detect the mRNA and protein expression and find the various rules of SH3GL2 gene in LSCC. Results The result of real-time PCR showed that the expression level of SH3GL2 mRNA in LSCC tissue was apparently down-regulated; immunohistochemical analysis showed that SH3GL2 protein was mainly located in cytoplasm, the rate of positive cells and SH3GL2 protein expression level were fluctuated with the pathological classification of LSCC; the result of Western blotting showed that SH3GL2 protein was down-regulated significantly in LSCC samples, especially in metastatic lymph nodes. Conclusions These results suggest that SH3GL2 is a LSCC related gene and its expression level is fluctuated with the pathological classification which indicate that SH3GL2 participates in the development and progression of LSCC. And it may be considered as a novel tumor marker to find both a new anti-oncogene and relative factors of invasion and metastasis of laryngeal carcinoma.

Cyclin G2抑制胃癌细胞增殖的实验研究

目的研究cyclinG2对体外培养胃癌细胞增殖能力的影响,探讨cyclinG2作为肿瘤干扰阻断药靶的有效性和临床使用性。方法应用脂质体转染技术将包含有cyclinG2cDNA的pIRES-G2和对照空质粒pIRES-neo分别转染胃癌细胞株SGC-7901,经G418(400μg/mL)选择性培养基筛选两周后获得转入cyclinG2基因的细胞克隆,通过细胞化学法和免疫蛋白印迹杂交检测外源性cyclinG2蛋白表达情况;倒置显微镜下观察克隆细胞形态,Giemsa染色后进行克隆计数。结果转染pIRES-G2获得的细胞克隆中cyclinG2蛋白的表达水平明显高于转染pIRESneo,表明cyclinG2基因被成功地转入SGC-7901细胞稳定表达;转染pIRES-G2组的阳性细胞集落数目明显减少,pIRESneo组的集落数为(87±3),pIRES-G2组的集落数为(53±4),占对照组的60.1%(P<0.01);与对照组相比,pIRES-G2组集落内细胞数目减少,细胞皱缩,形态变得不规则,胞质内出现大量颗粒和空泡。结论cyclinG2可显著抑制体外培养胃癌细胞的增殖,并可能和肿瘤细胞信号传导系统存在某种联系。

氯化镉诱发16HBE细胞恶性转化不同阶段P16基因甲基化研究

目的对氯化镉(CdCl2)诱发16HBE细胞系恶性转化不同阶段P16基因启动子区甲基化状况进行研究,探讨镉的表遗传致癌机制。方法从各组CdCl2恶性转化不同阶段及接种裸鼠成瘤的16HBE细胞中提取全基因组DNA,采取甲基化特异性PCR法(Methylation-specific PCR,MSP)检测该基因组P16基因启动子区的甲基化状况,与非转化的16HBE对照细胞进行比较,并用去甲基化因子5-Azac(5-Aza-2’deoxycytidine)处理有异常甲基化的细胞。结果CdCl2恶性转化及接种裸鼠成瘤的16HBE细胞P16抑癌基因启动子区CpG岛存在异常高甲基化现象,且随着细胞恶性程度的增加,甲基化现象有明显升高的趋势,同时发现有异常高甲基化的细胞经去甲基化处理后甲基化现象消失。结论P16基因启动子区的高甲基化导致抑癌基因表达关闭,无法正常发挥细胞增殖周期对细胞分裂和生长的负调控,造成细胞周期失控而导致无限制地细胞增殖,这可能是氯化镉诱导16HBE细胞恶性转化及接种裸鼠成瘤的一种表遗传致癌作用。研究结果可部分解释镉化合物的细胞转化作用及其可能的表遗传致癌机制,以及进一步对甲基化的表型逆转和药物治疗研究提供了重要依据。

食管鳞状细胞癌p16基因变异及其与肿瘤病理的关系

目的 探讨 p16基因与原发性食管鳞状细胞癌发生、发展的关系。 方法 应用聚合酶链反应 -单链构象多态分析方法检测 5 3例食管鳞状细胞癌组织中 p16基因的缺失和突变 ,分析 p16基因变异与食管鳞状细胞癌的临床病理关系。 结果 5 3例食管癌患者中 ,14例为 p16基因纯合性缺失 ,12例存在突变。p16基因变异在淋巴结转移、远处转移及临床分期方面差异有显著性 (P<0 .0 5 ) ;在性别、年龄、肿瘤长度及肿瘤浸润深度方面差异无显著性 (P>0 .0 5 )。 结论 p16基因的缺失与突变可能是食管鳞状细胞癌的晚期事件 ,它在食管癌的发生、发展中起重要作用。p16基因缺失与突变的检测可作为判断食管鳞状细胞癌恶性程度。

鼻咽癌P^(16)基因的异常表达及其临床意义的探讨

目的 :研究鼻咽癌抑癌基因P16存在的状态及其与鼻咽癌发生、发展的关系。方法 :采用免疫组织化学S P法对 6 0例鼻咽癌、38例鼻咽癌旁组织和 2 0例正常或炎性鼻咽组织P16基因的表达进行检测。结果 :鼻咽癌、癌旁、正常或炎性组织中P16基因总阳性率为 2 3 33%、73 6 8%和 80 0 0 % ,前者与后两者阳性之间比较均有显著性差异 (P <0 0 5 ,r=- 0 5 114 ) ,而后两者阳性率比较无显著性差异 (P >0 0 5 )。在 6 0例鼻咽癌中 ,有淋巴结转移 4 4例 ,P16基因阳性率 18 88% ,无淋巴结转移 16例 ,P16基因阳性率 5 0 0 0 % ,两者阳性率比较有显著性差异 (P <0 0 5 ,r =- 0 376 4 )。结论 :P16基因低表达或缺失与鼻咽癌的发生。

结肠癌组织中p16和CDK4的表达及相互关系

采用免疫组化法分析６０例结肠癌、结肠息肉和结肠炎标本中ｐ１６和ＣＤＫ４的蛋白表达。结肠癌和结肠息肉组织中ｐ１６表达随细胞分化程度降低而减少；结肠炎组织ｐ１６表达高于癌和息肉组，ＣＤＫ４在上述三种组织中表达无显著差异；推测结肠组织从癌前病变至癌变发展中有ｐ１６对ＣＤＫ４抑制失调。该检测方法亦为临床上区分癌与癌前病变及变化趋势提供辅助诊断方法。

微RNA在皮肤鳞状细胞癌中的作用研究进展

皮肤鳞状细胞癌(CSCC)是临床常见的皮肤癌症之一,其发生、发展可能与微RNA(miRNA)的异常表达有关。miRNA通过与靶基因信使RNA进行完全或不完全互补配对,实现转录后水平的调控。miRNA的表达在CSCC的增殖、迁移和侵袭过程中发挥重要作用,其中miR-31、miR-205、miR-221、miR-365和miR-506可促进肿瘤细胞增殖、迁移,促进CSCC发展,而miR-15b、miR-20a、miR-124/214、miR-125b、miR-148a、miR-199a-5p、miR-203、miR-361-5p、miR-497以及miR-3619-5p可抑制肿瘤细胞发展增殖、迁移,抑制CSCC发展。未来深入探索与CSCC相关miRNA的表达,寻找其靶基因,研究其作用机制可能为CSCC的治疗提供新途径。

诱导子宫内膜癌细胞DNA错配修复及肿瘤增殖抑制基因启动子CpG岛去甲基化修饰的研究

目的:研究在用5-杂氮-2′脱氧胞苷(5-Aza-CdR)诱导体外培养的人子宫内膜癌细胞株HEC-1-A中,DNA错配修复基因hMLH1、hMSH2及细胞周期调控基因p16启动子CpG岛的去甲基化,及对于该细胞的增殖抑制作用。方法:体外培养人子宫内膜癌细胞株HEC-1-A;利用MTT比色法检测5-Aza-CdR对于HEC-1-A的增殖抑制的影响;利用流式细胞术(FCM)分析药物处理前后HEC-1-A的细胞周期;利用重亚硫酸盐处理及甲基化特异性PCR(MS-PCR)检测hMLH1、hMSH2及p16基因启动子CpG岛的甲基化修饰;提取药物处理前后HEC-1-A细胞的总蛋白,利用Westernblotting检测hMLH1、hMSH2及p16蛋白的表达情况。结果:5-Aza-CdR对于人子宫内膜癌细胞株HEC-1-A体外增殖抑制呈明显的剂量依赖性[IC50为(0.71±0.05)μmol.L-1];FCM分析显示HEC-1-A细胞在5-Aza-CdR处理前后均呈现明显的细胞周期差异,并且药物组细胞表现为G0/G1期(P<0.05)阻滞。MS-PCR分析5-Aza-CdR显示著诱导hMLH1、hMSH2基因启动子CpG岛的去甲基化修饰。Western blotting检测发现IC50剂量的5-Aza-CdR处理HEC-1-A细胞前后,hMLH1、hMSH2及p16的表达量为(111.30±3.17)%、(76.99±1.19)%和(97.63±1.28)%明显高于空白对照组(P<0.05),提示该药物有提高上述蛋白表达的作用。结论:5-Aza-CdR可诱导人子宫内膜癌细胞株HEC-1-A中,hMLH1、hMSH2及p16启动子CpG岛的去甲基化修饰,从而有效抑制该细胞的体外增殖。