目的 探讨多聚胞嘧啶结合蛋白(PCBP)3过表达对结直肠癌(CRC)细胞增殖和迁移的影响.方法 收集2022年5至6月在嘉兴市第一医院行CRC切除术患者的CRC组织及癌旁正常组织标本各10份,比较CRC组织与癌旁正常组织中PCBP3 mRNA及蛋白表达水平.取人CRC细胞系HCT116、HT29,使用含有PCBP3 CDS区序列的过表达质粒分别转染HCT116和HT29细胞(设为HCT116过表达组、HT29过表达组),未经转染的HCT116和HT29细胞分别设为HCT116对照组、HT29对照组,检测PCBP3过表达对CRC细胞增殖、侵袭能力以及E-钙黏蛋白(E-cadherin)、裂解的胱天蛋白酶3(Cleaved Caspase-3)蛋白表达水平的影响.结果 CRC组织中PCBP3 mRNA及蛋白表达水平均明显低于癌旁正常组织(均P＜0.05).与对照组比较,PCBP3过表达可明显减弱HCT116和HT29细胞增殖和侵袭能力(均P＜0.05),上调HCT116和HT29细胞E-cadherin、Cleaved Caspase-3蛋白表达水平(均P＜0.05).结论 PCBP3过表达能抑制CRC细胞增殖和侵袭,可能通过上调E-cadherin、Cleaved Caspase-3蛋白表达水平发挥作用.

目的 探讨同源盒蛋白(HOX)B5对结直肠癌(CRC)细胞有氧糖酵解和恶性增殖的影响.方法 通过预实验选取CRC细胞株Caco2构建过表达HOXB5的细胞株,HCT116构建敲低HOXB5的细胞株;分别采用CCK-8法、平板克隆实验、糖代谢检测试剂盒、Western blot法检测过表达或敲低HOXB5对CRC细胞增殖能力、平板克隆形成能力、有氧糖酵解、糖酵解相关蛋白的影响.结果 在Caco2细胞中,过表达HOXB5能明显增强细胞增殖及平板克隆形成能力(均P<0.05),增加细胞葡萄糖消耗量以及乳酸、三磷酸腺苷(ATP)含量(均P<0.05),上调丙酮酸激酶同工酶2(PKM2)、乳酸脱氢酶A(LDHA)、葡萄糖转运蛋白1(GLUT1)蛋白表达水平(均P<0.05);在HCT116细胞中,敲低HOXB5能明显减弱细胞增殖及平板克隆形成能力(均P<0.05),减少细胞葡萄糖消耗量以及乳酸、ATP含量(均P<0.05),下调PKM2、LDHA、GLUT1蛋白表达水平(均P<0.05).结论 HOXB5通过促进CRC细胞有氧糖酵解来增强其恶性增殖.

目的 探讨老年女性直肠前突并直肠孤立性溃疡治疗体会.方法 选取2020年1月至2021年1月期间对3例确诊Ⅱ度以上直肠前突伴直肠孤立性溃疡老年女性患者,行吻合器直肠下端黏膜切除术(PPH术式或STARR术式)+直肠闭式修补术(Block术式),配以生物反馈调节及饮食、排便指导.结果 治疗后随访6～12个月无再便血,患者排便不畅感明显缓解,复查肠镜直肠溃疡病灶基本消失,手术后较手术前排粪造影力排相直肠前突程度明显减轻.结论 对于Ⅱ度以上直肠前突伴直肠孤立性溃疡老年女性患者,吻合器直肠黏膜切除术+经直肠闭式修补术是较佳的手术方式选择,配合生活习俗,排便、用药习惯调整,一般能取得理想疗效.

目的 分析直肠癌标本切缘及区域淋巴结组织中SATB1及EMT各标志基因表达与患者预后情况,探讨SATB1介导上调Snail/Slug信号通路促进直肠癌上皮间充质转化影响肿瘤预后的指导价值.方法 回顾性分析80例中低位直肠癌患者的临床资料,通过RT-qPCR及Western blot法检测SATB1及EMT各标志基因(E-cadherin、Snail及Slug)在环周切缘、下切缘、区域淋巴结、肿瘤组织及正常肠黏膜组织中表达,评估肿瘤切缘及区域淋巴结肿瘤微转移情况;应用单因素Kaplan-Meier检验及多因素分析COX风险比例模型对患者预后(包括无瘤生存时间、局部复发时间、远处转移时间三个控制点)进行相关性分析.结果 远处转移或局部复发患者相比无瘤生存患者SATB1、Snail及Slug表达明显增加(P<0.01),E-cadherin表达减少(P<0.05);Kaplan-Meier检验提示E-cadherin表达与无瘤生存时间呈正相关,与局部复发时间呈负相关;SATB1表达与无瘤生存时间呈负相关,与局部复发时间及远处转移时间均呈正相关;Snail及Slug表达与无瘤生存时间呈负相关,与局部复发时间呈正相关;多因素分析COX风险比例模型检验提示SATB1是影响直肠癌患者局部复发的独立危险因素.结论 直肠癌标本切缘及区域淋巴结组织中SATB1及EMT各标志基因(E-cadherin、Snail及Slug)的表达与直肠癌患者预后存在相关性,SATB1表达增加是影响肿瘤局部复发的独立危险因素;SATB1介导上调Snail/Slug信号通路促进上皮间充质转化对判断直肠癌患者预后具有积极的指导价值.

Here we report a molecular mass amplifying strategy to construct a fluorescence anisotropy sensor for the detection of microRNA. Because fluorescence anisotropy is less affected by biological matrix interferences, the proposed method can be used directly for the detection of targets in tumor cell lysates without any tedious sample pretreatment, and a good linearity in the 10 pM to 0.5 nM range with a detection limit down to 3.4 pM was achieved. The results established that our method can be used to design fluorescence anisotropy sensors for rapid, sensitive, and selective detection of miRNAs for early diagnosis of cancer.

目的 探讨锌指蛋白545(ZNF545)对结直肠癌上皮间质转化(EMT)和迁移侵袭的影响.方法 免疫印迹试验检测ZNF545在结直肠癌细胞系中的表达,并构建过表达和敲低ZNF545的细胞株;采用Transwell小室实验检测细胞的迁移和侵袭能力,免疫印迹试验检测EMT相关分子标志物[E钙黏蛋白(E-cadherin)、磷酸化锌指转录因子(slug)]的表达;细胞免疫荧光检测ZNF545表达变化对结直肠癌细胞E-cadherin表达的影响.结果 内源性ZNF545蛋白在SW480细胞中表达最高,在Lovo、Caco2细胞中的表达次之,在HCT116细胞中表达最低.过表达ZNF545可抑制HCT116细胞的迁移和侵袭能力(均P<0.01),抑制slug的表达(P<0.01),增加E-cadherin的表达(P<0.01);而敲低ZNF545的表达后,SW480细胞的迁移和侵袭能力明显增强(均P<0.01),可增加slug的表达(P<0.01),抑制E-cadherin的表达(P<0.01).在荧光显微镜下,过表达ZNF545可增加E-caherin表达;敲低ZNF545表达后抑制E-caherin表达.结论 ZNF545通过参与结直肠癌EMT过程调控结直肠癌细胞迁移和侵袭能力.

目的 探索泛素结合酶UBE2T在结直肠癌的表达及其生物学作用.方法 运用免疫组化法检测78对结直肠癌和对应癌旁正常组织中UBE2T的蛋白表达水平.脂质体转染法将UBE2T-siRNA及对照NC-siRNA转染结直肠癌细胞SW480后,采用细胞计数试剂盒(CCK-8)及平板克隆实验分析增殖情况;流式细胞仪检测细胞周期情况;Western blotting检测细胞中UBE2T、cyclin D1、cyclin E等蛋白表达情况.结果 与正常结直肠组织相比,结直肠癌组织中UBE2T表达水平增高(P＜0.001).癌组织中UBE2T异常高表达与不良病理指标相关:肿瘤大小(P=0.014)、组织分化程度(P=0.00S)、浸润深度(P=0.033)和TNM分期(P =0.014).CCK-8增殖实验和平板克隆形成实验都提示沉默UBE2T表达能显著抑制结直肠癌细胞的增殖.沉默UBE2T抑制周期蛋白cyclin D1和cyclin E的表达,促进结直肠癌细胞发生G1/S期细胞周期阻滞.结论 UBE2T在结直肠癌组织中异常高表达,并与不良病理指标相关.沉默UBE2T通过促进G1/S期细胞周期阻滞抑制结直肠癌细胞的增殖.

目的 探讨结直肠癌HOXA11基因启动子异常甲基化及其与临床病理特征之间的关系.方法 从89对结直肠癌组织和配对的正常肠黏膜组织中分别提取基因组DNA,采用甲基化特异PCR(MSP)检测HOXA11基因启动子区甲基化状态,比较结直肠癌及正常肠黏膜组织中HOXA11甲基化的发生率,并统计分析结直肠癌组织中甲基化水平与患者临床病理特征的关系.在体外实验中,采用甲基化转移酶抑制剂5-Aza-dC处理细胞,实时荧光定量PCR检测HOXA11 mRNA表达水平的变化.结果 结直肠癌组织中HOXA11基因启动子甲基化阳性率显著高于配对的正常肠黏膜组织(P<0.01).HOXA11基因启动子异常甲基化与直肠癌淋巴结转移(P<0.01)及TNM分期(P<0.01)密切相关.结直肠癌细胞经5-Aza-dC处理后HOXA11 mRNA表达水平显著增高(P<0.05).结论 在结直肠癌组织中,HOXA11基因启动子区异常甲基化抑制HOXA11基因的转录表达.HOXA11启动子区异常甲基化与结直肠癌患者的淋巴结转移及TNM分期密切相关.

目的 探讨锌指蛋白545(ZNF545)在结直肠癌组织中的表达及临床意义.方法 采用qRT-PCR法检测32对结直肠癌组织及癌旁正常组织中(来自本院行结直肠癌根治术的32例患者)ZNF545 mRNA相对表达量;进一步采用免疫组化法检测136对结直癌组织及配对癌旁正常组织(来自组织芯片)中ZNF545蛋白表达,并分析这一表达与患者临床特征及生存预后的关系.结果 结直肠癌组织中ZNF545 mRNA相对表达量明显低于配对癌旁正常组织,差异有统计学意义(P＜0.05).结直肠癌组织中ZNF545蛋白低表达(免疫组化评分≤3分)率为65.4％,明显高于配对癌旁正常组织的25.7％(P＜0.05);ZNF545蛋白低表达与患者肿瘤直径、浸润深度、TNM分期相关(均P＜0.05),与患者性别、年龄、肿瘤位置、分化程度、淋巴结转移无关(均P＞0.05).Kaplan-Meier生存分析结果显示,与ZNF545蛋白高表达的结直肠癌患者比较,ZNF545蛋白低表达患者的5年生存率明显降低,差异有统计学意义(P＜0.05).结论 ZNF545在结直肠癌组织中呈低表达,与患者肿瘤直径、浸润深度、TNM分期及生存预后有关.

目的 研究大黄素是否可增强5AzA-cdR对胰腺癌Panc1细胞抑癌基因p16、RASSF1A的去甲基化作用.方法 采用细胞增殖实验检测不同浓度大黄素对Panc1细胞的生长抑制情况,焦磷酸盐测序PCR(BSP)分别检测大黄素、5AzA-cdR及大黄素联合5AzA-cdR对Panc1细胞抑癌基因p16 、RASSF1A甲基化状态的影响,并用荧光定量PCR(FQ-PCR)和Western blot分别检测p16 、RASSF1A及甲基转移酶DNMT1、DNMT3a在mRNA和蛋白水平的表达情况.结果 大黄素以时间和浓度梯度依赖性抑制Panc1细胞生长.BSP结果显示大黄素具有微弱的去甲基化作用,5AzA-cdR具有一定程度的去甲基化作用,当两者联用时,去甲基化作用更加显著;FQ-PCR和Western blot结果显示大黄素与5AzA-cdR联用时,p16、RASSF1A的表达水平均较空白对照明显增高(均P＜0.05),DNMT1、DNMT3a的表达水平均较空白对照明显降低(均P＜0.05).结论 大黄素与5AzA-cdR联用可通过降低DNMT1和DNMT3a的表达水平来增强5AzA-cdR对胰腺癌抑癌基因p16、RASSF1A的去甲基化作用.

5-Aza-2'-deoxycytidine (5-Aza-CdR) is currently acknowledged as a demethylation drug, and causes a certain degree of demethylation in a variety of cancer cells, including pancreatic cancer cells. Emodin, a traditional Chinese medicine (TCM), is an effective monomer extracted from rhubarb and has been reported to exhibit antitumor activity in different manners in pancreatic cancer. In the present study, we examined whether emodin caused demethylation and increased the demethylation of three tumor-suppressor genes P16, RASSF1A and ppENK with a high degree of methylation in pancreatic cancer when combined with 5-Aza-CdR. Our research showed that emodin inhibited the growth of pancreatic cancer Panc-1 cells in a dose- and time-dependent manner. Dot-blot results showed that emodin combined with 5-Aza-CdR significantly suppressed the expression of genome 5mC in PANC-1 cells. In order to verify the effect of methylation, methylation-specific PCR (MSP) and bisulfite genomic sequencing PCR (BSP) combined with TA were selected for the cloning and sequencing. Results of MSP and BSP confirmed that emodin caused faint demethylation, and 5-Aza-CdR had a certain degree of demethylation. When emodin was combined with 5-Aza-CdR, the demethylation was more significant. At the same time, fluorescent quantitative PCR and western blot analysis results confirmed that when emodin was combined with 5-Aza-CdR, the expression levels of P16, RASSF1A and ppENK were increased more significantly compared to either treatment alone. In contrast, the expression levels of DNA methyltransferase 1 (DNMT1) and DNMT3a were more significantly reduced with the combination treatment than the control or either agent alone, further proving that emodin in combination with 5-Aza-CdR enhanced the demethylation effect of 5-Aza-CdR by reducing the expression of methyltransferases. In conclusion, the present study confirmed that emodin in combination with 5-Aza-CdR enhanced the demethylation by 5-Aza-CdR of tumor-suppressor genes p16, RASSF1A and ppENK by reducing the expression of methyltransferases DNMT1 and DNMT3a.

Emodin, a natural anthraquinone derivative isolated from Rheum palmatum, has been reported to inhibit the growth of pancreatic cancer cells through different modes of action; yet, the detailed mechanism remains unclear. In the present study, we hypothesized that emodin exerts its antitumor effect by participating in the regulation of the DNA methylation level. Our research showed that emodin inhibited the growth of pancreatic cancer PANC-1 cells in a dose- and time-dependent manner. Dot-blot results showed that 40 µM emodin significantly inhibited genomic 5 mC expression in the PANC-1 cells, and mRNA-Seq showed that different concentrations of emodin could alter the gene expression profile in the PANC-1 cells. BSP confirmed that the methylation levels of P16, RASSF1A and ppENK were decreased, while concomitantly the unmethylated status was increased. RT-PCR and western blotting results confirmed that the low expression or absence of expression of mRNA and protein in the PANC-1 cells was re-expressed following treatment with emodin. In conclusion, our study for the first time suggests that emodin inhibits pancreatic cancer cell growth, which may be related to the demethylation of tumor-suppressor genes. The related mechanism may be through the inhibition of methyltransferase expression.

左半结肠癌合并急性肠梗阻多见于老年人，老年人多合并其他疾病，对疾病敏感性差，就诊较晚，结肠梗阻已多为闭袢性梗阻。2005年1月至2014年3月本院对38例老年左半结肠癌合并急性肠梗阻患者，术中做结肠灌洗后行一期切除吻合，疗效满意。报告如下。

目的：探讨低位直肠癌前切除术吻合口瘘的病因及有效的防治措施。方法回顾性分析低位直肠癌患者行前切除术后发生吻合口瘘16例患者的临床资料。结果16例吻合口瘘患者均痊愈，无一例死亡，平均愈合时间15.4d（11~20d）。横结肠造瘘患者分别于3个月及5个月后将造瘘口还纳腹腔，治愈出院。结论吻合口瘘是低位直肠癌前切除术的常见并发症，应重视其发生的原因、预防措施及临床诊断，绝大部分可经非手术治疗治愈。