miR-203a通过靶向抑制CDK6调控膀胱癌细胞增殖及放射敏感性

目的 探讨miR-203a在膀胱癌(BC)细胞系(RT-112、T24、5637、UM-UC-3细胞)中的表达及对细胞增殖及放射敏感性的影响。方法 将miR-203a mimics、miR-203a inhibitor、CDK6siRNA、CDK6表达质粒及相应阴性对照(NC)转染入BC细胞中。实时荧光定量PCR检测miR-203a在RT-112、T24、5637、UM-UC-3细胞和人膀胱上皮永生化细胞系(SV-HUC-1)中的表达。CCK8实验研究miR-203a和CDK6对BC细胞系增殖的调控。克隆形成实验研究miR-203a和CDK6对BC细胞系放射敏感性的影响。荧光素酶报告基因实验验证miR-203a的靶基因。蛋白印迹法检测miR-203a对CDK6蛋白表达的影响。采用单因素方差分析进行多组间比较、t检验进行两组间比较。结果 与SV-HUC-1细胞相比,miR-203a在RT-112、T24、5637、UM-UC-3细胞中的表达明显降低(P<0.05)。与NC相比,过表达miR-203a抑制BC细胞系增殖(P<0.05),敲低miR-203a促进BC细胞系增殖(P<0.05)。与NC相比,过表达miR-203a增加BC细胞系的放射敏感性(P<0.05),敲低miR-203a减弱BC细胞系的放射敏感性(P<0.05)。CDK6是miR-203a的作用靶点。与NC组相比,过表达miR-203a显著降低了CDK6蛋白水平(P<0.05),敲低miR-203a显著上调了CDK6蛋白水平(P<0.05)。在转染miR-203a mimics的T24和UM-UC-3细胞中过表达CDK6后,与miR-203a mimics组相比,细胞增殖能力升高、放射敏感性降低(P<0.05);在转染miR-203a inhibitor的RT-112和5637细胞中沉默CDK6后,与miR-203a inhibitor组相比,细胞增殖能力降低、放射敏感性升高(P<0.05)。结论 miR-203a在BC细胞系中低表达,可作为抑癌基因抑制BC细胞系增殖和增强放射敏感性。     

lncRNA FOXD2-AS1通过靶向miR-506-5p调控宫颈癌细胞增殖与凋亡

[摘要] 目的: 探讨长链非编码RNA（lncRNA）FOXD2-AS1 是否通过靶向miR-506-5p 调控宫颈癌细胞增殖和凋亡。方法:体外培养人宫颈癌细胞系HeLa、Siha、Caski 与正常宫颈细胞株Ect1/E6E7,用qPCR检测细胞中FOXD2-AS1 和miR-506-5p 的表达水平。用脂质体转染技术分别构建抑制FOXD2-AS1 表达和过表达miR-506-5p 的宫颈癌细胞,用MTT实验和流式细胞术检测细胞的增殖和凋亡情况,WB实验检测细胞中增殖相关蛋白CyclinD1、p21 和p27 及凋亡相关蛋白Bcl-2、BAX和cleaved-capase-3的表达。用双荧光素酶报告基因实验验证FOXD2-AS1 是否靶向miR-506-5p,并分析同时抑制FOXD2-AS1 和miR-506-5p 表达对宫颈癌细胞增殖与凋亡的影响。结果：与Ect1/E6E7 细胞比较,宫颈癌HeLa、Siha 和Caski 细胞中FOXD2-AS1 表达水平显著升高,miR-506-5p 表达水平降低（均P<0.01）。抑制FOXD2-AS1 表达可显著抑制宫颈癌细胞中CyclinD1 蛋白和Bcl-2 蛋白表达,并促进p21、p27 和BAX、cleaved-capase-3 蛋白的表达,抑制细胞增殖并促进细胞凋亡（均P<0.01）。过表达miR-506-5p 可显著抑制宫颈癌细胞中CyclinD1 和Bcl-2 蛋白表达,促进p21 和BAX蛋白表达,抑制细胞增殖并促进细胞凋亡（均P<0.01）。双荧光素酶报告基因实验证实宫颈癌细胞中FOXD2-AS1 靶向负调控miR-506-5p 的表达（P<0.01）。抑制miR-506-5p 表达逆转了抑制FOXD2-AS1 表达对宫颈癌细胞增殖和凋亡的作用（P<0.01）。结论: FOXD2-AS1 通过靶向miR-506-5p 的表达调控宫颈癌细胞的增殖和凋亡。     

miR-26a/b调控p53/MDM2通路对胃癌细胞凋亡的影响

目的 探讨miR-26a/b调控p53/MDM2通路对胃癌细胞凋亡的影响。方法 实时荧光定量PCR检测胃癌细胞系MGC803、MKN-45和MKN-28中miR-26a/b的表达水平,荧光素酶报告系统分析miR-26a/b与MDM2 3’非翻译区（3’UTR）的结合情况;将化学合成miR-26a/b前体分子mimics和无关序列转染胃癌细胞MKN-45,化学合成miR-26a/b inhibitor转染胃上皮细胞GES-1后,Western blotting检测MDM2、p53及其下游分子p21和Bcl-2的表达,MTT法检测转染24、48、72 和96 h的细胞增殖情况;通过Annexin Ⅴ/PI双染检测miR-26a/b对MKN-45细胞凋亡情况。结果 与永生化胃上皮细胞GES-1相比,miR-26a/b在肿瘤细胞系MGC803、MKN-45和MKN-28中的表达均下调,在MKN-45中表达水平最低;荧光素酶活性检测显示,miR-26a/b过表达抑制MDM2 3’UTR报告载体（Wild）的荧光素酶活性,而对3’UTR突变型报告载体（Mutation）荧光素酶活性无明显影响。miR-26a/b抑制MKN-45细胞中MDM2表达并增强p53及下游分子表达,而在GES-1细胞中抑制miR-26a/b可以增强MDM2表达,降低p53及下游分子表达。MTT结果显示,miR-26a/b抑制MKN-45细胞的增殖,miR-26a/b inhibitor则明显促进GES-1细胞的增殖。通过AnnexinⅤ/PI检测细胞凋亡发现,miR-26a/b过表达的MKN-45细胞的凋亡率高于对照细胞（P＜0.01）。结论 miR-26a/b 能与MDM2 3’UTR 特异结合,调控p53/MDM2通路影响胃癌细胞的增殖及凋亡。     

rmhTNF-α对胃癌细胞系MKN45的生物学效应及其机制研究

摘 要：［目的］ 研究rmhTNF对胃癌细胞系MKN45的生物学效应及其机制。［方法］ 采用不同浓度（50、100、200IU/ml）重组改构人肿瘤坏死因子（rmhTNF）处理胃癌细胞MKN45,增殖/毒性检测试剂盒(CCK-8)观察其细胞增殖抑制率;实时荧光定量PCR（RT-PCR）法检测MKN45细胞p53异构体Δ133p53、STAT1及 STAT3 mRNA的表达变化。［结果］ CCK-8结果显示,随 rmhTNF 作用浓度增高（50、100、200IU/ml）,MKN45 细胞抑制率分别为17.133%,24.800%和31.733%,差异有统计学意义（F=16.246,P<0.01）。RT-PCR结果显示,随 rmhTNF 浓度增高,MKN45 细胞STAT1 mRNA表达上升（F=164.290,P<0.001）,STAT3、Δ133p53 mRNA表达下降（F=29.921,F=24.243;P均<0.001）。Pearson 相关性分析结果显示,Δ133p53 与STAT1表达呈负相关（r=-0.951,P<0.01）,与STAT3表达呈正相关（r=0.840,P<0.01）。［结论］MKN45细胞中,Δ133p53是STAT1、STAT3调控肿瘤细胞的共同靶基因,STAT1、STAT3-Δ133p53-p53通路可能是rmhTNF抑制胃癌细胞MKN45的机制。     

香菇多糖联合多西他赛对胃癌BGC823细胞增殖的影响

目的：观察香菇多糖单药及联合多西他赛抑制胃癌细胞BGC823增殖的作用.方法：分别用不同浓度的香菇多糖、多西他赛和香菇多糖联合多西他赛处理胃癌细胞BGC823.香菇多糖终浓度分别为1.560μg/ml,3.125 μg/ml,6.250 μg/ml,12.500μg/ml.多西他赛终浓度分别为0.625 μg/ml,1.250μg/ml,2.500μg/nl,5.000 μg/ml.MTT法检测各组细胞增殖情况;同时用Annexin Ⅴ/PI法检测各组细胞的凋亡.结果：单药香菇多糖能抑制BCG823细胞的增殖,不同浓度（由高到低）对BGC823细胞的增殖抑制率分别为67.7％,56.0％、42.1％和23.2％,与阴性对照组比较有统计学意义（P＜0.05）.不同浓度香菇多糖联合多西他赛能增强多西他赛对BGC823细胞增殖的抑制作用.单药香菇多糖能增加BGC823细胞的细胞凋亡率,与多西他赛联用使细胞凋亡率明显增加（P＜0.05）.结论：香菇多糖能抑制胃癌BGC823细胞的增殖,并能显著增强多西他赛抑制胃癌细胞BGC823的增殖作用.     

天花粉蛋白诱导胃癌细胞MKN-45凋亡的研究

目的 :研究天花粉蛋白（ trichosanthin,TCS）对胃癌细胞的作用,探讨其治疗胃癌的可能作用机制。方法 :采用生长曲线、集落形成实验、 MTT法、电镜、 TUNEL染色和流式细胞仪等技术,研究 TCS对胃癌细胞 MKN－ 45的增殖抑制和诱导凋亡作用。结果： TCS能够明显抑制胃癌细胞 MKN－ 45的生长和集落形成, 1μ g/ml和 0.1μ g/ml TCS作用 MKN－ 45后,细胞集落形成率分别为 (6.63± 1.31)％和 (14.68± 1.27)％,明显低于对照组的 (23.67± 2.76)％ (P     

丹参酮IIA 通过调控上皮间质转化抑制食管癌EC9706 和KYSE70 细胞的凋亡、侵袭和迁移

目的:探讨丹参酮IIA 对食管癌EC9706 和KYSE70 细胞侵袭和迁移的影响及其调控机制。方法:食管癌细胞系EC9706 和KYSE70 培养完成后分为4 组：对照组（加DMSO）和2、4、6 μg/ml 丹参酮组。采用CCK-8 法检测EC9706 和KYSE70 细胞增殖活力,流式细胞术检测细胞凋亡率,Transwell 实验检测细胞侵袭能力,划痕愈合实验检测细胞迁移能力,qRT-PCR和Western blotting 实验检测EMT相关蛋白E-cadherin、Snail-2、Vimentin 和N-cadherin mRNA和蛋白表达水平。结果：小于6 μg/ml的丹参酮IIA 不影响食管癌EC9706 和KYSE70 细胞增殖活力;4、6 μg/ml 丹参酮IIA 组细胞凋亡率明显高于对照组（均P<0.01）;2、4、6 μg/ml 丹参酮组每个视野下的侵袭细胞数及划痕愈合率明显低于对照组（均P<0.01）,且EC9706 和KYSE70 细胞形态由纺锤状的间充质形态转变为上皮形态。与对照组相比,2、4、6 μg/ml 丹参酮组E-cadherin 表达明显升高,Snail-2、Vimentin 和N-cadherin表达明显下降（均P<0.01）。结论：丹参酮IIA 通过抑制EMT促进食管癌EC9706 和KYSE70 细胞凋亡,并减弱其侵袭和迁移能力。     

miR-29c通过TNRC18 调控胃癌组织和细胞阿帕替尼耐药的机制

目的:探讨miR-29c 调控TNRC18 胃癌组织和细胞阿帕替尼耐药性的机制。方法：收集2015 年2 月至2017 年10 月武汉市中心医院具有完整资料的39 例胃癌和癌旁组织标本（其中21 例为阿帕替尼耐药患者、18 例为不耐药患者）,采用qRT-PCR检测miR-29c 在胃癌组织和细胞系中的表达水平。采用CCK-8、Transwell 和Annexin V-FITC/PI 双染流式术检测miR-29c 过表达/敲降对MGC-803/AP耐药细胞增殖、侵袭和凋亡影响,Western blotting 检测miR-29c 调控TNRC18 表达,双荧光素酶报告基因验证miR-29c 与TNRC18 的靶向作用关系。结果：miR-29c 在3 种胃癌细胞系和阿帕替尼耐药癌患者组织中均低表达。双荧光素酶报告基因证实miR-29c 靶向作用TNRC18 并下调其表达水平。miR-29c 通过靶向下调TNRC18 抑制阿帕替药耐药的MGC-803/AP细胞的增殖、侵袭并促进细胞凋亡（均P<0.05 或P<0.01),进而降低胃癌细胞MGC-803/AP对阿帕替尼的耐药性。体内实验同样证实,miR-29c 通过靶向抑制TNRC18 降低胃癌对阿帕替尼的耐药性。结论：miR-29c/TNRC18 分子轴在胃癌组织和细胞MGC-803/AP对阿帕替尼耐药中发挥着一定作用,过表达miR-29c可逆转MGC-803/AP细胞对阿帕替尼耐药。     

rmhTNF联合顺铂抑制人胃癌细胞增殖的作用机制

摘 要：［目的］探讨rmhTNF联合顺铂抑制人胃癌细胞增殖的作用机制。［方法］不同浓度rmhTNF（50,100,200IU/ml）单独或联合顺铂（4μg/ml）作用于人胃癌细胞系MKN45和SGC7901,细胞增殖/毒性检测试剂盒（CCK-8试剂盒）测定细胞抑制率;巢式逆转录多聚酶链反应（RT-PCR）测定细胞中p53β和caspase-3 mRNA的表达变化。［结果］ CCK-8法结果显示,MKN45胃癌细胞中联合组细胞抑制率高于单独组,且随rmhTNF浓度的增加而增加,差异有统计学意义;而SGC7901胃癌细胞无此现象且单独rmhTNF对SGC7901胃癌细胞增殖抑制不明显,差异无统计学意义（F=1.01,P>0.05） 。单独rmhTNF组中MKN45胃癌细胞抑制率随药物浓度增加而增加,差异有统计学意义（F=35.40,P<0.001）。RT-PCR结果显示,在MKN45胃癌细胞系中单独组和联合组caspase-3 mRNA表达均增加,且联合组均高于单独组并随rmhTNF浓度的增加而增加,组间差异有统计学意义（F=93.889,P<0.05）;p53β在单独rmhTNF组中未见明显改变,差异无统计学意义（F=0.006,P>0.05）,rmhTNF联合顺铂作用时可明显上调p53β的表达,并且随rmhTNF浓度的增加而增加,差异有统计学意义（F=18.577,P<0.001 ）。在SGC7901胃癌细胞中未见p53β表达;而caspase-3 mRNA的表达趋势与MKN45相同,组间差异有统计学意义（F=1409.656,P<0.05）。Person相关性分析显示,p53β与caspase-3表达呈正相关（r=0.766,P<0.001）,细胞抑制率与caspase-3的表达呈正相关（r=0.978,P<0.001）。［结论］ rmhTNF和顺铂对胃癌细胞系MKN45的协同抑制作用机制可能是通过p53β上调caspase-3。     

miR-34a通过靶基因Notch1调控子宫内膜癌细胞的增殖和凋亡

目的:探讨miR-34a通过靶基因Notch1调控子宫内膜癌细胞的增殖和凋亡。方法:通过荧光定量PCR检测miR-34a在子宫内膜癌和癌旁组织中的表达,并通过双荧光素酶报告基因检测miR-34a的靶基因,子宫内膜癌Ishikawa细胞转染miR-34a mimics及对照miR-NC,并转染Notch表达载体,通过MTT和流式细胞术分别检测miR-34a和Notch1对子宫内膜癌Ishikawa细胞增殖和凋亡的影响,并通过Western blot检测miR-34a对Notch1蛋白表达的影响。结果:miR-34a在人子宫内膜癌组织中表达显著下调（P<0.05）,双荧光素酶报告基因实验证实miR-34a能与Notch1 3' UTR结合,miR-34a mimics能够显著抑制细胞活力（P<0.05）,促进细胞凋亡（P<0.05）,并且miR-34a mimics显著抑制Notch1蛋白表达（P<0.05）。结论:miR-34a能够通过调控靶基因Notch1的表达,抑制子宫内膜癌细胞的增殖,促进细胞凋亡。     

白介素6在雌激素促进小鼠肺腺癌进展中的作用及其机制

目的 探讨白介素6（interleukin 6, IL-6）在雌激素（estrogen, E2）促进小鼠肺腺癌进展中的作用及其可能的机制。方法 采用乌拉坦诱导雌性昆明小鼠建立肺腺癌模型,进行如下分组处理（每组9只）：空白对照组、E2组、雌激素抑制剂（E2 inhibitor, E2I）组、E2+E2I组、IL-6组、E2+IL-6组。小鼠饲养16周后处死检测小鼠体重变化、成瘤情况、肿瘤分级、肺脏器指数等指标,酶联免疫法（ELISA）检测对照组血清标本中E2/IL-6的含量,实时荧光定量PCR（Real-Time PCR, RT-PCR）检测对照组肿瘤标本中ERβ/IL-6的mRNA表达水平,采用免疫印迹法检测各组ERβ、Akt、MAPK,p-ERβ、p-Akt、p-MAPK表达水平。结果 肺部出现结节的昆明小鼠占小鼠总数的比例（即称为成瘤率）为87.04%（47/54）,小鼠肺部结节做病理切片后证实为肺腺癌的小鼠数目占小鼠总数的比例（即成癌率）为70.37%（38/54）,肺结节数、肿瘤指数、肺脏器指数等统计指标在E2组显著高于E2+E2I组、E2I组、E2+IL-6组、IL-6组及空白对照组（P均<0.05）;对照组小鼠血清中E2与IL-6呈高度相关性（P均<0.05）,各组小鼠肺癌组织中Akt、MAPK、ERβ、p-AKt、p-MAPK、p-ERβ的表达水平及ERβ的mRNA表达水平与肿瘤统计指标表达的趋势相一致（P均<0.05）。结论 在E2促进小鼠肺腺癌进展过程中IL-6具有下调ERβ信号通路作用,提示IL-6可能抑制E2促进肺腺癌进展。     

Interleukin-6 released by colon cancer-associated fibroblasts is critical for tumour angiogenesis: anti-interleukin-6 receptor antibody suppressed angiogenesis and inhibited tumour–stroma interaction

Background:

Interleukin-6 (IL-6) has an important role in cancer progression, and high levels of plasma IL-6 are correlated with a poor prognosis in a variety of cancers. It has also been reported that tumour stromal fibroblasts are necessary for steps in cancer progression, such as angiogenesis. There have been few reports of a correlation between fibroblast actions and IL-6 levels. In this study, we examined the correlation between cancer stromal fibroblasts and IL-6 and the utility of IL-6 as a therapeutic target in human colon cancer.

Methods:

The expression levels of IL-6 and VEGF of fibroblasts and cancer cell lines were evaluated using real-time PCR and ELISA. The anti-angiogenic effect of inhibiting IL-6 signalling was measured in an angiogenesis model and animal experiment.

Results:

We demonstrate that stromal fibroblasts isolated from colon cancer produced significant amounts of IL-6 and that colon cancer cells enhanced IL-6 production by stromal fibroblasts. Moreover, IL-6 enhanced VEGF production by fibroblasts, thereby inducing angiogenesis. In vivo, anti-IL6 receptor antibody targeting stromal tissue showed greater anti-tumour activity than did anti-IL6 receptor antibody targeting xenografted cancer cells.

Conclusion:

Cancer stromal fibroblasts were an important source of IL-6 in colon cancer. IL-6 produced by activated fibroblasts induced tumour angiogenesis by stimulating adjacent stromal fibroblasts. The relationship between IL-6 and stromal fibroblasts offers new approaches to cancer therapy.     

ANXA4和ANXA6在肝门部胆管癌中的表达及临床意义

目的 探讨膜联蛋白A4（ANXA4）和膜联蛋白A6（ANXA6）在肝门部胆管癌（HC）组织中的表达及两者与HC临床病理特征和预后的关系。方法 采用组织芯片与免疫组织化学技术检测2005年至2007年49例HC组织及10例癌旁组织中ANXA4和ANXA6的表达情况,分析两者表达与HC临床病理特征及预后的关系。结果 ANXA4和ANXA6在HC组织中的阳性表达率分别为63.3％（31／49）和69.4％（34／49）,明显高于癌旁组织的20.0％（2／10）和30.0％（3／10）,差异均有统计学意义（P＜0.05）。ANXA4蛋白表达与淋巴结转移有关（P＜0.05）,ANXA6表达则与肿瘤大小、淋巴结转移及TNM分期有关（P＜005）。ANXA4阳性表达者的中位总生存时间（OS）和无进展生存时间（PFS）分别为14个月和12个月,明显短于阴性表达者的46个月和39个月（P＜0.05）;ANXA6阳性表达者的中位OS和PFS分别为14个月和12个月,明显短于阴性表达者的42个月和39个月（P＜0.05）。Cox多因素分析显示,ANXA6、浸润深度是影响OS的独立因素,浸润深度是影响PFS的独立因素。结论 ANXA4和ANXA6与HC的发生、发展有关,且ANXA6是潜在的评估预后的肿瘤标志物。     

The Protective Effect of Antioxidants in Areca Nut Extract-Induced Oral Carcinogenesis

Objective: Oral submucous fibrosis (OSF) is the premalignant disorder associated with fibrosis and epithelial atrophy. Areca Nut (AN) is the most significant risk factors for OSF. However, the molecular mechanism behind AN induced OSF remains unclear, and there exists no effective treatment for the malignant disorder. We aimed to investigate whether AN-extract causes epithelial-mesenchymal transition (EMT) in oral keratinocytes, and evaluated the therapeutic potential of antioxidants. Methods: The HPV16 E6/E7-transfected immortalized human oral keratinocytes (IHOK) were employed in the present study. For the preparation of AN-extract, dried AN was dissolved in distilled water overnight. The solution was centrifuged and the supernatant was collected for further use. For the determination of change in cytokine levels, ELISA was performed. To investigate EMT-related protein expression and phenotype, immunoblot and immunofluorescence were performed. Results: Among tumor-promoting cytokines (Gro-α, IL-6 and IL-8), IL-6 was remarkably increased by AN in IHOK. AN-extract induced EMT phenotypes, such as cell elongation, up-regulation of vimentin and snail. After treatment with neutralizing antibody of IL-6, AN-induced snail expression was reduced remarkably. Collectively, AN-extract induced IL-6 expression and mediated EMT. The use of antioxidants (EGCG, glutathione and NAC) significantly reduced IL-6 expression in AN-treated IHOK. Also, AN-decreased E-cadherin and increased vimentin were reversed by antioxidants, indicating that the effectiveness of antioxidants in inhibiting IL-6-induced EMT by AN. Conclusion: AN promotes EMT and antioxidants interrupt AN-induced-EMT in oral keratinocytes. Consequently, it is proposed that antioxidants could prevent AN-induced carcinogenesis and function as a prototype for developing therapeutic interventions of OSF.     

N6-methyladenosine (m6A) regulatory gene divides hepatocellular carcinoma into three subtypes

BackgroundThe N6-methyladenosine (m6A) plays an important role in epigenetic modification and tumor progression, but the modulations of m6A in hepatocellular carcinoma (HCC) have not been determined while the relationship between m6A regulation and immune cell infiltration remains unclear.MethodsThis study investigated the modification patterns of m6A by analyzing HCC samples from The Cancer Genome Atlas (TCGA) database and the Gene Expression Omnibus (GEO) dataset, and performed molecular typing based on the characteristics of immune cell infiltration. The m6Ascore was also constructed to quantify m6A modifications and predict the immunotherapy response and prognosis of HCC patients.ResultsOf the 364 samples, 31 (8.52%) were genetically altered in the m6A regulatory gene, with the highest frequency of mutations in HNRNPC, ZC3H13, and LRPPRC. Three distinct molecular subtypes of m6A were identified in 590 HCC samples, which were associated with different immune cell infiltrates: immunodepletion type, immune activation type, and immune immunity type. According to the construction of the m6Ascore system in the m6A genotype, HCC patients could be divided into high and low groups. The m6A modified pattern, characterized by immune immunity and immune failure, showed a lower score and a better prognosis. However, the immune-activated type of m6A had a higher score and a poorer prognosis. Further analysis showed that the m6Ascore was correlated with tumor mutation burden (TMB), and the higher the TMB, the worse the prognosis. m6Ascore was also correlated with the expression of cytotoxic T-lymphocyte-associated protein 4 (CTAL-4), and the higher the score, the higher the expression of HCC in patients.ConclusionsHCC has a unique m6A modification pattern, and 3 different m6A subtypes help to classify HCC, provide knowledge of drug regimens for immunotherapy, and can be used to predict treatment response and prognosis.     

CHARACTERIZATION OF A HUMAN HERPES VIRUS－6（HHV－6） AND EPSTEIN－BARR VIRUS（EBV） ASSOCIATED LEUKEMIC CELL LINE，J6－1

ＣＨＡＲＡＣＴＥＲＩＺＡＴＩＯＮＯＦＡＨＵＭＡＮＨＥＲＰＥＳＶＩＲＵＳ－６（ＨＨＶ－６）ＡＮＤＥＰＳＴＥＩＮ－ＢＡＲＲＶＩＲＵＳ（ＥＢＶ）ＡＳＳＯＣＩＡＴＥＤＬＥＵＫＥＭＩＣＣＥＬＬＬＩＮＥ，Ｊ６－１ＷｕＫｅｆｕ吴克复；ＪａｎｏｓＬｕｋａ；Ｓｈａｎｔ...     

Serum ICAM-1 in renal cell cancer patients: Possible significance for presence of metastatic disease

Background Intercellular adhesion molecule-1 (ICAM-1) facilitates cell-to-cell adhesion through lymphocyte function associated antigen-1 (LFA-1). ICAM-1 also exists as a soluble form (sICAM-1) and the level of sICAM-1 is known to increase in the presence of malignant tumors. Previously, we reported frequent ICAM-1 expression in renal cell cancer (RCC). sICAM-1 may suppress antitumor immune reactions by blocking LFA-1 on lymphocytes. Methods Serum sICAM-1 and IL-6, possible RCC autocrine growth factors, were examined in 35 RCC patients before and after surgery. In situ expression of sICAM-1 in RCC tissue and the degree of macrophage and lymphocyte infiltration were evaluated semiquantitatively using immunohistochemistry techniques. Results Nine pre- and 12 postoperative patients had elevated sICAM-1 levels. RCC patients with elevated sICAM-1 levels revealed a high ICAM-1 expression and/or high degree of lymphocyte/macrophage infiltration. Elevation or persistence of high sICAM-1 levels after nephrectomy correlated with the presence of metastatic disease (P=0.02); sICAM-1 was elevated in 5 of the 7 (71%) patients with metastasis compared with 7 of the 28 (25%) without evidence of metastatic disease. In some patients, however, the sICAM-1 level fluctuated without regard to the presence or absence of tumor after operation. Elevated sICAM-1 levels were also associated with high IL-6 levels. Conclusion Elevated sICAM-1 levels in RCC patients are not necessarily produced by RCC cells. However, in some cases, elevated sICAM-1 levels may be useful in detecting metastatic disease and aid in the development of treatment strategies and/or postnephrectomy follow-up therapies.     

Inhibition of 1, 2-Dimethylhydrazine-Induced Mucin-Depleted Foci and O6-Methylguanine DNA Adducts in the Rat Colorectum by Boiled Garlic Powder

The scavenging capacity of reactive oxygen species, such as hydroxyl radicals, is reported not to decreasein boiled garlic (an odorless garlic preparation). We therefore examined the modifying effect of boiled garlicpowder (BGP) on 1,2-dimethylhydrazine-induced mucin-depleted foci (MDF) and aberrant crypt foci (ACF),preneoplastic lesions, in the rat colorectum. Male F344 rats (5 weeks old) were fed a basal diet, or experimentaldiets containing 5% or 1% BGP for 5 weeks. One week later, all rats were injected s.c. with DMH (40 mg/kg, onceweekly for 2 weeks). At 10 weeks of age, all the rats were sacrificed, and the colorectum was evaluated for MDFand ACF. In rats given DMH and the 5% or 1% BGP diets (Groups 2 and 3), the numbers of MDF decreasedsignificantly in a dose-dependent manner, compared with the DMH and basal diet value (Group 1) (p<0.01). Thenumbers of ACF in Group 2, but not Group 3, showed a non-significant tendency to decrease. Next, the effectsof BGP on the formation of DMH-induced O6-methylguanine (O6-MeG) DNA adducts in rats were studied. MaleF344 rats (5 weeks old) were fed the basal diet, or 10% BGP diet for 5 weeks. All rats were injected i.p. once with40 mg/kg DMH at the end of week 5. The animals were sacrificed 6 hours after DMH injection to analyze theO6-MeG DNA adducts in the colorectal mucosa. Dietary administration of BGP significantly inhibited the O6-MeG DNA adduct levels in the colorectal mucosa, compared with the controls (p<0.01). These results suggestedthat BGP may exert chemopreventive effects against colon carcinogenesis at least in the initiation stage.     

Tumor suppressor dual-specificity phosphatase 6 (DUSP6) impairs cell invasion and epithelial-mesenchymal transition (EMT)-associated phenotype

Suppressive effects of DUSP6 in tumorigenesis and EMT-associated properties were observed. Dual-specificity phosphatase (DUSP6) is a MAP kinase phosphatase (MKP) negatively regulating the activity of ERK, one of the major molecular switches in the MAPK signaling cascade propagating the signaling responses during malignancies. The impact of DUSP6 in EMT and its contribution to tumor dissemination has not yet been characterized. Due to differences in tumor microenvironments affecting cell signaling during cancer progression, DUSP6 may play varying roles in tumor development. We sought to examine the potential role of DUSP6-mediated tumorigenesis and EMT-associated properties in two aerodigestive tract cancers, namely, esophageal squamous cell carcinoma (ESCC) and nasopharyngeal carcinoma (NPC). Significant loss of DUSP6 was observed in 100% of 11 ESCC cell lines and 71% of seven NPC cell lines. DUSP6 expression was down-regulated in 40% of 30 ESCC tumor tissues and 75% of 20 NPC tumor tissues compared to their respective normal counterparts. Suppressive effects of DUSP6 in tumor formation and cancer cell mobility are seen in in vivo tumorigenicity assay and in vitro colony formation, three-dimensional Matrigel culture, cell migration and invasion chamber tests. Notably, overexpression of DUSP6 impairs EMT-associated properties. Furthermore, tissue microarray analysis reveals a clinical association of DUSP6 expression with better patient survival. Taken together, our study provides a novel insight into understanding the functional impact of DUSP6 in tumorigenesis and metastasis of ESCC and NPC.     

miR-186 Represses Proliferation,Migration, Invasion,and EMT of Hepatocellular Carcinoma via Directly Targeting CDK6

The present study aimed to investigate the effect of miR-186 on proliferation, migration, invasion, and epithelial–mesenchymal transition (EMT) of hepatocellular carcinoma (HCC). In this work, miR-186 was downregulated in HCC tissues and cells, and low miR-186 level helped predict the occurrence of vascular invasion and poor prognosis in patients with HCC. miR-186 overexpression inhibited cell proliferation and tumor growth in nude mice, repressed migration and invasion abilities, and enhanced apoptosis in HCC cells. miR-186 also retarded progression of EMT. miR-186 directly bound to the 3 -untranslated regions of cyclin-dependent kinase 6 (CDK6) to inhibit its expression. Overexpression of CDK6 markedly reversed inhibitory effects of miR-186 on proliferation, apoptosis, migration, and invasion of HCC cells. Conversely, inhibition of CDK6 exerted synergic effect on the biological functions of miR-186. In conclusion, miR-186 represses proliferation, migration, invasion, and EMT, and induces apoptosis through targeting CDK6 in HCC, which may provide a new therapeutic target for HCC.