黄连素对脱氧胆酸诱导的人结肠癌细胞株增殖抑制作用及机制

研究表明，胆汁酸特别是脱氧胆酸（DCA）与结直肠癌（CRC）的发生密切相关。DCA可促进结肠癌细胞增殖，其机制可能与其增强激活蛋白-1（AP-1）活性、诱导环氧合酶-2（COX-2）表达有关。黄连素（BER）又名小檗碱，已有研究发现其对某些类型的肿瘤细胞有抑制作用。本实验采用细胞培养方法，观察BER对DCA诱导的HT-29细胞增殖、AP-1及COX-2表达的影响，旨在阐明BER抗结直肠癌的分子机制。     

姜黄素对胆汁酸诱导的人结肠癌细胞HT-29增殖的抑制作用

/L、作用6 h时和浓度≥0.5μmol/L、作用12 h时,即可抑制DCA诱导的COX-2 mRNA表达;姜黄素浓度≥1μmol/L、作用24～48 h和浓度>5 μmol/L、作用6～12 h,即可抑制DCA诱导的COX-2蛋白表达和PGE2合成.结论 姜黄素可以浓度和时间依赖模式抑制DCA诱导的HT-29细胞的增殖、COX-2 mRNA和蛋白的表达以及PGE2的合成,这可能是姜黄素抑制HT-29细胞增殖的机制之一.     

脱氧胆酸调节环氧合酶-2对结肠癌细胞增殖的影响

目的 通过观察不同浓度脱氧胆酸对结肠腺癌细胞SW1116生长的效应，以及在相应状态下细胞内环氧合酶(COX)-2蛋白表达量的改变，探讨脱氧胆酸钠对结肠癌细胞的作用机制。方法 用MTT法测定细胞增殖活性；免疫组化及Western blot方法检测细胞内COX-2蛋白的表达。结果 10～100μmol／L的脱氧胆酸钠具有明显的促进结肠腺癌细胞生长的作用；大于100μmol／L时则表现出抑制作用。脱氧胆酸钠在10、50和100μmol／L的浓度下均可促进COX-2蛋白的表达，10μmol／L的效应可以持续72h，但后两者在48h后COX-2蛋白表达开始下降。结论 脱氧胆酸对结肠癌细胞SW1116增殖的影响呈双向调节作用，脱氧胆酸促进COX-2蛋白表达可能是其促进结肠癌细胞增殖的作用途径。     

PARP-1在脱氧胆酸钠促结肠癌细胞增殖中的作用

结肠癌的发生与肠腔内脱氧胆酸引起的DNA损伤有关,聚腺苷二磷酸核糖聚合酶-1（PARP-1）对DNA损伤具有修复作用.目的：探讨PARP-1在脱氧胆酸钠促结肠癌细胞增殖中的作用及其可能机制.方法：选用不同浓度的脱氧胆酸钠、PARP-1抑制剂5-氨基异喹啉酮（5-AIQ）或两者联合分别作用于人结肠腺癌细胞株HT-29.MTT实验检测细胞增殖情况,流式细胞仪检测细胞周期和细胞凋亡,免疫细胞化学染色检测环氧合酶-2（COX-2）、caspase-3、PARP-1蛋白表达.结果：10～50 μmol/L脱氧胆酸钠能促进HT-29细胞增殖,增加COX-2、PARP-1蛋白表达,减低caspaae-3蛋自表达.100μmol/L 5-AIQ单独作用对HT-29细胞增殖无明显影响,但能减低COX-2、PARP-1蛋白表达.与单独作用相比,10μmol/L脱氧胆酸钠与100 μmol/L 5-AIQ联合能显著抑制HT-29细胞生长,阻滞细胞周期,诱导细胞凋亡,COX-2、PARP-1蛋白表达减低,caspase-3蛋白表达无明显变化.结论：COX-2与PARP-1在脱氧胆酸钠促结肠癌细胞增殖的过程中可能存在相互作用,PARP-1抑制剂5-AIQ可能用于预防脱氧胆酸诱发的结肠癌.     

脱氧胆酸诱导大肠癌发生的研究

胆汁酸中的脱氧胆酸可以促进肿瘤的形成,脱氧胆酸诱发的细胞凋亡在大肠癌的发生中起着重要作用。提高粪便中胆汁酸含量可增加患大肠癌的风险,表明脱氧胆酸可以促进大肠癌的发生、发展。脱氧胆酸作为一种促癌因子在大肠癌的发生中起很大作用,同时通过多种机制诱发大肠癌的发生。     

牛磺酸脱氧胆酸诱导HepG2细胞凋亡及坏死的研究

目的:探讨牛磺酸脱氧胆酸(TDCAO)对HepG2细胞凋亡和坏死的影响。方法:应用台盼蓝拒染试验判断细胞坏死,应用HE染色、Hoechst 33 258染色、电镜和DNA电泳对细胞凋亡定性;应用流式细胞仪对细胞凋亡定量。结果:TDCA 200 μmol/L孵育15 h可抑制细胞生长及增殖;400 μmol/L孵育 12 h可诱导显著 HepG2细胞凋亡,凋亡率为50.35±2.20％;600μmol/L孵育6 h即显著抑制细胞生长和增殖,随孵育时间延长脱落坏死细胞明显增多;800μmol/L细胞不能增殖,孵育6 h即可引起细胞坏死。结论:TDCA与HepC2细胞孵育时,即可引起细胞坏死,又可诱导细胞凋亡,取决于浓度及孵育时间的长短;在低浓度,TDCA对细胞的毒性主要表现在为抑制细胞生长增殖和诱导细胞凋亡,在高浓度则以引起细胞坏死为主。     

脱氧胆酸在人结肠癌细胞株HCT116迁移和侵袭中的作用及其机制研究

背景:脱氧胆酸(DCA)是参与肠肝循环的一种次级胆汁酸。近年研究发现DCA可促进结肠癌细胞增殖,然而其在结肠癌细胞迁移和侵袭中的作用尚未明确。目的:探讨DCA在人结肠癌细胞株HCT116迁移和侵袭中的作用及其机制。方法:以DCA刺激HCT116细胞较长时间(1~28 d),以DMSO处理的细胞作为阴性对照。采用CCK-8实验检测细胞增殖,划痕实验检测细胞迁移力,Transwell实验检测细胞侵袭力,蛋白质印迹法检测上皮型钙黏蛋白(E-cad)、神经型钙黏蛋白(N-cad)和维生素D受体(VDR)表达。结果:经12.5μmol/L DCA作用的HCT116细胞,增殖、迁移和侵袭力均较DMSO组显著增强(P0.05)。DCA组E-cad和N-cad表达水平随DCA作用时间的延长而升高,第28 d时明显高于DMSO组;DCA组VDR表达水平在第1 d时较DMSO组有所上升,其后呈下降趋势,第28 d时较DMSO组明显降低。结论:较长时间(7 d)的DCA刺激可促进结肠癌细胞迁移和侵袭,其机制可能与上调N-cad表达和下调VDR表达有关。     

普伐他汀对人结肠癌细胞增殖及COX-2蛋白表达的影响

目的体外观察普伐他汀对人结肠癌细胞HT-29、Ls-174-T细胞增殖、细胞周期及COX-2蛋白表达的影响。方法采用噻唑蓝（MTT）法观察普伐他汀对HT-29和Ls-174-T细胞增殖的影响，流式细胞仪（FCM）研究普伐他汀对细胞周期的作用，免疫细胞化学观察COX-2蛋白的表达。结果体外普伐他汀可抑制HT-29和Ls-174-T细胞增殖，各处理组内G0／G1期细胞增多，但诱导凋亡不明显，体外普伐他汀可减少HT-29和Ls-174-T细胞株COX-2蛋白的表达。结论体外普伐他汀对HT-29和Ls-174-T细胞增殖有抑制作用，该作用可能与使细胞生长阻滞于G0／G1期及抑制COX-2蛋白表达有关。     

鹅脱氧胆酸衍生物HS-1200诱导人肝癌细胞株凋亡的作用及机制

鹅脱氧胆酸衍生物HS-1200可有效地抑制肝肿瘤细胞株BEL7402的生长,其作用随药物浓度提高和作用时间延长而增强,呈一定的剂量、时间依赖性;鹅脱氧胆酸衍生物HS-1200对人肝肿瘤细胞株BEL7402有显著的生长抑制作用,该生长抑制作用与HS-1200诱导肝肿瘤细胞凋亡有关;HS-1200对正常肝细胞株无生长抑制及诱导凋亡的作用.HS-1200诱导凋亡的机制可能是HS-1200提升Bax而降低Bcl-2的表达,从而使线粒体膜通透性增高,使细胞色素C由线粒体释放入胞质,活化easpase-9,活化的caspase-9切割pro-caspase-3生成caspase-3,从而诱导凋亡;但上述凋亡过程caspase-8特异性抑制剂未改变HS-1200诱导的凋亡,因而HS-1200诱导肝肿瘤细胞凋亡途径为内源性凋亡途径.     

脂联素对结肠癌HT-29细胞的增殖及凋亡研究

目的探讨脂联素对结肠癌HT-29细胞的增殖、凋亡的影响。方法以不同浓度脂联素干预细胞,MTT法检测细胞增殖能力;AnnexinV／PI双染后流式细胞仪检测细胞凋亡。结果随着脂联素浓度的不断升高和培养时间的延长,HT-29细胞的生长逐渐受到抑制,脂联素可以诱导细胞凋亡。结论脂联素可抑制HT-29细胞增殖,诱导细胞凋亡。     

EphA2 Up-regulation induced by deoxycholic acid in human colon carcinoma cells,an involvement of extracellular signal-regulated kinase and p53-independence

Purpose The EphA2 receptor protein tyrosine kinase gene has been shown to be over-expressed or functionally altered in a number of human tumors, including colon cancer, but little is known about the regulation of this new oncoprotein. In order to explore the mechanism of EphA2 up-regulation in cancer cells, we examined the change of expression of EphA2 gene induced by deoxycholic acid (DCA) and elucidated its possible pathways in human colon cancer cells.Methods Western blot and RT-PCR were used to assess the protein expression and messenger RNA in several colon cancer cell lines, which harbor various p53 status. The inhibition study to interfere the MAPK pathway was performed by using various chemicals and by transfecting dominant negative mutant plasmids.Results Up-regulation of EphA2 induced by DCA was observed in a dose- and time-dependent fashion both in mRNA and protein levels. This regulation is constant regardless of p53 status including wild, mutant or knocked out in the colon cell lines used. This induction was in part blocked by either erk1/2 inhibitors or dominant negative mutants erk1/2 plasmids.Conclusions These results suggest that DCA induced up-regulation of EphA2 in colon cancer cells is due to activation of erk1/2 cascade, and is p53-independent. Taken together with the roles of EphA2 and DCA in tumorigenesis, which have been independently reported, our observation will provide a new mechanistic basis of DCA commitment in carcinogenesis.     

Effects of inositol hexaphosphate on proliferation of HT-29 human colon carcinoma cell line

AIM: To investigate the effects of inositol hexaphosphate (IP6) on proliferation of HT-29 human colon carcinoma cell line. METHODS: Cells were exposed to various concentrations (0, 1.8, 3.3, 5.0, 8.0, 13.0 mmol/L) of IP6 for a certain period of time. Its effect on growth of HT-29 cells was measured by MTT assay. The expressions of cell cycle regulators treated with IP6 for 2 d were detected by immunocytochemistry. RESULTS: IPe inhibited the HT-29 cell growth in a dose- and time-dependent manner. Analysis of cell cycle regulator expression revealed that IPe reduced the abnormal expression of P53 and PCNA and induced the expression of P21. CONCLUSION: IPe has potent inhibitory effect on proliferation of HT-29 cells by modulating the expression of special cell cycle regulators.     

舒林酸对结肠腺癌HT-29细胞增殖调控的实验研究

目的 观察舒林酸对结肠腺癌ＨＴ－２９细胞增殖的影响,探讨其作用机制。方法 采用ＭＴＴ比色法观察舒林酸对ＨＴ－２９细胞增殖的影响;采用流式细胞仪检测舒林酸对ＨＴ－２９细胞周期的影响,同时结合ＤＮＡ电泳和透射电镜观察舒林酸对ＨＴ－２９细胞有无促凋亡作用。结果 舒林酸可抑制ＨＴ－２９细胞增殖,使Ｇ０／Ｇ１期细胞比例增高,Ｓ期比例降低,７２ｈ后,ＨＴ－２９细胞凋亡分别上升至１２．５＾、１５．４＾和２４．２     

黄连素抑制结肠癌细胞环氧合酶-2的作用

目的 探讨黄连素对结肠癌细胞系HT—29的作用及其相关机制,为阐明黄连素作为一种新的结肠癌化学治疗药物进行理论上的准备并提供相关实验结果。方法 黄连素0．1、0．3、3．0、30．0μmol／L加入到HT—29结肠癌细胞系培养液中,每天测量各有关值。同时以NS—398为环氧合酶(COX-2)活性抑制剂,对比观察黄连素对COX—2活性的影响。采用氮蓝四唑盐实验法检测细胞的生长和增殖,逆转录—PCR法检测COX—2 mRNA,免疫细胞化学法检测COX—2蛋白表达,ELISA法检测前列腺素(PG)E2的含量。结果 黄连素在浓度大于0．3μmol／L时则有明显的量效关系。黄连素在浓度大于0．3μmol／L时对COX—2 mRNA水平和蛋白的表达有明显的抑制作用。黄连素对PGE2的生成有显著抑制作用。结论 黄连素抑制COX—2在mRNA水平和蛋白质水平的表达,同时也抑制COX—2活性,进而抑制PGE2的生成,这可能为黄连素抑制HT—29细胞生长和增殖的机制之一。     

Modulation of colon cancer cell invasiveness induced by deoxycholic acid]

BACKGROUND/AIMS: Deoxycholic acid (DCA), a secondary bile acid, has been implicated to promote colon cancer growth and progression. However, its molecular mechanisms are largely unknown. In this study, we investigated the effects of DCA on proliferation, migration, and invasiveness of colon cancer cells (HT-29). METHODS: HT-29 cells were incubated with either medium (control) only or DCA for 24-48 hours. Time courses of RT-PCR for vascular endothelial growth factor (VEGF) and hypoxia-inducible factor (HIF)-1alpha mRNA expression, Western blotting for VEGF and matrix metalloproteinase (MMP)-9, zymography for MMP-9 activation, and wound-migration assay were determined after various concentrations of DCA (0-80 microM) treatment. Moreover, these experiments were reassessed after pretreatments (2-6 hours) with specific inhibitors of various signal pathways. RESULTS: DCA enhanced HIF-1alpha mRNA expression, VEGF mRNA and VEGF protein expression, MMP-9 protein expression/activation, and cell migration ability in a dose-related manner. DCA-induced VEGF protein expression was inhibited by pretreatment with NS-398 (COX-2 inhibitor), PDTC (NF-kappaB inhibitor), or tauroursodeoxycholic acid (TUDC). DCA-induced cell migration ability was inhibited by pretreatment of GF109203X, a protein kinase C inhibitor. DCA-induced MMP-9 protein expression/activation was inhibited by pretreatment with SB203580, U0126, or PDTC. CONCLUSIONS: DCA significantly upregulates invasive and angiogenic potentials of human colon cancer cells through multiple signal transduction pathways.