辛伐他汀抑制人乳腺癌MCF-7细胞内多能干细胞标志物Oct3/4 Nanog和Sox-2表达

  目的  研究辛伐他汀（simvastatin,SIM）对人乳腺癌MCF-7细胞内多能干细胞标志物Oct3/4、Nanog和Sox-2表达的影响。  方法  应用实时荧光定量聚合酶链式反应（qRT-PCR）技术、免疫荧光染色方法,流式细胞仪和免疫印迹（Western blot）方法检测SIM对MCF-7细胞内多能干细胞标志物表达的影响。  结果  qRT-PCR结果显示,10、50和100 μmol/L SIM作用于MCF-7细胞48h后,能显著抑制细胞内Oct3/4、Nanog和Sox-2基因表达,与对照组相比,差异有统计学意义（P < 0.05）,而SIM 1 μmol/L浓度组和对照组相比差异无统计学意义（P>0.05）。SIM 50、100 μmol/L浓度组和10 μmol/L浓度组相比,抑制Oct3/4和Nanog的表达差异有统计学意义（P < 0.05）,而对Sox-2的表达抑制,SIM 10、50 μmol/L和100 μmol/L各浓度组间差异无统计学意义（P>0.05）。免疫荧光染色显示经10 μmol/L SIM处理48 h后MCF-7细胞核内Oct3/4、Nanog和Sox-2蛋白表达减弱,部分细胞核无表达。流式细胞检测显示MCF-7细胞经10 μmol/L SIM处理48 h后,Oct3/4阳性细胞数、Nanog阳性细胞数和Sox-2阳性细胞数显著减少（P < 0.05）,Western blot进一步证实经10 μmol/L SIM处理48 h后MCF-7细胞核内Oct3/4、Nanog和Sox-2蛋白表达显著减少（P < 0.05）。  结论  SIM在体外能有效地抑制人乳腺癌MCF-7细胞内多能干细胞标志物Oct3/4、Nanog和Sox-2的表达,为SIM应用于癌症治疗提供实验依据。      

肝癌细胞系中Oct4与Wnt/β-catenin 和TGF-β信号通路的相互影响

 目的 探讨干细胞相关基因Oct4与Wnt/β-catenin 及TGF-β信号通路在肝癌细胞系中的相互作用。方法 应用RT PCR法检测Oct4、Wnt/β-catenin及TGF-β信号通路相关基因β-catenin、Wnt10b、TCF3及 ELF、Smad3和 Smad4在肝癌组织及细胞系中的表达;使用siRNA 沉默人肝癌HepG2细胞Oct4和TCF3的表达,实时荧光定量RT PCR法检测Wnt10b、β-catenin、 TCF3及 ELF、Smad3和 Smad4等基因的表达变化。结果 Oct4和β catenin、Wnt10b、TCF3及 ELF、Smad3和 Smad4在肝癌组织及细胞系中同时表达; siRNA-Oct4 沉默人肝癌HepG2细胞Oct4后,Oct4表达明显下调,β-catenin、Wnt10b随之下调达40%~50%左右,而TCF3表达升高到3倍左右;同时ELF、Smad3和 Smad4均下降到原来的1%以下;而siRNA-TCF3 沉默TCF3后,Oct4的表达也升高2~3倍;ELF升高亦达2~3倍,Smad3和 Smad4也略有升高。结论 Oct4和β-catenin、Wnt10b、TCF3及 ELF、Smad3和 Smad4在肝癌组织及细胞系中同时表达提示彼此之间有相互作用。RNAi实验证明Oct4对Wnt/β-catenin及TGF-β信号通路的成员有调控作用;Oct4 与TCF3之间的负反馈作用值得深入研究。     

通过 CRISPR/Cas9 技术抑制 TFDP3 基因对前列腺癌 PC3 细胞生物学功能的影响

[ 摘 要 ] 目的：通过 CRISPR/Cas9 技术构建前列腺癌 PC3 细胞 TFDP3 基因敲除的稳转株,探讨抑制 TFDP3 表达对 PC3 细 胞周期、凋亡、迁移和侵袭能力的影响。方法：通过生物信息学筛选 sgRNA,通过 CRISPR/Cas9 技术、构建抑制 TFDP3 基因表达 的 sgRNA-Cas9 共转染慢病毒,感染 PC3 细胞后筛选获取稳转细胞株。通过流式细胞术对 TFDP3 基因敲除（knock-out,KO） 实验组与空白对照组进行细胞周期和凋亡检测,并进一步通过划痕实验和 Transwell 实验进行细胞迁移和侵袭能力检测。 结果：通过生物信息学筛选获得 3 条 sgRNA,其中 sgRNA2 有明显的抑制前列腺癌细胞基因表达的功能;通过 CRISPR/ Cas9 技术成功构建了基于 CRISPR/Cas9 介导的 TFDP3 低表达的 PC3 细胞稳转株。抑制 TFDP3 基因表达后,相比于对照组, KO 组中 G0/G1 期细胞百分比增加、G2/M 期细胞百分比下降（P<0.05 或 P<0.01）,细胞凋亡率显著升高（P<0.05）,细胞迁移率 明显下降 [24 h 迁移率：(44.00±1.60)% vs (65.00±4.40)%,P<0.01],穿过聚碳酸酯膜的侵袭细胞数明显下降 [（185.89±11.71）vs （248.33±11.95）个,P<0.01]。结论：通过 CRISPR/Cas9 技术抑制 TFDP3 基因表达后,PC3 细胞发生周期阻滞、凋亡率也有所增 加、迁移和侵袭能力显著减弱,提示 TFDP3 是一个前列腺癌促癌基因。     

利用CRISPR/Cas9技术构建CYP2E1基因敲除的HEK293FT细胞系

目的：利用CRISPR/Cas9系统在HEK293FT细胞系中敲除CYP2E1基因并探讨其在检测化学物毒性中的应用。方法：设计3对分别靶向CYP2E1基因第2、第3和第7外显子的sgRNA序列并构建PX461表达载体;将携带靶向CYP2E1的sgRNA表达载体转染至HEK293FT细胞中,通过SURVEYOR检测分析sgRNA的切割效率;利用有限稀释法获得CYP2E1敲除细胞株,通过实时荧光定量PCR和蛋白印迹检测细胞株中CYP2E1的敲除效果;并检测其对N-（4-羟基苯基）乙酰胺和1,2-二氯乙烷的细胞毒性效应的影响。结果：SURVEYOR检测分析靶向CYP2E1基因第3外显子的sgRNA的切割效率为23%;成功构建了2株CYP2E1基因敲除的细胞株,实时荧光定量PCR结果显示CYP2E1 mRNA表达下降,蛋白印迹结果显示CYP2E1蛋白无表达;CYP2E1基因敲除会显著降低N-（4-羟基苯基）乙酰胺和1,2-二氯乙烷的细胞毒性。结论：通过CRISPR/Cas9构建出CYP2E1稳定敲除的肾细胞系,为研究CYP2E1的功能和作用机制提供了有效的工具。     

Oct4、Stat3在卵巢浆液性/黏液性囊腺癌中的表达及临床意义

目的:探讨Oct4、Stat3在卵巢浆液性/黏液性囊腺癌中的表达及其相关性。 方法:应用组织芯片,采用免疫组化SP法检测60例卵巢浆液性/黏液性囊腺癌组织中Oct4、Stat3的表达。结果:Oct4在卵巢浆液性/黏液性囊腺瘤中的阳性表达率为35.0%（7/20）,低于卵巢浆液性/黏液性囊腺癌中的83.3%（50/60）,两者有显著统计学差异（P<0.01）。卵巢浆液性/黏液性囊腺癌中Oct4的阳性表达率与患者年龄、临床分期、组织学分化程度、患者组织学类型均无相关性（P>0.05）。Stat3在卵巢浆液性/黏液性囊腺癌阳性表达率为71.7%（43/60）,明显高于卵巢浆液性/黏液性囊腺瘤中的35.0%（7/20）,两者有显著统计学差异（P<0.01）。Stat3在Ⅰ、Ⅱ期癌中的阳性表达率低于Ⅲ、Ⅳ期癌（P<0.05）,在中、低分化癌中的阳性表达率高于高分化癌（P<0.05）,与患者组织学类型无关。卵巢浆液性/黏液性囊腺癌中Oct4表达与Stat3表达呈正相关（P<0.05）。结论:Oct4、Stat3高表达可能与卵巢浆液性/黏液性囊腺癌的发生、发展密切相关,且相互间可能具有协同作用。     

耐药喉癌Hep-2/VCR细胞ATP7B表达与顺铂耐药的相关性

目的 检测copper transporting P-type adenosine triphosphatase(ATP7B)在人喉癌耐药细胞株中的表达,并研究其与喉癌细胞顺铂耐药的关系.方法 人喉鳞癌Hep-2细胞和Hep-2/长春新碱(vincristine,VCR)耐药细胞分别给予顺铂作用24小时和48小时,应用细胞增殖/毒性检查试剂盒(cell counting kit-8,CCK8)细胞活力试验、末端脱氧核苷酸转移酶介导的dUTP缺口末端标记测定法[terminal dexynueleotidyl transferase (TdT)-mediated dUTPnick end labeling,TUNEL]细胞凋亡实验分别检测细胞增殖和凋亡.分别用免疫荧光染色和实时RT-PCR检测ATP7B蛋白和mRNA在细胞中的表达.结果 50 μmol/L和l00μmol/L的顺铂作用24小时后,Hep-2细胞活力下降为71.0％,而Hep-2/VCR细胞活力无下降,差异有统计学意义(P＜0.01).50 μmol/L顺铂能够诱导Hep-2和Hep-2/VCR细胞表达ATP7B.Hep-2细胞在作用后3小时表达增强,6小时达到峰值,12小时下调(P＜0.05);而Hep-2/VCR细胞在3小时表达增强,6小时达到峰值(P＜0.05),持续高表达24小时无明显变化.结论 喉癌耐药细胞Hep-2/VCR的ATP7B蛋白的高表达与喉癌细胞的顺铂耐药有关.     

EGFR/CerbB-2/MAPK介导乳腺癌三苯氧胺耐药细胞的生长

目的探讨乳腺癌三苯氧胺(TAM)获得性抵抗细胞的生长调节途径及三苯氧胺(TAM)获得性抵抗的发生机制.方法构建TAM抵抗的人乳腺癌细胞系(MCF-7/TAMR),RT-PCR、Westem blot及免疫细胞化学方法比较MCF-7/TAMR与MCF-7/WT细胞系中EGFR(CerbB-1)、CerbB-2、CerbB-3、CerbB-4 mRNA及蛋白表达的不同.结果与MCF-7/WT细胞相比,MCF-7/TAMR细胞中表皮生长因子受体EGFR的mRNA增加6倍(P＜0.05),蛋白增加3倍(P＜0.05);CerbB-2的mRNA增加3倍(P＜0.05),蛋白增加1.5倍(P＜0.05);CerbB-3在两种细胞中的表达相似;CerbB-4未能检测到;MCF-7/TAMR细胞中MAPK活化水平增高;基础条件下,MCF-7/TAMR细胞中磷酸化的EGFR/CerbB-2,EGFR/CerbB-3异二聚体与细胞外信号调节激酶磷酸化水平增高有关;Herceptin处理MCF-7/TAMR细胞,阻滞了CerbB-2受体异二聚体的形成及磷酸化,降低了细胞外信号调节激酶的活性,明显抑制了细胞的生长.结论表皮生长因子受体特异性配体的自分泌释放作用诱导EGFR/CerbB-2二聚体形成及下游细胞外信号调节激酶(MAPK)的活化引起MCF-7/TAMR细胞的生长.     

维甲酸联合他莫昔芬对人乳腺癌细胞株作用的研究

目的:探讨他莫昔芬(TAM)与维甲酸(ATRA)联合对人乳腺癌他莫昔芬耐药株的作用.方法:在体外培养条件下,分别或联合应用ATRA和TAM作用于MCF-7人乳腺癌他莫昔芬耐药株(MCF-7/T)及敏感组(MCF-7/S).MTT比色法分析细胞生长抑制作用;用流式细胞仪(FCM)检测细胞周期分布、凋亡率及用药前后Bcl-2、Bax、Fas和FasL蛋白的变化.结果:TAM能抑制ER阳性MCF-7/S的生长,阻滞细胞周期于G0/G1期,并可诱导细胞凋亡,TAM不能抑制MCF-7/T的生长; ATRA预处理细胞24 h后,TAM抗乳腺癌细胞MCF-7/S的作用增强,且恢复了MCF-7/T对TAM的敏感性.ATRA与TAM联用后,MCF-7/T细胞Bcl-2蛋白表达下调,细胞Bax、Fas和FasL蛋白表达水平未见明显变化.结论:体外条件下,TAM通过影响细胞周期、诱导细胞凋亡而发挥抗ER阳性MCF-7/S作用;视黄酸能加强TAM对激素敏感细胞MCF-7/S的抗乳腺癌作用,恢复耐受细胞MCF-7/T对TAM的敏感性.同时恢复TAM对MCF-7/T的促凋亡作用.     

米非司酮逆转人乳腺癌细胞MCF-7/ADR耐多柔比星机制的探讨

目的 探讨米非司酮对乳腺癌耐药细胞株MCF-7/ADR的逆转耐药作用.方法 以亲本乳腺癌细胞MCF-7和耐多柔比星(阿霉素)乳腺癌细胞MCF-7/ADR为研究对象,分别应用MIF进行干预后,流式细胞仪检测MIF作用前后瘤细胞P-gp的表达、细胞内阿霉素蓄积量的变化以及细胞周期的分布.结果 (1)10 μmol/L MIF作用72小时后,MCF-7/ADR细胞P-gp表达率[(23.21±1.80)％]明显高于MCF-7细胞[(19.37±2.37)％,P＜0.05].(2)5 μmol/L ADR处理后,MCF-7/ADR细胞内ADR蓄积量为(47.13±4.11)％,低于MCF-7细胞[(60.24±2.61)％,P＜0.05].(3) 10 μmol/L MIF联合5 μmol/L ADR处理细胞,MCF-7/ADR和MCF-7细胞内ADR的蓄积量分别为(82.72±2.42)％及(88.63±2.75)％ (P ＞0.05);但均较单用ADR时升高(均P＜0.01).(4) MIF作用前,MCF-7/ADR细胞G0/G1期比例[(77.21±3.10)％]高于MCF-7细胞G0/G1期比例[(59.05±2.16)％,P＜0.05];MCF-7/ADR细胞S期比例明显低于MCF-7细胞(P＜0.05).经10 μmol/L MIF作用后,MCF-7细胞G0/G1期比例(75.28±2.53)％较MIF作用前明显升高(P＜0.05);S期比例则较MIF作用前显著降低(P＜0.05);MCF-7/ADR细胞G0/G1期比例和S期比例分别为(80.13±2.72)％及(13.52±1.03)％,与MIF作用前比较差异均无统计学意义(均P＞0.05);两种瘤细胞的G2/M期比例与MIF作用无关(P＞0.05).结论 (1)MIF可以逆转MCF-7/ADR的耐药性,其作用机制与降低细胞P-gp含量、增加细胞内ADR蓄积量有关.(2) 10 μmol/L浓度的MIF对MCF-7/ADR细胞周期分布影响不大.     

SUMOylation of HSP27 by small ubiquitin-like modifier 2/3 promotes proliferation and invasion of hepatocellular carcinoma cells

Primary hepatocellular carcinoma (PHC) is a major health problem worldwide and is one of the 10 most commonly diagnosed cancers in China. Heat shock protein 27 (HSP27) were found to be overexpressed in a wide range of malignancies including PHC, however, post-translational modification of HSP27 still needs exploration in PHC. Recently, SUMOylation, an important post-translational modification associating with the development of many kinds of cancers has been intensively studied. In the current study, mRNA and protein level of HSP27 in archived tumor samples representing various pathological characteristics of PHC were examined, and modification of HSP27 by SUMO2/3 was investigated. HSP27 were expressed abundantly in patients' tumor tissues, and found to be associated with pathological progression. Besides, HSP27 was also elevated significantly in liver cancer cell lines Huh7 and HepG2 compared with human hepatocyte cells L02. Furthermore, knockdown of HSP27 was found to be associated with the decreased proliferation and invasion ability in Huh7 and HepG2 cells. Immunofluorescence assay showed that HSP27 and SUMO2/3 were co-localized in the subcellular, and co-immunoprecipitation verified the interaction between HSP27 and SUMO2/3. Overexpression of SUMO2/3 upregulated the HSP27 protein level and promotes Huh7 and HepG2 cell proliferation and invasion, and vice versa when the SUMO2/3 was knockdown. Taken together, increased protein level of HSP27 through SUMO2/3-mediated SUMOylation plays crucial roles in the progression of PHC, and this finding may shed light on developing potential therapeutic targets for PHC.     

Hsp90抑制剂减弱肝癌耐药细胞株对索拉非尼诱导铁死亡的耐受性北大核心

目的:检测热休克蛋白90(heat shock protein 90,Hsp90)抑制剂协同索拉非尼(Sorafenib)对肝癌耐药细胞的作用机制,并研究其对Sorafenib诱导肝癌耐药细胞铁死亡的增敏效应。方法:MTT法检测格尔德霉素(Geldanamycin)联用Sorafenib在抑制HepG2/Sorafenib细胞增殖中的联合效应;流式细胞术检测HepG2/Sorafenib及HepG2细胞株在Sorafenib作用下,细胞内活性氧(reactive oxygen species,ROS)水平的变化,以及Geldanamycin对细胞内ROS的协同诱导效应;FerroOrange染色法检测HepG2/Sorafenib及HepG2细胞株在Sorafenib作用下,细胞内二价铁离子水平变化;RT-PCR及Western blot实验检测Geldanamycin对铁死亡相关基因SLC7A11及GPX4等转录和表达的调节作用。结果:Geldanamycin可增强HepG2/Sorafenib耐药细胞株对Sorafenib的敏感性,经30 nmol/L浓度的Geldanamycin预处理后,Sorafenib的半数抑制率IC_(50)由原来的(2.6±0.1)×10^(3) nmol/L降为(37.2±6.9)nmol/L,变化差异显著(P<0.01)。当Sorafenib浓度高于32 nmol/L后,二者的联用指数CI小于0.2,表明Geldanamycin与Sorafenib联用处理HepG2/Sorafenib耐药细胞株具有强烈的协同效应;HepG2/Sorafenib耐药细胞株经30 nmol/L Geldanamycin预处理后,其细胞内的ROS水平可重新被Sorafenib诱导,相同浓度下与无Geldanamycin处理组比较,增强了4.2倍(P<0.01);30 nmol/L Geldanamycin可显著增强Sorafenib诱导HepG2/Sorafenib细胞内二价铁离子水平提高,提示其协同Sorafenib促细胞铁死亡作用;Geldanamycin能通过Akt-Nrf2途径下调SLC7A11蛋白的转录,并影响GPX4蛋白的表达。结论:抑制Hsp90能减弱肝癌耐药细胞株对Sorafenib诱导铁死亡的耐受性,其机制可能是通过阻断Akt/Nrf2介导的SLC7A11转录下调实现的。     

Tripartite motif-containing 3 (TRIM3) inhibits tumor growth and metastasis of liver cancer

Background:Reduced expression of tripartite motif-containing 3 (TRIM3) has been reported to be involved in the pathogenesis of human glioblastoma.In our previous research,we found that TRIM3 expression was markedly reduced in human primary hepatocellular carcinoma (HCC) tissues and that low TRIM3 expression was associated with short survival of HCC patients.However,the role of TRIM3 in liver cancer remains unknown.This study aimed to investigate the function of TRIM3 in liver cancer cells.Methods:The protein levels of TRIM3 in five liver cancer cell lines (SK-Hep1,Hep3B,Huh7,HepG2,Bel-7402) and one normal liver cell line (L02) were detected with Western blotting.HepG2 and Bel-7402 cells with IowTRIM3 expression were infected with recombinant lentiviruses overexpressing TRIM3 (LV-TRIM3),whereas Huh7 and Hep3B cells with high TRIM3 expression were transfected with TRIM3-targeted small interfering RNA (siTRIM3).The functions of TRIM3 in the proliferation,colony formation,cell cycle,migration,invasion,and apoptosis of the above cell lines were examined.The effect of TRIM3 on tumor growth and metastases in nude mice was also investigated.Results:TRIM3 was overexpressed in HepG2 and Bel-7402 cells with LV-TRIM3 infection,which further reduced proliferation,colony formation,migration,and invasion of both cell lines.Cell cycle analysis showed thatTRIM3 overexpression induced G0/G1 phase arrest in HepG2 and Bel-7402 cells.Moreover,apoptosis was not increased in HepG2 or Bel-7402 cells overexpressing TRIM3.Contrarily,silencing TRIM3 expression in Huh7 and Hep3B cells by siTRIM3 led to significantly decreased percentages of both cells in the G0/G1 phase and promoted cell proliferation,colony formation,migration,and invasion.In vivo experiment results confirmed thatTRIM3 overexpression suppressed tumor growth and metastasis.Conclusions:TRIM3 plays a tumor-suppressing role in the regulation of liver cancer development by reducing cell proliferation through cell cycle arrest at the G0/G1 phase.     

Sodium glucose cotransporter 2 inhibitor canagliflozin attenuates liver cancer cell growth and angiogenic activity by inhibiting glucose uptake

Sodium‐glucose cotransporter 2 inhibitors (SGLT2‐Is) comprise a new class of antidiabetic agents that inhibit glucose reabsorption in the renal proximal tubules. Although a recent report demonstrated the potential ability of SGLT2‐Is to attenuate cancer growth of SGLT2‐expressing cancer cells, little is known about the effects of SGLT2‐Is on hepatocellular carcinoma (HCC). Here, we investigate the anti‐cancer properties of a SGLT2‐I, canagliflozin, against human liver cancer cells. SGTL2 mRNA and protein expression were detected in Huh7 and HepG2 cells, although not in HLE as well as primary human hepatocytes and hepatic stellate cells. Canagliflozin exerted antiproliferative effects on SGLT2‐expressing Huh7 and HepG2 cells in a dose‐dependent manner by inhibiting glycolytic metabolism including glucose uptake, lactate and intracellular ATP production. This agent also induced G2/M arrest and apoptosis with inhibited phosphorylation of ERK, p38 and AKT and cleavage of caspase3. Xenograft tumor growth assay showed that oral administration of canagliflozin (10 mg/kg/day) significantly reduced subcutaneous tumor burdens in a glycemic status‐independent manner, and attenuated intratumor vascularization in Huh7‐ and HepG2‐derived xenograft tumors in BALB/c nude mice. In vitro, canagliflozin suppressed the increased human umbilical vein endothelial cell (HUVEC) proliferation and tubular formation which were observed in Huh7 or HepG2 co‐cultures. By contrast, canagliflozin had no effect on tumor growth and intratumor angiogenesis in SGLT2‐null HLE‐derived xenograft models. These results indicate that SGLT2‐I therapy is a potential new strategy for the treatment of HCC.     

肝癌多药耐药细胞株的建立及其多药耐药机理的研究

 为建立人肝癌多药耐药细胞株并研究其多药耐药的机理，本文应用BEL－7402细胞株，通过不断提高培养液中阿霉素（Doxorubicin）的浓度，长期筛选培养，得到肝癌多药耐药株BEL7402/DoX。经MTT法检测BEL－7402对长春新碱（VCR）等8种抗癌药的抗性提高了27－1100倍不等。以流式细胞技术检测了此细胞株表面MDR1蛋白P－gp、多药耐药相关蛋白MRP及谷胱甘肽硫转移系统（GSH/GST）的表达；用RT－PCR方法检测了MDR及MRP基因表达水平。发现BEL7402/Dox细胞表面P－gp表达为93.5～97.4％；MRP的表达为84.7～90.2％；RT－PCR证实此细胞株中有MDR及MRPmRNA的高表达；未发现GSH/GST的表达升高。     

沉默环状RNA hsa_circ_0000591表达对肝癌细胞增殖及迁移的影响

目的探讨环状RNA hsa_circ_0000591在肝细胞癌(hepatocellular carcinoma,HCC)组织以及肝癌细胞系中的表达,及其对肝癌细胞增殖和迁移的影响。方法收集2014—2018年广西医科大学附属肿瘤医院手术切除的84例HCC组织及其相应癌旁正常组织标本,采用qRT-PCR检测hsa_circ_0000591的表达,并分析其与预后的关系。将hsa_circ_0000591沉默慢病毒及阴性对照慢病毒感染肝癌Hep3B细胞,记为siRNA组和NC组。采用CCK-8实验、平板克隆形成实验检测细胞增殖能力;划痕实验检测细胞迁移能力;Transwell实验检测细胞侵袭和迁移能力。结果qRT-PCR检测结果显示,hsa_circ_0000591在HCC组织中的表达高于癌旁组织(P<0.001),在肝癌细胞系Li-7、HepG2、Hep3B、Huh7及人体正常肝细胞系HL-7702中的表达差异有统计学意义(P<0.05),其中在Hep3B细胞中的表达量高于其余肝癌细胞系及正常肝细胞系HL-7702(均P<0.05)。与NC组比较,沉默hsa_circ_0000591后肝癌Hep3B细胞中的hsa_circ_0000591表达下调(P<0.05),细胞增殖、迁移和侵袭能力降低(P<0.05)。结论hsa_circ_0000591在肝癌中表达上调,沉默hsa_circ_0000591可抑制肝癌细胞增殖和迁移,hsa_circ_0000591可能是肝癌潜在的分子靶点。     

Farnesyltransferase inhibitor, ABT-100, is a potent liver cancer chemopreventive agent.

PURPOSE: Treatment of hepatocellular carcinoma raised on cirrhotic liver represents a major endeavor because surgery and chemotherapeutic management fail to improve the clinical course of the disease. Chemoprevention could represent an important means to inhibit the process of hepatocarcinogenesis. Farnesyltransferase inhibitors are a class of drugs blocking the growth of tumor cells with minimal toxicity towards normal cells. EXPERIMENTAL DESIGN: In the present study, we investigated the effects of a novel farnesyltransferase inhibitor, ABT-100, on human liver cancer cell lines, HepG2 and Huh7, and on an animal model of hepatocarcinogenesis. RESULTS: ABT-100 inhibited HepG2 and Huh7 cell growth as well as the invading ability of Huh7 on Matrigel. In HepG2 and Huh7 cells, ABT-100 inhibited growth factor-stimulated phosphoinositide 3-kinase and Akt/protein kinase B activity. Furthermore, ABT-100 inhibited Akt-dependent p27(Kip1) phosphorylation and this event was associated with increased levels of p27(Kip1) in the nucleus and reduced activity of the cyclin-dependent kinase 2. Moreover, ABT-100 treatment resulted in a significant reduction in tumor incidence and multiplicity. CONCLUSIONS: Taken together, these findings identify a mechanism of ABT-100 function and show the efficacy of ABT-100 as a chemopreventive agent of hepatocellular carcinoma.     

鸦胆子油对人肝癌细胞增殖的抑制作用及机制

目的:探讨鸦胆子油对人肝癌HepG2和Huh7细胞的增殖抑制作用及机制。方法:采用MTT法分析不同浓度鸦胆子油的增殖抑制作用,并用流式细胞仪进行细胞周期和细胞凋亡分析。结果:鸦胆子油能够使HepG2细胞和Huh7细胞的存活率降低,HepG2细胞24h和48h的IC50值分别为59.41μl/L和45.35μl/L,Huh7细胞24h和48h的IC50值为273.43μl/L和103.52μl/L。细胞周期实验中细胞G0/G1期比例升高。细胞凋亡实验中HepG2和Huh7细胞早期凋亡和晚期凋亡的比例均有增加,但晚期凋亡的细胞比例增加更大,分别为(61.47±7.74)%和(69.34±6.42)%。 结论:鸦胆子油能够抑制HepG2细胞和Huh7细胞的增殖,且通过阻滞细胞周期于G0/G1期诱导细胞凋亡。