羟基脲对人脑胶质瘤U251细胞放化疗增敏作用的研究

目的:探讨羟基脲(HU)联合替莫唑胺(TMZ)加放疗(RT)对人脑胶质瘤U251细胞放化疗(CRT)敏感性的影响。方法:体外培养U251细胞,采用CCK8实验检测不同浓度HU、TMZ及不同条件处理后细胞的增殖能力;流式细胞术检测细胞凋亡及细胞周期分布情况;Transwell小室、划痕实验评估细胞侵袭、迁移能力变化;We...     

替莫唑胺通过调控miR-216a/PRKCA抑制胶质瘤细胞U251增殖和侵袭作用的研究

目的:探讨替莫唑胺抑制胶质瘤细胞U251增殖和侵袭的作用,推测其作用机制是通过微小RNA-216a（microRNA-216a,miR-216a）/蛋白激酶Cα(protein kinase C-alpha,PRKCA)进行调控。方法:体外培养胶质瘤细胞U251,用不同剂量替莫唑胺染毒48 h后,构建野生型PRKCA 3' UTR-荧光素酶报告载体及检测荧光素酶活性,检测替莫唑胺对胶质瘤细胞U251增殖和侵袭的影响及miR-216a和PRKCA表达的影响。结果:miR-216a mimics使野生型PRKCA 3' UTR-荧光素酶报告载体的活性下降了47.52%,差异具有统计学意义（P＜0.05）;miR-216a mimics使突变型PRKCA 3' UTR-荧光素酶报告载体的活性下降不显著,差异无统计学意义（P＞0.05）。与空白对照组比较,低、中、高剂量组胶质瘤细胞U251增殖率、侵袭细胞数、PRKCA mRNA相对表达量和PRKCA蛋白表达量均降低,miR-216a mRNA相对表达量均增高,差异具有统计学意义（P＜0.05）;随着替莫唑胺剂量的增加,胶质瘤细胞U251增殖率、侵袭细胞数、PRKCA mRNA相对表达量和PRKCA蛋白表达量随之降低,miR-216a mRNA相对表达量随之增高,差异具有统计学意义（P＜0.05）。结论:替莫唑胺可以通过促进miR-216a的表达而抑制PRKCA的表达,降低胶质瘤细胞U251增殖和侵袭。     

FUBP1基因在脑胶质瘤中的表达及其对U251细胞凋亡的影响

目的：研究FUBP1基因在脑胶质瘤中的表达及其对胶质瘤U251细胞凋亡的影响。方法：Real-time PCR检测35例脑胶质瘤及相应癌旁组织中FUBP1 mRNA的表达。设计并合成FUBP1基因特异性的siRNA,转染U251细胞。转染后,western blot检测转染后FUBP1蛋白的表达,双标流式细胞法检测细胞凋亡率。结果：与癌旁组织相比,FUBP1 mRNA在脑胶质瘤组织中表达明显上调。转染后,U251细胞中的FUBP1蛋白表达明显降低,同时,凋亡率明显增加。结论：FUBP1基因在脑胶质瘤中存在表达异常,其表达降低能够促进细胞凋亡。     

FUBP1基因在脑胶质瘤中的表达及其对U251细胞凋亡的影响

目的:研究FUBP1基因在脑胶质瘤中的表达及其对胶质瘤U251细胞凋亡的影响。方法:Real-time PCR检测35例脑胶质瘤及相应癌旁组织中FUBP1 mRNA的表达。设计并合成FUBP1基因特异性的siRNA,转染U251细胞。转染后,western blot检测转染后FUBP1蛋白的表达,双标流式细胞法检测细胞凋亡率。结果:与癌旁组织相比,FUBP1 mRNA在脑胶质瘤组织中表达明显上调。转染后,U251细胞中的FUBP1蛋白表达明显降低,同时,凋亡率明显增加。结论:FUBP1基因在脑胶质瘤中存在表达异常,其表达降低能够促进细胞凋亡。     

p53基因对胶质瘤细胞系U251生长抑制作用的研究

背景与目的：肿瘤抑制基因p53是调节多种与细胞周期、凋亡、DNA修复等有关基因表达的转录因子。p53基因在大约30％的胶质瘤中发生突变，在胶质瘤的发生和发展中起重要作用。本文主要探讨野生型p53基因过表达对脑胶质瘤细胞系U251细胞生长抑制的机制。方法：通过p53腺病毒表达载体pAdCMV-p53及空载体pAdCMV-lacZ分别感染U251细胞系，RT-PCR及Westem blot方法检测转染效率；并通过MTT检测生长抑制率、流式细胞仪检测细胞周期及TUNEL检测分析细胞凋亡等指标观察p53基因对U251细胞生长的影响。结果：MOI为100时，野生型p53基因的过表达可引起U251细胞G0、G1期阻滞、诱导U251细胞凋亡以及引起U251细胞生长抑制。结论：p53基因可以通过细胞周期G0、G1期阻滞及诱导细胞凋亡抑制胶质瘤细胞系U251的生长。     

RORα miRNA转染对DADS抑制胶质瘤U251细胞增殖的影响

背景与目的:维甲酸相关孤核受体α(retinoid acid receptor related orphan receptor α,ROR α)可能参与肿瘤的调控.本实验室前期研究发现,二烯丙基二硫(diallyl disulfide,DADS)可抑制人胶质瘤U251细胞增殖,其抑制增殖作用可能与诱导ROR α蛋白表达上调有关.为了明确ROR α在DADS抑制人胶质瘤细胞增殖中的作用,本研究采用miRNA干扰技术抑制ROR α表达,观察其对DADS抑制U251细胞增殖的影响.方法:首先将ROR α miRNA转染人胶质瘤U251细胞,运用Western blot检测转染前后ROR α蛋白表达情况.实验分为未转染组、脂质体转染组和ROR α miRNA转染组及分别经30 mg/L DADS处理后的3组,共6组.MTT法检测ROR α表达下调对DADS抑制胶质瘤U251细胞增殖的影响.结果:Western b1ot结果显示,转染ROR α miRNA细胞的ROR α蛋白表达(0.09±0.05)明显低于未转染组(0.81±0.11)和脂质体转染组(0.89±0.15),其蛋白表达下调了89.5%(P＜0.05).MTT结果显示,U251细胞增殖活性在RORα miRNA转染后48hA570值为(0.98±0.15),高于未转染组(0.47±0.11)和脂质体转染组(0.45±0.10)(P＜0.05),其增殖率高达108.5%.并且ROR α miRNA转染削弱了DADS对U251细胞的增殖抑制作用,转染ROR α miRNA细胞加入DADS后的细胞增殖率(A570值为0.69±0.20)明显高于DADS处理的未转染组(0.28±0.13)和脂质体转染组(0.25±0.12)(P＜0.05),其抑制率由40.4%下降为29.6%.结论:miRNA干扰ROR α表达可促进U251细胞增殖,并且削弱了DADS对U251细胞的增殖抑制作用,说明ROR α参与了DADS抗胶质瘤U251细胞增殖的作用.     

miR-192在胶质瘤U251细胞中的表达及对其增殖、迁移及凋亡能力的影响

目的 探讨miR-192在胶质瘤U251细胞中的表达及对其增殖、迁移及凋亡等生物学能力的影响.方法 采用RT-PCR检测胶质瘤细胞株U251、LN18、U373和正常人脑胶质细胞株中HEB的表达水平.采用脂质体转染法将miR-192类似物(miR-192 mimics)和阴性对照(miR-negtive control...     

shRNA结合131I-免疫脂质体抑制胶质瘤细胞U251增殖的初步研究

目的 探索RNA干扰与131I-免疫脂质体联用在胶质瘤疾病治疗中的潜在价值.方法 在合成pGPU6-GFP-Neo-EGFRvⅢ-shRNA表达载体的基础上,结合核素内放射性131I免疫脂质体,进行细胞实验探索其对胶质瘤细胞U251增殖凋亡的影响.结果 成功构建pGPU6-GFP-Neo-EGFRvⅢ-shRNA载体;同时细胞实验结果显示,shRNA和131I免疫脂质体联用有效抑制胶质瘤细胞U251增殖和促进其凋亡,抑制率达70％.结论 shRNA结合131I免疫脂质体能高效地促凋亡和抑制细胞增殖,为其在胶质瘤治疗中的应用奠定了初步基础.     

长链非编码RNA MEG3靶向miR-21对胶质瘤U251细胞增殖、侵袭和迁移的影响

摘 要：［目的］ 探究长链非编码RNA人母系表达基因3 (maternally expressed gene 3,MEG3)对人胶质瘤细胞U251增殖、侵袭和迁移能力的影响及机制。［方法］ RT-PCR检测MEG3和miR-21在胶质瘤组织、癌旁组织中、正常星形胶质细胞NHAs和胶质瘤细胞U251中的表达;用pcDNA-MEG3 (pc-MEG3)转染U251细胞,RT-PCR检测MEG3和miR-21的表达;生物信息及荧光素酶报告实验预测并验证MEG3和miR-21的关系;MTT检测细胞增殖能力,Transwell和划痕实验检测细胞侵袭和迁移能力;免疫印迹检测增殖细胞核抗原 (proliferating cell nuclear antigen,PCNA)、基质金属蛋白酶-2(metalloproteinase-2,MMP-2) 和MMP-9的表达。［结果］ 与癌旁组织比较,MEG3在胶质瘤组织中表达水平明显降低(t=23.169,P<0.001),miR-21水平明显升高(t=14.965,P=0.002);与NHAs组比较,U251组细胞MEG3表达水平明显降低(t=13.145,P<0.001),miR-21表达水平显著升高（t=12.483,P<0.001);pcMEG3 能显著上调MEG3的表达水平并抑制miR-21表达(t=8.129,P<0.001;t=11.705,P<0.001);miR-21 mimic能显著促进miR-21表达并能降低MEG3 野生质粒 (MEG3 wt) 的活性(t=6.460,P<0.001;t=7.742,P=0.004);pc-MEG3能显著降低U251细胞增殖倍数和PCNA的表达水平(F=96.45,P<0.001;t=5.337,P<0.001),miR-21 mimic能显著减弱pc-MEG3对细胞增殖及PCNA表达的抑制作用(t=7.073,P<0.001;t=4.609,P<0.001);同时,pc-MEG3还能显著降低U251细胞的划痕闭合率和侵袭细胞数(t=5.014,P<0.001;t=10.664,P<0.001),并抑制MMP-2和MMP-9的表达(t=3.360,P=0.007;t=3.453,P=0.006）;miR-21 mimic能明显减弱pc-MEG3对细胞侵袭、迁移及MMP-2和MMP-9表达的抑制作用(t=2.498,P=0.032;t=4.298,P=0.002;t=4.612,P<0.001;t=5.137,P<0.001)。［结论］ MEG能通过靶向miR-21减弱胶质瘤U251细胞的增殖、侵袭和迁移能力。     

过表达 CD133 对脑胶质瘤U251细胞生物学行为的影响

目的： 构建稳定表达人 CD133 基因的脑胶质瘤U251细胞株,并探讨 CD133 对U251细胞生物学行为的影响。 方法： 将人 CD133 全长cDNA构建入逆转录病毒表达载体pEGZ-Term ,包装成逆转录病毒pEGZ-Term-CD133,进而感染脑胶质瘤U251细胞株。流式细胞术及Real-time PCR检测感染后U251细胞CD133分子的表达。细胞计数法、神经球形成实验观察CD133过表达对U251细胞体外的增殖和神经球形成的影响。裸鼠皮下成瘤法检测感染后U251细胞的体内致瘤性。 结果： 成功构建pEGZ-Term-CD133逆转录病毒表达载体,并获得稳定表达 CD133 的U251细胞。相比U251-mock、U251细胞,U251-CD133细胞高表达 CD133 mRNA \[（7 400.2±5 003.4) vs (2.0±1.1)、(1.0±2.2),均P=0.0007）和蛋白。感染pEGZ-Term-CD133对U251细胞的体外增殖并无影响（P＞0.05）;但在无血清神经干细胞培养条件下,U251-CD133细胞所形成的神经球数量显著高于U251-mock和U251细胞\[（34.0±7.5） vs（14.6±2.3）、（11.5±1.3）个,均P<0.01\]。接种量为1×105 个细胞时,U251-CD133细胞在裸鼠体内的成瘤时间（32 d）少于U251-mock细胞（38 d）、成瘤率更高（100% vs 30%）,在第41天时,肿瘤体积显著增大\[（180.3±146.8） vs （4.0±0.0）mm3,P=0.003\]。 结论： CD133分子不影响脑胶质瘤U251细胞的体外增殖,但可促进U251细胞神经球的形成和致瘤性。     

Inhibitor of cyclooxygenase-2 induces cell-cycle arrest in the epithelial cancer cell line via up-regulation of cyclin dependent kinase inhibitor p21

Cyclooxygenase-2 is the rate-limiting enzyme in synthesis of prostaglandins and other eicosanoids. Prior reports have shown that inhibition of cyclooxygenase-2 activity, either by selective inhibitors or by antisense oligonucleotide, results in suppression of growth of squamous cell carcinoma cell lines which express high cyclooxygenase-2 levels, such as NA, a cell line established from a squamous cell carcinoma of the tongue. To investigate the mechanisms by which cyclooxygenase-2 inhibitors suppressed growth of these cells, the effects of NS-398, the selective cyclooxygenase-2 inhibitor, on cell-cycle distribution were examined. NS-398 induced G0/G1 cell-cycle arrest in NA cells which expressed cyclooxygenase-2. G0/G1 arrest induced by NS-398 was accompanied by up-regulation of cyclin-dependent kinase inhibitor p21, but not by up-regulation of the other cyclin-dependent kinase inhibitors. Transfection with p21 antisense oligonucleotide inhibited cell-cycle arrest induced by NS-398. Accumulation in G0/G1 was also observed in NA cells transfected with cyclooxygenase-2 antisense oligonucleotide. On the other hand, NS-398-treated NA cells showed a loss of plasma membrane asymmetry, a marker of early events in apoptosis. However, NS-398 did not induce other morphological and biochemical changes related to apoptotic cell death. These results suggest that cyclooxygenase-2 inhibitor induces G0/G1 cell-cycle arrest in NA cells by up-regulation of p21. Our results also suggest that NS-398 is not sufficient to complete the whole process of apoptosis in NA cells, although it induces an early event in apoptosis.     

Inhibition of human vascular endothelial cells proliferation by terbinafine

We have demonstrated previously that terbinafine (TB), an oral antifungal agent used in the treatment of superficial mycosis, suppresses proliferation of various cultured human cancer cells in vitro and in vivo by inhibiting DNA synthesis and activating apoptosis. In our study, we further demonstrated that TB at a range of concentrations (0-120 microM) dose-dependently decreased cell number in cultured human umbilical vascular endothelial cells (HUVEC). Terbinafine was not cytotoxic at a concentration of 120 microM, indicating that it may have an inhibitory effect on the cell proliferation in HUVEC. The TB-induced inhibition of cell growth rate is reversible. [(3)H]thymidine incorporation revealed that TB reduced the [(3)H]thymidine incorporation into HUVEC during the S-phase of the cell-cycle. Western blot analysis demonstrated that the protein levels of cyclin A, but not cyclins B, D1, D3, E, CDK2 and CDK4, decreased after TB treatment. The TB-induced cell-cycle arrest in HUVEC occurred when the cyclin-dependent kinase 2 (CDK2) activity was inhibited just as the protein level of p21 was increased and cyclin A was decreased. Pretreatment of HUVEC with a p21 specific antisense oligonucleotide reversed the TB-induced inhibition of [(3)H]thymidine incorporation. Taken together, these results suggest an involvement of the p21-associated signaling pathway in the TB-induced antiproliferation in HUVEC. Capillary-like tube formation and chick embryo chorioallantoic membrane (CAM) assays further demonstrated the anti-angiogenic effect of TB. These findings demonstrate for the first time that TB can inhibit the angiogenesis.     

In vitro and in vivo studies of the anticancer action of terbinafine in human cancer cell lines: G0/G1 p53-associated cell cycle arrest

Terbinafine (TB) (Lamisil), a promising oral antifungal agent used worldwide, has been used in the treatment of superficial mycosis. In our study, we demonstrated that TB dose-dependently decreased cell number in various cultured human malignant cells. Flow cytometry analysis revealed that TB interrupts the cell cycle at the G0/G1 transition. The TB-induced cell cycle arrest in colon cancer cell line (COLO 205) occurred when the cyclin-dependent kinase (cdk) system was inhibited just as the levels of p53, p21/Cip1 and p27/Kip1 proteins were augmented. In the TB-treated COLO 205, the binding between p53 protein and p53 consensus binding site in p21/Cip1 promoter DNA probe was increased. Pretreatment of COLO 205 with p53-specific antisense oligodeoxynucleotide decreased the TB-induced elevations of p53 and p21/Cip1 proteins, which in turn led to arrest in the cell cycle at the G0/G1 phase. Moreover, in the p53 null cells, HL60, TB treatment did not induce cell cycle arrest. Taken together, these results suggest an involvement of the p53-associated signaling pathway in the TB-induced antiproliferation in COLO 205. We further examined whether administration of TB could affect the growth of tumors derived from human colon cancer cells in an in vivo setting. COLO 205 cells implanted subcutaneously in nude mice formed solid tumor; subsequent intraperitoneal injections of TB (50 mg/kg) led to obvious decline in tumor size, up to 50-60%. In these tumors, increases in the p21/Cip1, p27/Kip1 and p53 proteins and the occurrence of apoptosis were observed. Combined treatment with TB and nocodazole (ND), a clinically used anticancer agent, potentiated the apoptotic effect in COLO 205. These findings demonstrate for the first time that TB can inhibit the proliferation of tumor cells in vitro and in vivo.     

Inhibitory effects of evodiamine on the growth of human prostate cancer cell line LNCaP

Evodiamine, isolated from a Chinese herbal drug named Wu-Chu-Yu, possesses many biological functions. Recently, it has been reported that Wu-Chu-Yu exerts an antiproliferative effect on several cancers. Prostate carcinoma initially occurs as an androgen-dependent tumor and is the second leading cause of cancer death in American males. In the present study, the effect of evodiamine on the growth of androgen-dependent prostate cancer cell line LNCaP in vitro was examined. Based on [3-(4,5-dimethylthiazol-2-yle)2,5-diphenyltetrazolium bromide] (MTT) assay, evodiamine significantly inhibited the growth of LNCaP cells in a concentration-dependent manner. A significant and concentration-dependent inhibitory effect of evodiamine on LNCaP cell growth was observed at 24 hr and persisted for 96 hr. The examination of lactate dehydrogenase (LDH) assay showed that the cytotoxic effects of evodiamine on LNCaP cells were concentration dependent. Furthermore, we examined the influences of evodiamine on cell death and cell cycle. The flow cytometric analysis of evodiamine-treated cells indicated a block of G2/M phase and an elevated level of DNA fragmentation. The G2/M arrest reached a maximum at 24 hr after evodiamine treatment. The G2/M arrest was accompanied by an elevated p34(cdc2) kinase activity and an increase in the protein expression of cyclin B1 and phosphorylated form of p34(cdc2) (Thr 161). Examination of TUNEL showed that evodiamine-induced apoptosis was observed at 24 hr and extended for 72 hr. Evodiamine elevated caspase-3, and caspase-9 activities and the processing of caspase-3 and caspase-9. These results suggested that evodiamine inhibits the growth of prostate cancer cell line, LNCaP, through an accumulation of cell cycle at G2/M phase and an induction of apoptosis.     

Characteristics of antitumour activity of cepharanthin against a human adenosquamous cell carcinoma cell line

Cepharanthin is one of the biscoclaurine alkaloids widely used for treatment of many acute and chronic diseases; snakebite, bronchial asthma, alopecia areata, leukopenia during radiation therapy or anticancer treatment. Recently, it has been reported that cepharanthin exerts antitumour effects by increasing immunological competence of the host or apoptosis-inducing activity. In this study, we examined the antitumour effects of cepharanthin against a human adenosquamous cell carcinoma cell line (TYS). Treatment of TYS cells with cepharanthin (1020 μg/ml) resulted in a significant suppression of cell growth. Moreover, it was found by the flow cytometry analysis, nick end labelling or agarose gel electrophoresis, that G1 arrest and DNA fragmentation occurred in cepharanthin-treated cells. In addition, it was detected that induction of p21^WAF1 protein and activation of caspase 3 protype, which is one of Interleukin-1β converting enzyme (ICE) family proteases, were detected by Western blotting. The TYS tumour-bearing nude mice were treated with cepharanthin, which was administered subcutaneously (20 mg/kg/day). The cepharanthin treatment results in a significant suppression of tumour growth and an induction of apoptosis. These findings suggest that cepharanthin induces G1 arrest via expression of p21^WAF1 and apoptosis through caspase 3.     

吉非替尼对胃癌细胞株放射增敏的作用

背景与目的:表皮生长因子受体(epidermal growth factor receptor,EGFR)在绝大部分人类上皮肿瘤中都有表达,其表达高低与放疗抗拒有关.我们检测EGFR酪氨酸激酶抑制剂吉非替尼(gefitinib)对高表达胃癌细胞株放射增敏的作用,并初步探讨其机制.方法:Western blot法测定7株人胃癌细胞株(MKN45、SGC7901、SNU-1、N87、AGS、SNU-16、KATO-Ⅲ)中EGFR蛋白的表达,选取2株EGFR相对高表达的胃癌细胞用于后续实验.采用MTT法测定吉非替尼的半数抑制浓度(50%inhibition concentration,IC50),克隆形成实验计算细胞存活率,拟合生存曲线并计算放射生物学参数,流式细胞仪检测吉非替尼联合放疗的凋亡率及细胞周期分布.结果:选取7株胃癌细胞中EGFR表达最高的MKN45和SGC7901细胞,发现其存活率均随吉非替尼浓度或放射剂量的增加而明显下降(P＜0.05).MTT法检测吉非替尼对MKN45及SGC7901细胞的IC50分别为0.4mmol/L和0.8 mmol/L.MKN45细胞在0.1×IC50及0.2×IC50剂量下的增敏比(SER)分别为1.102和1.154,SGC7901则为1.092和1.176.吉非替尼或照射均可增加凋亡率,减少S期细胞比例及增加G2/M期细胞比例(P＜0.01).结论:吉非替尼序贯照射应用可提高EGFR高表达胃癌细胞的放射敏感性并阻碍细胞增殖、促进凋亡和干扰细胞周期分布.吉非替尼有望成为EGFR高表达胃癌的放射增敏剂.     

粉防己碱对人肝癌细胞株7402的放射增敏作用

目的:观察粉防己碱(Tet)对人肝癌细胞株7402的放射增敏作用。方法:以人肝癌细胞株7402为研究对象,应用MTT比色法、克隆形成实验检测Tet的细胞抑制效应;应用克隆形成实验观察Tet对7402细胞株的放射敏感性的影响;流式细胞术测定照射后7402细胞株细胞周期的再分布并观察Tet能否去除放射引起的细胞周期阻滞;Western blot检测CyclinB1、Cdc2和Cdc25C磷酸化形式的表达水平;细胞分裂指数实验观察Tet对照射后细胞分裂指数的影响。结果:不同浓度的Tet作用于人肝癌7402细胞株24h后,其细胞毒性呈剂量依赖性。Tet(0.5μg/ml)能明显降低放射后7402细胞的克隆形成率,其放射增敏比(SERDq)为1.76。流式细胞术结果显示,照射明显导致7402细胞株G2期阻滞,Tet能够去除放射引起的7402细胞G2期阻滞。Western blot显示细胞在受到X射线照射后,CyclinB1表达水平明显降低,Cdc2和Cdc25磷酸化形式的表达水平明显增加,分裂指数也明显降低;经Tet处理后,CyclinB1表达水平明显增高,Cdc2和Cdc25磷酸化形式的表达水平明显下降,分裂指数则增高。结论:Tet对7402细胞株有放射增敏作用,其作用机制可能与Tet去除放射引起的G2/M期阻滞有关。     

Molecular mechanisms of G0/G1 cell-cycle arrest and apoptosis induced by terfenadine in human cancer cells

Terfenadine (TF), a highly potent histamine H1 receptor antagonist, has been shown to exert no significant central nervous system side effects in clinically effective doses. In this study, we demonstrated that TF induced significant growth inhibition of human cancer cells, including Hep G2, HT 29, and COLO 205 cells, through induction of G(0)/G(1) phase cell-cycle arrest. The minimal dose of TF induced significant G(0)/G(1) arrest in these cells was 1-3 microM. The protein levels of p53, p21/Cip1, and p27/Kip1 were significantly elevated, whereas the kinase activities of cyclin-dependent kinase 2 (CDK2) and CDK4 were inhibited simultaneously in the TF-treated cells. On the other hand, significant apoptosis, but not G(0)/G(1) arrest, was induced in the HL 60 (p53-null) or Hep 3B (with deleted p53) cells when treated with TF (3-5 microM). To clarify the roles of p21/Cip1 and p27/Kip1 protein expression, which was involved in G(0)/G(1) arrest and apoptosis induced by TF in human cancer cells, antisense oligodeoxynucleotides (ODNs) specific to p21/Cip1 and p27/Kip1 were used, and the expression of the p21/Cip1 and p27/Kip1 were monitored by immunoblotting analysis. Our data demonstrated that the percentage of the apoptotic cells detected by annexin V/PI analysis in the TF-treated group was clearly attenuated by pretreatment with p27/Kip1-specific ODNs. These results indicated that p27/Kip1 (but not p21/Cip1) protein indeed played a critical role in the TF-induced apoptosis. We also demonstrated that the TF-induced G(0)/G(1) cell-cycle arrest effect was not reversed by TF removal, and this growth inhibition lasted for at least 7 d. Importantly, the occurrence of apoptosis and cell growth arrest was not observed in the TF-treated normal human fibroblast, even at a dose as high as 25 microM. Our study showed the molecular mechanisms for TF-induced cell growth inhibition and the occurrence of apoptosis in human cancer cells.