角质细胞生长因子对囊肿衬里上皮细胞细胞周期及其调控蛋白表达的影响

目的:研究角质细胞生长因子(KGF)对常染色体显性多囊肾病(ADPKD)囊肿衬里上皮细胞细胞周期及其调控蛋白的影响,以初步探讨ADPKD囊肿发生、发展的细胞学机制。 方法:应用MTT法检测KGF对ADPKD囊肿衬里上皮细胞的促增殖作用;应用流式细胞仪观察KGF对ADPKD囊肿衬里上皮细胞细胞周期的影响;应用免疫组化结合病理图文检测KGF对囊肿衬里上皮细胞细胞周期调控蛋白Cyclin D1、P21 waf1表达影响。 结果:①KGF能明显促进ADPKD囊肿衬里上皮细胞增殖,影响细胞周期,使G0~G1期细胞减少,S期细胞增多;②体外培养的ADPKD囊肿衬里上皮细胞存在Cyclin D1和 P21waf1蛋白,经图文分析, 50 ng/ml KGF刺激后,Cyclin D1蛋白表达(0.41±0.04)较对照组(0.30±0.01)明显增强,P21waf1的表达(0.32±0.02)较对照组(0.37±0.03)明显减弱(P<0.01)。 结论:Cyclin D1和 P21waf1是ADPKD囊肿衬里上皮细胞细胞周期G1期进展的调控蛋白;KGF可能通过刺激Cyclin D1蛋白的表达,抑制P21waf1蛋白的产生,从而进一步促进细胞通过G1~S调控点,致使ADPKD囊肿衬里上皮细胞明显增殖。     

肾上腺髓质素对大鼠肝星状细胞表达细胞周期蛋白的影响

目的探讨肾上腺髓质素(ADM)对活化肝星状细胞(HSC)表达细胞周期蛋白的影响。方法采用不同浓度的ADM对HSC-T6细胞进行干预12h、24h和48h。采用MTT法检测HSC-T6细胞增殖;采用SABC法检测细胞增殖核抗原(PCNA)、细胞周期蛋白D1(cyclin D1)和细胞周期素依赖性激酶抑制因子P21waf1的表达。结果在ADM10-7mol/L浓度下作用12h,24h,48h,细胞生长抑制率逐渐增加(分别是7.88%,17.58%,18.79%);在ADM10-7mol/L作用24h时,HSC的PCNA、cyclin D1和P21waf1表达的平均光密度分别为0.387±0.023、0.454±0.029和0.508±0.011,而对照组分别为0.246±0.030、0.209±0.014和0.401±0.033(t值分别为5.149、5.752和5.299,P<0.05),差异有显著性。相关性分析表明,PCNA和cyclinD1的变化呈正相关趋势(r=0.834,P<0.05),而PCNA和P21waf1呈负相关趋势(r=-0.770,P<0.05),cyclinD1和P21waf1呈负相关趋势(r=-0.796,P<0.05)。结论肾上腺髓质素可抑制HSC-T6细胞增殖。ADM作用HSC可能通过调控HSC的细胞周期,抑制PCNA和cyclinD1表达,促进P21waf1表达,从而发挥抑制HSC增殖作用。     

腺病毒介导的野生型PTEN抑制活化肝星状细胞系HSC-T6增殖的机制

目的 通过检测腺病毒介导的野生型第10号染色体缺失的磷酸酶张力蛋白同源物基因(PTEN)对体外活化肝星状细胞(HSC)细胞周期的影响,探讨过表达的野生型PTEN抑制活化HSC增殖的可能机制. 方法 体外培养肝星状细胞系HSC-T6,以腺病毒为载体将野生型PTEN基因瞬时转染活化HSC,实验分为空白对照组、Ad-GFP组和Ad-PTEN组.流式细胞术测定HSC细胞周期时相,Western blot及实时定量PCR检测HSC的PTEN表达量,Western blot检测HSC的细胞周期素D1 (cyclin D1)和细胞周期素依赖性激酶4(CDK4)表达水平.多组间比较采用单因素方差分析,组间比较采用Tukey检验. 结果 携带野生型PTEN基因的腺病毒(AD-PTEN)成功转染HSC,Ad-PTEN组G0/G1期HSC细胞数(67.68％±2.75％)较对照组(53.01％±2.37％)及Ad-GFP组(53.85％±3.0％)明显增高(F=38.59,P＜0.01);S期HSC细胞数(14.42％±1.81％)较对照组(22.17％±1.99％)及Ad-GFP组(21.54％±1.74％)明显降低(F=18.85,P＜ 0.01);G2/M期HSC细胞数(17.90％±2.70％)较对照组(24.82％±3.81％)及Ad-GFP组(24.62％±3.15％)明显降低(F=7.17,P＜0.01).腺病毒感染HSC后72h,Ad-PTEN组cyclin D1的相对表达量(1.12±0.07)较对照组(1.45±0.05)及Ad-GFP组(1.47±0.08)明显降低(F=42.86,P＜0.01),CDK4相对表达量(1.05±0.07)较对照组(1.41±0.03)及Ad-GFP组(1.43±0.06)也明显降低(F=67.01,P＜ 0.01).结论 过表达的野生型PTEN通过下凋cyclin D1和CDK4的表达,明显抑制体外活化HSC细胞周期的G1/S期转化,阻滞HSC细胞周期时相于G0/G1期,进而抑制其增殖.     

PHB1对人肝癌细胞增殖的影响及其作用机制

目的探讨抗增殖蛋白1(prohibitin1,PHB1)对人肝癌细胞增殖的影响及其作用机制。方法构建PHB1真核表达重组质粒(pEGFP-PHB1)和PHB1靶向siRNA质粒(shRNA-PHB1)转染人肝癌细胞HepG2和SMMC-7721,诱导PHB1在人肝癌细胞中表达上调和下调,采用MTT、流式细胞学、荧光定量PCR和免疫印迹等技术检测人肝癌细胞增殖、细胞周期,以及细胞周期关键调控分子的表达情况。结果高表达PHB1可阻滞HepG2和SMMC-7721细胞于G0/G1期[(67.28±2.94)%比(56.71±2.56)%,t=6.64,P=0.00;(69.48±3.82)%比(60.43±2.59)%,t=4.80,P=0.00],使S期比例下降[(14.74±1.45)%比(24.13±1.92)%,t=9.54,P=0.00;(13.73±1.26)%比(25.50±2.30)%,t=10.99,P=0.00],抑制细胞增殖;周期调控分子p53和p21CIP1mRNA和蛋白质水平显著升高,而Cyclin A2、Cyclin E1和CDK2 mRNA和蛋白质水平显著降低(P0.01)。低表达PHB1可促使HepG2和SMMC-7721细胞趋于S期[(31.65±2.71)%比(24.68±1.28)%,t=5.69,P=0.00;(31.02±2.49)%比(25.88±2.40)%,t=3.64,P=0.005],促进细胞增殖;p53、p21CIP1、Cyclin A2、Cyclin E1、CDK2 mRNA和蛋白质水平与PHB1高表达时相反(P0.01)。结论高表达PHB1可以阻滞人肝癌细胞周期于G0/G1期,进而抑制细胞增殖,其作用机制可能与p53介导的G0/G1期相关细胞周期蛋白有关。     

过氧化物酶体增殖物活化受体γ配体15d-PGJ2对肝星状细胞增殖活化的影响

目的观察过氧化物酶体增殖物活化受体γ(PPARγ)的天然配体15d-PGJ2对HSC增殖及活化的影响,以探讨PPARγ在HSC活化过程中的作用。方法采用MTT法和RT-PCR方法观察5μmol/L及10μmol/L 15d-PGJ2对体外培养的HSC自发活化及血小板衍生生长因子(PDGF)引起的HSC增殖及活化的影响。结果以5μmol/L 15d-PGJ2处理原代HSC 3 d后,可明显抑制HSC活化标志物α-平滑肌肌动蛋白的表达,而PPARγ的表达较未处理组明显增高(0.64±0.03对比0.09±0.01,t=36.0517,P<0.01);15d-PGJ2可剂量依赖性地抑制PDGF引起的HSC增殖;经5μmol/L和10μmol/L 15d-PGJ2预处理后再用PDGF干预,则PPARγ的表达较单用PDGF干预组明显增高(分别为0.03±0.02对比0.60±0.03,t=42.6616,P<0.01;以及0.03±0.02对比0.69±0.04,t=33.83,P<0.01),而HSC的活化指标α-平滑肌肌动蛋白、α1(I)型胶原及单核细胞趋化蛋白-1的表达则受抑制。结论激活PPARγ可调控HSC的促纤维化和促炎症作用,促进PPARγ的表达可能成为抗肝纤维化的新手段。     

粉防己碱对大鼠静止期肝星状细胞细胞周期的影响

目的观察粉防己碱对大鼠静止期肝星状细胞（HSCs）细胞周期的影响及机制。方法分离正常大鼠HSCs,培养48 h后给予粉防己碱（1.6μmol/L）和（或）TGF-β1（5μg/L）处理3 d。流式细胞仪检测细胞周期分布;应用RT-PCR法检测Cyclin D1、Cyc-lin E、p21WAF1/CiP1和p27Kip1表达;使用Western blot法检测Cyclin D1和PPARγ蛋白水平。结果粉防己碱使G0/G1期和G2/M期细胞增多,S期细胞数显著减少,抑制TGF-β1介导的G0/G1期细胞含量下调和S期细胞含量升高,上调p21WAF1/CiP1mRNA表达,抑制Cyclin D1而维持PPARγ蛋白在一定水平。结论在静止期HSC中,粉防己碱通过抑制Cyclin D1、上调p21WAF1/CiP1和维持PPARγ表达使HSCs出现G0/G1期停滞和S期向G2/M期转换加速。     

Hedgehog-Gli 1信号通路对肝星状细胞增殖和凋亡的调控作用

目的 研究肝星状细胞(HSC)中hedgehog(Hh)信号通路成员Shh、patched、smoothened(Smo)和Gli-1的表达,及Hh通路抑制剂环耙明对HSC-T6细胞株增殖、凋亡及其激活的影响.方法 体外培养HSC株,提取细胞总RNA,用RT-PCR扩增Shh、patched、Smo、Gli-1基因;用环耙明作用HSC-T6后采用四甲基偶氮唑盐法检测其在体外对HSC-T6的抑制情况,流式细胞仪检测其对HSC-T6细胞周期的影响,碘化丙啶和膜联蛋白-Ⅴ双染荧光标记法及提取细胞DNA检测HSC-T6细胞凋亡,荧光定量聚合酶链反应检测Gli-1,转化生长因子β1、血小板衍生生长因子、Bcl-2 mRNA的表达水平.结果 HSC-T6中表达有Hh通路家族成员.环耙明在50、100、150、200,250 μmol/L作用下,A值分别为1.55±0.07、1.19 ±0.05,0.78±0.06、0.48±0.03、0.38±0.04,正常对照组为1.74±0.03,环耙明组与正常对照组相比,F=636.81,P<0.01,差异有统计学意义.流式细胞术测出干预后环耙明G0/G1期细胞所占比例为65.08%±1.50%,正常对照组为55.41%±2.54%,两组比较,t=-8.05,P<0.01.药物干预后提取DNA及碘化丙啶和膜联蛋白-Ⅴ双染荧光标记均检测出HSC-T6细胞凋亡.荧光定量结果表明,经环耙明干预后,HSC-T6细胞中Gli-1、转化生长因子β1、血小板衍生生长因子、Bcl-2 mRNA的目的基因相对表达量分别0.28±0.05、0.13±0.04、0.07±0.04、0.17±0.02,正常对照组为1,各基因表达量较正常对照组均明显降低,t值分别为23.09、31.34、43.87和59.10,P值均<0.01,差异有统计学意义.结论 HSC-T6中存在Hh信号通路,环耙明能抑制HSC增殖及增殖周期,促进活化的HSC凋亡,其抑制HSC活化可能是通过抑制Hh-Gli-1信号通路的结果.     

姜黄素对人肝星状细胞中神经生长因子及其低亲和力受体表达的影响

目的观察姜黄素对人肝星状细胞(HSC)中神经生长因子(NGF)及其低亲和力受体P75NTR表达的影响,探讨姜黄素逆转肝纤维化的可能机制。方法体外培养人HSC株LX-2,分别给予0~80μmol/L不同浓度的姜黄素作用于LX-2,MTT法检测细胞增殖;选取30~50μmol/L姜黄素分别作用于LX-2,细胞免疫化学染色检测NGF和P75NTR表达,计算阳性细胞率。结果 10~20μmol/L姜黄素作用下LX-2增殖与对照组相比,差异无统计学意义(P0.05);在30~80μmol/L浓度范围内,LX-2增殖显著降低(P0.05);在30~50μmol/L浓度范围内,NGF蛋白表达率分别为(46.59±8.15)%、(90.75±4.72)%、(83.32±4.78)%,对照组为(27.11±5.50)%;P75NTR为(37.17±4.71)%、(88.74±6.27)%、(83.79±4.70)%,对照组为(24.33±5.17)%,差异均有统计学意义(P0.01)。结论姜黄素可诱导人HSC中NGF和P75NTR的蛋白表达,这可能是姜黄素逆转肝纤维化的机制之一。     

姜黄素对血管平滑肌细胞增殖及cyclinD1和p21waf1/cip1蛋白表达的影响

目的 观察姜黄素（curcumin）抑制大鼠血管平滑肌细胞(VSMC)增殖和诱导细胞周期停滞的作用,以及对细胞周期蛋白cyclinD1,p21waf1/cip1表达的影响。方法 采用组织贴块法体外培养大鼠VSMC,MTT法测定姜黄素对VSMC增殖的抑制作用,流式细胞仪分析细胞周期分布,Western印迹法检测cyclinD1,p21waf1/cip1蛋白的变化。结果 MTT示不同浓度姜黄素(7.5～120 μmol/L)在24～72 h范围内,浓度和时间依赖性抑制VSMC增殖;30 μmol/L以上浓度姜黄素显著抑制增殖的VSMC细胞周期进程,使S及G2/M期减少(P<0.05),G0/G1期增加(P<0.05);30 μmol/L的姜黄素可抑制cyclinD1表达,促进p21waf1/cip1蛋白表达。结论 姜黄素具有明确的抑制VSMC增殖和细胞周期停滞的作用,其与cyclinD1,p21waf1/cip1蛋白变化有关。     

姜黄素对肝星状细胞增殖与凋亡的影响

目的　观察姜黄素对体外培养肝星状细胞 (HSC)增殖与凋亡的影响。方法　不同浓度姜黄素处理HSC株HSC T6 ,MTT法检测细胞增殖 ,流式细胞仪、透射电镜和琼脂糖凝胶电泳法检测细胞凋亡。结果　在 2 0～ 10 0 μmol/L浓度范围内 ,姜黄素可剂量依赖性地抑制HSC增殖 (P <0 .0 1)。 2 0、4 0、6 0 μmol/L姜黄素处理HSC 2 4h后 ,细胞周期分析发现S期细胞减少 ,G2 /M期细胞显著增加 (P <0 .0 1) ;流式细胞术检测到明显的亚G1峰 ,各组的凋亡指数 (% )分别是 15 .3± 1.9,2 6 .7± 2 .8,37.6± 4 .4 ,与对照组 (1.9± 0 .6 )相比 ,差异有显著性 (P <0 .0 1) ;4 0 μmol/L姜黄素作用 12、2 4、36、4 8h ,凋亡指数 (% )分别是 12 .0± 2 .4、2 6 .7± 3.5、33.8± 1.8和 4 9.3± 1.6 ,与对照组相比 ,差异有显著性 (P <0 .0 1) ;透射电镜观察到细胞皱缩、核染色质浓缩沿核膜排列和凋亡小体形成等 ;琼脂糖凝胶电泳可见到明显的DNA梯带形成。结论　姜黄素可显著抑制HSC增殖 ,使细胞周期停滞于G2 /M期 ,并诱导其凋亡 ,其作用具有时间和剂量依赖性     

细胞外信号调节激酶信号通路对乙醛刺激的大鼠肝星状细胞周期的影响

目的 观察特异性丝裂原细胞外信号反应激酶1(MEK 1)阻断剂(PD98059)对乙醛刺激的大鼠肝星状细胞(HSC)增殖及细胞周期的影响,并探讨其作用机制。 方法 用不同浓度的PD98059对乙醛刺激的HSC进行处理;以四甲基偶氮唑蓝法检测细胞增殖,流式细胞仪检测细胞周期,逆转录聚合酶链反应方法检测HSC内细胞周期蛋白-D1(Cyclin D1)mRNA和细胞周期蛋白依赖性激酶(CDK4)mRNA的表达。 结果 20、50、100μmol/L的PD98059均能显著且剂量依赖性地抑制乙醛刺激的HSC增殖,3组A值分别为0.109±0.020、0.081±0.010、0.056±0.020,与乙醛组A值0.146±0.030相比较,F=31.385,P<0.05;20、50、100 μmol/L的PD98059可显著抑制乙醛刺激的HSC由G1期进入S期,G0/G1期细胞百分比逐渐升高,3组G0/G1期细胞百分比分别为(61.9±6.3)％、(64.1±3.3)％、(70.9±4.8)％,与乙醛组(55.2±4.4)％相比较,F=16.402,P<0.05;50、100μmol/L的PD98059能显著抑制乙醛刺激的HSC内Cylin D1 mRNA表达,2组平均光强度比值分别为0.56±0.04,0.46±0.03,与乙醛组0.65±0.07相比较,F=68.758,P<0.05;50、100μmol/L的PD98059能显著抑制乙醛刺激的HSC内CDK4 mRNA表达,2组平均光强度比值分别为0.39±0.07,0.33±0.05,与乙醛组0.50±0.06相比较,F=29.406,P<0.05。 结     

Blocking effects of genistein on cell proliferation and possible mechanism in human gastric carcinoma

AIM: To study the blocking effects of genistein on cell proliferation cycle in human gastric carcinoma cells (SGC-7901) and the possible mechanism. METHODS: MTT assay was applied in the detection of the inhibitory effects of genistein on cell proliferation. Flow cytometry was used to analyze the cell cycle distribution. Immunocytochemical technique and Western blotting were performed to detect the protein expression of cyclin D_1, cyclin B_1 and p21~(waf1/cip1). RESULTS: Genistein significantly inhibited the growth and proliferation of human gastric carcinoma cells (SGC-7901). Seven days after treatment with different concentrations of genistein (2.5, 5.0, 10.0, 20.0 μg/mL), the growth inhibitory rates were 11.2%, 28.8%, 55.3%, 84.7% respectively and cell cycles were arrested at the G(2)/ M phase. Genistein decreased cyclin D_1 protein expression and enhanced cyclin B_1 and p21~(waf1/cip1) protein expression in a concentration-dependent manner. CONCLUSION: The growth and proliferation of SGC-7901 cells can be inhibited by genistein via blocking the cell cycle, with reduced expression of cyclin D_1 and enhanced expression of cyclin B_1 and p21~(waf1/cip1) protein in the concentration range of 0-20 μg/mL.     

雷帕霉素对高糖诱导肾小球系膜细胞增殖的影响

目的 探讨雷帕霉素对高糖诱导肾小球系膜细胞（GMC）增殖的影响及其可能涉及的途径。方法高糖培养GMC,以不同剂量雷帕霉素干预,MTT及流式细胞仪等方法观察雷帕霉素对细胞增殖和周期的影响,RT-PCR、Western印迹观察细胞Cyclin D1、CyclinE和p27^KIP1的变化。结果高糖刺激下,GMC增殖明显;雷帕霉素能抑制这一作用,且呈剂量依赖性,并下调CyclinD1、CyclinE的基因及蛋白水平,上调p27^KIP1的蛋白表达;与对照组相比,高糖组G0／G1期细胞减少,S期细胞比例增加（P均＜0．05）;雷帕霉素干预后,G0／G1期细胞升高,S期细胞比例减少（P均＜0．05）。结论雷帕霉素可抑制高糖状态下GMC的增殖,且呈剂量依赖性;其可能机制是通过下调CyclinD1、CyclinE及上调p27^KIP1,参与了G1／S期阻滞。     

汉防己甲素抑制PANC-1细胞增殖作用的机制探讨

目的：探讨汉防己甲素对人胰腺癌细胞株PANC-1细胞增殖的影响及其相关机制。方法：采用MTT法观察汉防己甲素对人胰腺癌细胞株PANC-1细胞增殖的影响;流式细胞术检测其对PANC-1细胞周期及凋亡的影响;实时定量PCR检测细胞周期和凋亡相关基因P21^cip/waf1、cdc25A、c-Myc、E2F1和survivin mRNA水平的变化;Western blot检测P21^cip/waf1蛋白水平的改变。结果：MTT结果显示汉防己甲素对PANC-1细胞增殖抑制作用呈明显的剂量和时间依赖性。经汉防己甲素处理后,流式细胞术检测到明显的G1期细胞阻滞和细胞凋亡。实时定量PCR结果显示,汉防己甲素处理早期,P21^cip/waf1mRNA水平明显增加,而cdc25A、c-Myc、2F1、survivin mRNA水平均下降。Western blot结果表明P21^cip/waf1蛋白水平在汉防己甲素处理早期升高。结论：我们的研究表明汉防己甲素能有效的抑制人胰腺癌细胞株PANC-1细胞增殖。该过程可能通过上调21^cip/waf1mRNA和蛋白的表达,下调cdc25A、c-Myc、E2F1mRNA的表达,致使PANC-1细胞G1期阻滞;也可能通过下调survivin mRNA的表达,诱导PANC-1细胞凋亡。     

Effects of the tyrosine protein kinase inhibitor genistein on the proliferation,activation of cultured rat hepatic stellate cells

AIM：Hepatic stellate cell(HSC)plays a pivotal role in liver fibrosis and is considered as the therapeutic target for the treatment of hepatic fibrosis,Tyrosine protein kinase plays an important role in the proliferation,activation of HSC.The purpose of the study is to investigate the effects of the tyosine protein kinase inhibitor genistein on the proliferation and activation of cultured rat HSC.METHODS:Rat HSC were isolated from Wistar rats by in situ perfusion of collagenase and pronase and single-step density Nycodenz gradient,Culture-activated HSC were serum-starved and incubated with10^-9to10^-5mol/L concentration of genistein for 24,48or 72h,In PDGF-induced HSC proliferation,HSC were stimulated with10μg&#183;L^-1PDGF-BBfo15min,and thentreated with genistein for the same time.Cell proliferation was measured by MПassay and based on flow cytometric analysis of cell cycle.The a-smooth muscle actin(α-SMA）expression in HSC was studied with confocallaser microscopy and flow cytometry.c-fos,c-jun and cyclinD1expression in HSCwas also detected by flow cytometry.RESULTS:Genistein inhibited basal and PDGF-induced proliferation of HSCat the concentration of 10^-8to10^-5mol/L,and treatment with10^-7mol/L concentration of genistein for 48h inhibited the HSCproliferation significantly(the inhibition rate was 70.3%,P&lt;0.05).Immunofluorescence detected by confocal laser microscopy and flow cytometry showed that treatment with10^-7mol/L genistein for48h suppressed the expression of α-SMA significantly in HSC(the specific fluorescence intensity were60.2&#177;21.5vs35.3&#177;11.6and12.8&#177;10.4vs9.54&#177;6.39,respectively,bothP&lt;0.05).The intensity of c-fos,c-jun and cyclinD1 expression of HSCs treated with 10^-7mol/L genistein for 48h was also significantly decreased compared with the controls.CONCLUSION;Genistein influences proliferation of HSC,suppresses the expression of α-SMA in HSC and tinhibits the intensity of c-fos,c-jun and cyclinD1 expression of HCSs,Genistein has therapeutic potential against liver fibrosis.     

Effects of tachyplesin on the regulation of cell cycle in human hepatocarcinoma SMMC-7721 cells

AIM: To investigate the effects of tachyplesin on the cell cycle regulation in human hepatcarcinoma cells. METHODS: Effects of tachyplesin on the cell cycle in human hepatocarcinoma SMMC-7721 cells were assayed with flow cytometry. The protein levels of p53, p16, cyclin D1 and CDK4 were assayed by immunocytochemistry. The mRNA levels of p21(WAF1/CIP1) and c-myc genes were examined with in situ hybridization assay. RESULTS: After tachyplesin treatment, the cell cycle arrested at G0/G1 phase, the protein levels of mutant p53, cyclin D1 and CDK4 and the mRNA level of c-myc gene were decreased, whereas the levels of p16 protein and p21(WAF1/CIP1) mRNA increased. CONCLUSION: Tachyplesin might arrest the cell at G0/G1 phase by upregulating the levels of p16 protein and p21(WAF1/CIP1) mRNA and downregulating the levels of mutant p53, cyclin D1 and CDK4 proteins and c-myc mRNA, and induce the differentiation of human hepatocacinoma cells.     

Overexpression of cyclin D2 in chronic B-cell malignancies

Tumor progression in B-cell chronic lymphocytic leukemia (B-CLL) is thought to result from the gradual accumulation of small resting G0/G1 phase lymphoid cells rather than the proliferation of actively dividing cells. The recent identification of G1 cyclins that are likely to control both the progression through G0 and G1 phase and the G1/S transition prompted us to study the mRNA expression of D-type cyclins in the peripheral blood lymphocytes from 34 patients with B-CLL, 7 patients with lymphoplasmacytic lymphoma (LPL), and 2 patients with mantle cell lymphoma (MCL). Cyclin D2 mRNA was, on average, 5- to 10- fold overexpressed in most of the samples studied (B-CLL, 29/34; LPL, 7/7; MCL, 0/2) as compared with normal resting B lymphocytes, in which cyclin D2 mRNA was barely detectable. In situ hybridization with cyclin D2 digoxigenin-labeled mRNA probe showed that all the cells from a given sample were stained with approximately the same intensity. Cyclin D3 was never detected in any of the samples tested, whereas cyclin D1 was expressed in only the 3 cases (1 LPL and 2 MCL) bearing a t(11;14) translocation. A trisomy 12 was found in 4 of 19 (21%) B-CLL or LPL cases for which cytogenetic analysis was available. Although the cyclin D2 gene has been mapped to chromosome 12p13, there was no apparent correlation between trisomy 12 and the level of cyclin D2 expression. Cell cycle analysis by flow cytometry after staining with propidium iodide consistently showed that more than 96% of the cells were in G0/G1 phase, whatever the importance of cyclin D2 overexpression was, and that cyclin D2 overexpression in B-CLL was not associated with any modifications of the cell cycle repartition. No consistent overexpression of cyclin D2 was found in acute myeloid leukemias. In conclusion, overexpression of cyclin D2 mRNA was found to be an almost constant feature in B-CLL and LPL. Therefore, it led us to hypothesize, with the support of data from some transfection experiments previously reported in murine hematopoietic cell lines, that cyclin D2 might play a role in B-CLL pathogenesis, possibly by preventing cells from programmed cell death.