川芎嗪对IL-1β诱导的兔原代软骨细胞增殖和凋亡影响

目的 观察川芎嗪对白细胞介素-1β诱导的软骨细胞的增殖及凋亡的影响.方法 兔原代软骨细胞培养及鉴定,用IL-1β10ng/ml和/或不同浓度川芎嗪共培养兔原代软骨细胞48h后,利用流式细胞仪检测各组软骨细胞的周期及凋亡率;利用MTT法检测软骨细胞的生长状态.结果 与对照组比较,IL-1β诱导下软骨细胞的凋亡率显著增加,差异具有显著统计学意义(P＜0.01);加入川芎嗪能明显降低IL-1β诱导的软骨细胞凋亡率,有显著统计学意义(P＜0.01);与对照组比较,IL-1β诱导下软骨细胞被明显阻滞在G1期(P＜0.01);加入川芎嗪能明显降低IL-1β对软骨细胞G1期的阻滞作用,细胞增殖指数明显增加(P＜0.05或P＜0.01).结论 川芎嗪对IL-1β诱导的兔原代软骨细胞抑制凋亡并促进生长.     

新型手性抗肿瘤药物德氮吡格（TNBG）体外抗肿瘤活性初步研究

目的观察手性旋光异构体德氮吡格（TNBG）对6株人体肿瘤细胞增殖的影响,初步探讨其抗肿瘤作用机制。方法运用MTT法检测手性TNBG对6株肿瘤细胞生长抑制情况;油红O染色法观察QGY-7701细胞中脂质聚集情况;流式细胞仪检测手性TNBG对人肝癌细胞株QGY-7701细胞周期分布和凋亡影响。结果手性TNBG能不同程度抑制肿瘤细胞增殖,且呈剂量依赖性;油红O染色提取显示QGY-7701细胞内脂质量与给药剂量呈正相关;流式细胞仪检测到有s期细胞增加,并且手性TNBG还能诱导QGY-7701细胞发生凋亡。结论手性TNBG在体外具有良好的抗肿瘤活性,其抗肿瘤作用机制可能是通过促使肿瘤细胞产生脂质聚集,抑制肿瘤细胞增殖、诱导其凋亡从而达到抗肿瘤效应。总体看（＋）TNBG体外抗肿瘤活性比（-）TNBG更强。     

枸杞多糖对人肺腺癌 A549细胞增殖和凋亡的影响

目的探讨枸杞多糖(LBP)对体外培养的人肺腺癌细胞 A549的增殖抑制作用及其可能的作用机制.方法用不同浓度的LBP处理 A549细胞,MTT法检测24、48、72h时间点 LBP对 A549细胞的生长抑制率,实验设为对照组和实验组(1/2IC50作用48小时),MTT法绘制生长曲线、细胞计数计算倍增时间、流式细胞仪检测凋亡率及其细胞周期、RT PCR检测 SurvivinmRNA的变化、Westernblot检测 CyclinB1蛋白的变化,transwell体外侵袭实验观察药物对细胞体外侵袭的影响.结果 MTT显示不同浓度的LBP均能明显抑制 A549细胞的增殖且成剂量 效应关系,实验组细胞的倍增时间、凋亡率与对照组相比,均有统计学意义(P<0.05);LBP使细胞阻滞在 G2期,SurvivinmRNA表达和 CyclinB1蛋白的表达均降低,与对照组相比差异有显著性(P<0.05).结论 LBP可抑制 A549细胞的增殖,其机理可能与 LBP使 SurvivinmRNA表达下降引起细胞凋亡及 CyclinB1蛋白的表达降低造成细胞周期阻滞及抑制细胞的侵袭能力有关     

EGCG通过TGF-β1信号通路抑制PLA-802细胞的机制研究

目的 观察表没食子儿茶素-3-没食子酸酯(EGCG)对人横纹肌肉瘤细胞株PLA-802的抑制作用,以及对细胞内TGF-β1/Smad4的表达的影响,探索EGCG抑制横纹肌肉瘤细胞生长的机制.方法 体外培养人横纹肌肉瘤细胞株PLA-802,并用不同浓度的EGCG作用不同的时间.用MTT检测EGCG对PLA-802细胞生长情况的影响,用流式细胞术检测细胞周期的变化情况,RT-PCR和Westernblot分别检测细胞内TGF-β1和Smad4的mRNA和蛋白水平的表达.结果 MTT结果显示EGCG显著降低了PLA-802细胞的存活率((P＜0.05).流式细胞结果表明EGCG明显降低了S期而增加了G1期(P＜0.05).而TGF-β1和其下游因子Smad4的mRNA和蛋白水平的表达也明显受到EGCG的抑制,且这种抑制作用呈浓度-时间依赖性(P＜0.05).结论 EGCG发挥其抑制PLA-802细胞的作用可通过抑制TGF-β1信号通路,这或许将为临床治疗横纹肌肉瘤提供新的思路.     

青蒿水提液对肺癌A549细胞的影响

目的研究青蒿水提液对肺癌A549细胞株增殖的影响和诱导凋亡的情况。方法不同浓度青蒿水提液作用于细胞不同时间,四甲基氮噻唑蓝（MTT）法检测吸光度值（A490nm）并计算增殖抑制率;AnnexinV-FITC／PI荧光染色后流式细胞仪检测细胞凋亡率;并以荧光显微镜观察细胞形态改变情况;蛋白质印迹法分析细胞凋亡相关蛋白Bax、Bcl-2的表这。结果青蒿水提液呈时间和剂量依赖性抑制A549细胞增殖;荧光显微镜下A549细胞出现不同时期凋亡特征性改变;流式细胞仪检测细胞凋亡率随着药物浓度增加而升高;A549细胞株的Bax蛋白表达量增多、Bcl-2蛋白表达量下降。结论青蒿水提液促进体外培养的A549细胞株增殖抑制并诱导凋亡,其机制可能与A549细胞Bax表达上调和Bcl-2表达下调有关。     

依那普利对高糖诱导的大鼠肾小球系膜细胞增殖及分泌TGF-1β、LN的影响

目的 观察依那普利（Ena）对高糖诱导的大鼠肾小球系膜细胞（GMCs）增殖及分泌转化生长因子-β1（TGF-β1）和层粘连蛋白（LN）的影响.方法 大鼠GMCs分别培养于正常浓度葡萄糖（NG,5.5mmol/L）、高浓度葡萄糖（HG,20mmol/L）及高糖和不同浓度（10-7mol/L、10-6mol/L、10-5mol/L）的Ena中,同时设空白对照.分别采用四甲基偶氮唑蓝（MTT）法和酶联免疫吸附试验（ELISA）、放射免疫法（RIA）检测不同时间（24 h、48 h、72 h）GMCs增殖情况和细胞培养上清液中TGF-β1、LN的含量.结果 与空白对照组比较,正常糖组和高糖组GMCs增殖增加,TGF-β1、LN分泌增多（P〈0.001）,高糖组更明显（P〈0.001）.与高糖组比较,高、中、低浓度的Ena干预均可逆转上述变化.且在一定浓度范围内,Ena无明显细胞毒性.结论 高糖可致GMCs增殖和TGF-β1、LN分泌增多,Ena可抑制高糖诱导的GMCs增殖与分泌TGF-β1、LN,这可能是其减少细胞外基质（ECM）积聚的机理之一,并有利于延缓糖尿病肾病肾小球硬化（GS）的进展.     

Curcumin通过Wnt信号通路抑制乳腺癌细胞MCF-7增殖的研究

目的 观察姜黄素(Curcumin)对乳腺癌细胞MCF-7增殖的影响,以及对细胞内Wnt信号通路的影响,探索Curcumin可能存在的抑制乳腺癌细胞增殖的分子机制.方法 体外培养人乳腺癌细胞MCF-7,并用不同浓度的Curcumin作用不同的时间.用MTT检测Curcumin对MCF-7细胞生长情况的影响;流式细胞仪观察经Curcumin作用后细胞周期的改变;RT-PCR和Westernblot分别检测细胞内β -catenin和下游靶基因CyclinD1的mRNA和蛋白水平的表达.结果 MTT结果显示Curcumin可以抑制MCF-7的增殖,并具有剂量-时间依赖性.在浓度为20 μmol·L-1时,对细胞生长的抑制作用最为明显.流式细胞仪观察细胞周期的结果提示,Curcumin能够阻止MCF-7细胞由G1期进入S期,提高Go/G1期细胞的百分比.RT-PCR和Western blot结果显示,Curcumin显著降低了细胞内β-catenin和CyclinD1的mRNA和蛋白水平的表达,且呈剂量-时间依赖性.结论 Curcumin能够抑制MCF-7细胞胞浆内β -catenin蛋白进入胞核,阻断Wnt信号转导通路.进而抑制下游靶基因CyclinD1的表达,阻止MCF-7由G1期进入S期,有效抑制了MCF-7细胞的增殖.     

胃癌侧群细胞耐药性研究及干细胞相关基因检测

目的研究胃癌侧群（Side Population,sP）细胞对化疗药物5-Fu（氟尿嘧啶）的耐药性及可能机制,并检测干细胞相关基因Nanog、Musashi-1及cD44的表达情况。方法选择人胃癌细胞株sGc-7901,以荧光染料H0echst 33342染色,维拉帕米桔抗对照,应用流式细胞仪分选sP细胞和nonsP细胞。细胞耐药实验比较sP细胞与nonsP细胞对化疗药物5．Fu的耐药性差异;westem_h10t检测ABcG2和bcl-2蛋白表达情况;流式细胞仪分析细胞周期;荧光定量PcR检测两组细胞中干细胞相关基因Nanog、Musashi-1及cD44mRNA的表达差异。结果胃癌细胞株sGc．790l中sP细胞的比例为2．8％,sP细胞对5-Fu的耐药存活率明显高于non-sP细胞（P〈0．05）,与nonsP细胞相比,sP细胞高表达耐药蛋白ABcG2和抗凋亡蛋白bcl-2,有更多的细胞处于c0／G1期（P〈0．05）,并高表达干细胞相关基因Musashi-1和cD44。结论胃癌sGC_7901细胞株中sP细胞对化疗药物5．Fu的耐药性明显高于nonsP细胞,其耐药机制可能与sP细胞高表达耐药蛋白ABcG2和抗凋亡蛋白bcl-2,有更多细胞处于G0／Gl期有关;Musashi-1和cD44可能是相对特异性的胃癌干细胞标志物。     

齐墩果酸抑制肝癌细胞QGY增殖和诱导凋亡的作用研究

目的研究齐墩果酸对人肝癌细胞QGY增殖的作用及与细胞内钙离子浓度（[Ca2＋]i）关系。方法将浓度分别为40、80、100μg／ml齐墩果酸作用肝癌H细胞QGY24h后,DAPI染色,以荧光显微镜观察细胞形态变化;以11组不同浓度齐墩果酸（5—400μg／ml）作用QGY细胞24h后,用四甲基偶氮唑蓝（Myr）法检测QGY增殖情况;分别以不同浓度齐墩果酸（80、100、120μg／ml）作用QGY细胞24h后,流式细胞仪检测细胞周期改变、细胞凋亡率和[Ca2＋]i。结果细胞增殖被抑制并发生凋亡：不同浓度齐墩果酸能够抑制QGY细胞株增殖,且在5—120μg／mL范围内呈剂量依赖性,药物作用细胞24h、48的Ic50分别为76．27μg／mL和66．56μg／mL;处理组细胞周期在s期产生阻滞、细胞内[Ca2＋]i较对照组显著增加,细胞凋亡率和[Ca2＋]i与药物浓度存依赖关系。结论齐墩果酸能够抑制肝癌细胞QGY增殖和诱导其凋亡;诱导凋亡可能与细胞内[Ca2＋]i增加有关。     

环磷酰胺抑瘤作用及含药血清对肿瘤细胞凋亡水平的影响

目的:采用小鼠腋下接种瘤株的方法,建立肝癌荷瘤小鼠动物模型;利用血清药理学的方法,观察环磷酰胺(CTX)舍药血清体外的抑制肿瘤细胞生长及调亡作用的研究.方法 :小鼠肝癌细胞混悬液用生理盐水按1:1进行稀释制成含瘤腹水混悬液,在给药前24 h,每只小鼠腋窝皮下接种0.2 ml.CTX对小鼠肝癌抑瘤实验连续给药10 d,测定肿瘤作用端粒酶活性与细胞凋亡表达.结果:(1)CTX能够明显抑制肝癌肿瘤的生长(P＜0.01);(2)30%、20%和10%含药血清高中低剂量含药血清对HepG2肿瘤细胞具有明显的抑瘤作用,Hochest染色肿瘤细胞出现凋亡形态;而流式细胞仪检测可见含药血清高中低剂量的促肿瘤凋亡率,明显高于正常血清组.CTX能够明显升高肝癌荷瘤小鼠体内bc1-2的水平(P＜0.01);(3)CTX能够明显促进肝癌细胞的凋亡.结论 :CTX具有抑制小鼠肝癌的作用;能够明显促进肝癌细胞的凋亡;CTX的抑瘤作用可能与升高动物体内的bcl-2水平及促进癌细胞的凋亡有关,从而表现抗肿瘤的作用.     

解聚复肾宁对糖尿病肾病大鼠TNF—OL、MCP-1的影响

目的观察解聚复肾宁（Jiejufushenning,JJFSN）对糖尿病（Diabetes mellitus,DM）大鼠肾组织肿瘤坏死因子-α（Tumorneerosisfactor-alpha,TNF-α）、单核细胞趋化蛋白-1（Monocyte chemoattractant protein-1,MCP-1）和血清TNF-α的影响,探讨其肾保护作用机制。方法腹腔注射链脲佐菌素（Streptozotocin,STZ）建立SD大鼠DM模型,将成模DM大鼠随机分为3组：DM模型组（B组）、JJFSN组（C组）、厄贝沙坦（Irbesartan）组（D组）,另设正常对照组（A组）。采用相应干预措施处理12周。常规方法测定12周末各组大鼠空腹血糖（FBG）、血尿素氮（BUN）、血肌酐（Scr）、肾重／体重（KW／BW）、24h尿蛋白（24huFro）;放免法测血清TNF-α含量：免疫组织化学方法测肾组织TNF—Ot、MCP-1的表达;PAS染色评估细胞外基质;电镜观察肾组织超微结构改变。结果与A组相比．B组大鼠FBG、BUN、Scr、KW／BW、24huPro及血清TNF-α升高（P〈0．05）,肾组织TNF—Ot、MCP-1表达明显增高（P〈0．05）;肾组织超微结构明显异常;C组和D组上述指标显著改善（P〈0．05）,肾组织超微结构异常改善。结论竹FSN能延缓DM大鼠肾损害进程,其机制可能与抑制TNF-α、MCP-1上调有关。     

Divergent effects of the major mast cell products histamine, tryptase and TNF-alpha on human fibroblast behaviour

Fibroblast proliferation is a key process in tissue remodeling and mast cells (MCs) are thought to play a crucial role. Having established that the three major MC products, tryptase, histamine and TNF-alpha (TNF) are normally present in human skin MCs, which are in close proximity to dermal fibroblasts, we studied their individual effects on cell cycle-controlled human dermal fibroblasts (HFFF2). These cells express receptors (H1, PAR2, TNFR1/2) for the major MC mediators, but only tryptase or a PAR2 agonist peptide stimulated proliferation and gene expression. TNF was antimitotic, and histamine, while elevating intracellular Ca²⁺ levels at high concentrations, did not affect proliferation. We conclude that MC products but also composition and numbers of respective receptors on fibroblasts are crucially responsible for fibroproliferative events. Received: 28 June 2005; received after revision 28 September 2005; accepted 6 October 2005     

Regulation of G1 phase progression by growth factors and the extracellular matrix

Cell cycle progression is regulated by both intracellular and extracellular control mechanisms. Intracellular controls ensure that cell cycle progression is stopped in response to irregularities such as DNA damage or faulty spindle assembly, whereas extracellular factors may determine cell fate such as differentiation, proliferation or programmed cell death (apoptosis). When extracellular factors bind to receptors at the outside of the cell, signal transduction cascades are activated inside the cell that eventually lead to cellular responses. We have shown previously that MAP kinase (MAPK), one of the proteins involved in several signal transduction processes, is phosphorylated early after mitosis and translocates to the nucleus around the restriction point. The activation of MAPK is independent of cell attachment, but does require the presence of growth factors. Moreover, it appears that in Chinese hamster ovary cells, a transformed cell line, growth factors must be present early in the G1 phase for a nuclear translocation of MAPK and subsequent DNA replication to occur. When growth factors are withdrawn from the medium immediately after mitosis, MAPK is not phosphorylated, cell cycle progression is stopped and cells appear to enter a quiescent state, which may lead to apoptosis. Furthermore, in addition to this growth-factor-regulated decision point in early G1 phase, another growth-factor-sensitive period can be distinguished at the end of the G1 phase. This period is suggested to correlate with the classical restriction point (R) and may be related to cell differentiation.     

解聚复肾宁对糖尿病大鼠肾脏HGF表达的影响

目的观察解聚复肾宁（JJFSN）对糖尿病大鼠肾HGF表达的影响及肾保护作用。方法SD大鼠腹腔注射链尿佐菌素法建立糖尿病模型,随机分为正常对照组、模型组、解聚复肾宁治疗组、依那普利治疗组、JJFSN＋依那普利治疗组。12周后,检测各组空腹血糖（FBG）、尿素（BU）、24h尿β2微球蛋白（β2-MG）、肌酐清除率（Ccr）;免疫组化法检测肾组织HGF表达,放射免疫法测定大鼠血清胰岛素（Ins）含量,透射电镜观察肾脏超微结构。结果与模型组比较,各治疗组大鼠FBG、BU、β2-MG明显降低,Ccr明显升高,血清Ins含量增加,肾组织HGF表达明显上调,肾组织病理变化减轻,其中以J＋Y组改善最显著（P〈0．05）。结论JJFSN对DM大鼠肾脏形态争功能有明显保护作用,其机制可能与其上调肾脏HGF表达、保护胰岛β细胞、延缓肾脏纤维化进程有关。     

Cell cycle progression by the repression of primary cilia formation in proliferating cells

In most cell types, primary cilia protrude from the cell surface and act as major hubs for cell signaling, cell differentiation, and cell polarity. With the exception of some cells ciliated during cell proliferation, most cells begin to disassemble their primary cilia at cell cycle re-entry. Although the role of primary cilia disassembly on cell cycle progression is still under debate, recent data have emerged to support the idea that primary cilia exert influence on cell cycle progression. In this review, we emphasize a non-mitotic role of Aurora-A not only in the ciliary resorption at cell cycle re-entry but also in continuous suppression of cilia regeneration during cell proliferation. We also summarize recent new findings indicating that forced induction/suppression of primary cilia can affect cell cycle progression, in particular the transition from G0/G1 to S phase. In addition, we speculate how (de)ciliation affects cell cycle progression.     

Culture in low levels of oxygen enhances in vitro proliferation potential of satellite cells from old skeletal muscles

The proliferation ability of satellite cells (considered the 'stem cells' of mature myofibers) declines with increasing age when cultured under standard cell culture conditions of 21% oxygen. However, actual oxygen levels in the intact myofiber in vivo are an order of magnitude lower. No studies to date have addressed the issue of whether culturing satellite cells from old muscles under more 'physiologic' conditions would enhance their proliferation and/or differentiation ability. Therefore, we analyzed satellite cells derived from 31-month-old rats in standard cultures with 21% O₂ and in lowered (∼3%) O₂. Under the lowered O₂ conditions, we noted a remarkable increase in the percentage of large-sized colonies, activation of cell cycle progression factors, phosphorylation of Akt, and downregulation of the cell cycle inhibitor p27^Kip1. These data suggest that lower O₂ levels provide a milieu that stimulates proliferation by allowing continued cell cycle progression, to result ultimately in the enhanced in vitro replicative life span of the old satellite cells. Such a method therefore provides an improved means for the ex vivo generation of progenitor satellite cell populations for potential therapeutic stem cell transplantation. Received 20 April 2001; received after revision 28 May 2001; accepted 31 May 2001     

Regulation of the cell cycle following DNA damage in normal and Ataxia telangiectasia cells

A proportion of the population is exposed to acute doses of ionizing radiation through medical treatment or occupational accidents, with little knowledge of the immedate effects. At the cellular level, ionizing radiation leads to the activation of a genetic program which enables the cell to increase its chances of survival and to minimize detrimental manifestations of radiation damage. Cytotoxic stress due to ionizing radiation causes genetic instability, alterations in the cell cycle, apoptosis, or necrosis. Alterations in the G1, S and G2 phases of the cell cycle coincide with improved survival and genome stability. The main cellular factors which are activated by DNA damage and interfere with the cell cycle controls are: p53, delaying the transition through the G1-S boundary; p21^WAF1/CIPI, preventing the entrance into S-phase; proliferating cell nuclear antigen (PCNA) and replication protein A (RPA), blocking DNA replication; and the p53 variant protein p53as together with the retinoblastoma protein (Rb), with less defined functions during the G2 phase of the cell cycle. By comparing a variety of radioresistant cell lines derived from radiosensitive ataxia talangiectasia cells with the parental cells, some essential mechanisms that allow cells to gain radioresistance have been identified. The results so far emphasise the importance of an adequate delay in the transition from G2 to M and the inhibition of DNA replication in the regulation of the cell cycle after exposure to ionizing radiation.