增生性瘢痕退行性变过程中的细胞凋亡和相关基因表达

增生性瘢痕发生的重要机制之一是成纤维细胞不能进入正常的程序性死亡,导致其异常增殖使细胞外基质过度沉积。半胱氨酸天冬氨酸蛋白酶(caspases)的活化形式过度表达可诱导细胞凋亡。本实验对增生性瘢痕退行性变的不同时期采用原位缺口末端标记(TUNEL)法作细胞凋亡检测,探讨细胞凋亡及caspase-3在该过程中的作用机制。     

硫利达嗪对胃癌细胞生长的抑制作用与机制研究

目的:探讨硫利达嗪对胃癌细胞体外生长的抑制作用及其机制。方法:不同浓度的硫利达嗪作用于胃癌SGC-7901细胞24 h后,分别用MTT法与流式细胞仪检测胃癌细胞的增殖与凋亡情况,以及用Western blot检测凋亡相关蛋白bax、bcl-2、caspase-3的表达情况。结果:硫利达嗪作用后,SGC-7901细胞的增殖明显抑制,凋亡率明显增加,抗凋亡蛋白bcl-2表达下调、促凋亡蛋白bax表达上调、caspcase-3蛋白表达上调,且均呈浓度依赖性(均P0.05)。结论:硫利达嗪对人胃癌细胞体外的生长有明显抑制作用,该作用可能与其并活化caspase-3依赖的凋亡途径有关。     

肾必宁冲剂加S-9对肾小球系膜细胞凋亡及其相关因素的影响

目的：探讨肾必宁冲剂对肾小球系膜细胞凋亡及凋亡相关基因ICE和BcI-2表达的影响，旨在阐明其治疗系膜增生性肾病有效的作用机制，为临床利用该药治疗以系膜增生为核心病理环节的肾脏疾病提供实验理论依据。方法：采用肾小球系膜细胞培养方法进行体外 研究，有不同剂量与S－9共同孵育的肾必宁冲剂及含肾必宁大鼠血清刺激4－6代系细胞16h,MTT法测定系膜细胞增殖指数；TUNEL法定量观察凋亡细胞；琼脂糖凝胶电泳法观察细胞核小体DNA的断裂现象，逆转录－聚合酶链反应技术（RT－PCR）检测凋亡相关基因ICE和bcI-2的表达变化。结果：加入肾必宁冲剂（S－9）16h后，系膜细胞出现了典型的凋亡征象，诸如细胞内深棕色颗粒形成、核固缩、碎裂和DNA梯形条带等，凋亡率显高于空白对照组（P＜0．01）；同时肾必宁各组ICE表达亦明显高于空白血清对照组，而抑制凋亡的bcI-2基因各组均表达不明显。结论：肾必宁冲剂可诱导系膜细胞凋亡，促进肾小球增殖性病变消散，并可通过调控ICE和bcI-2基因影响凋亡。     

天冬氨酸特异性半胱氨酸蛋白酶活化在喜树碱诱导膀胱癌细胞凋亡中的作用

目的：探讨天冬氨酸特异性半胱氨酸蛋白酶(Caspase)活化,在DNA拓扑异构酶Ⅰ抑制剂喜树碱(CPT)诱导人膀胱癌细胞凋亡中的作用。方法：将体外培养的膀胱癌RT4和MGH细胞用CPT处理,用流式细胞仪检测处理前及处理后24、36、48h等不同时段细胞凋亡的变化,用蛋白免疫印迹方法检测CPT处理后不同时间段caspase-9和caspase-3活性的改变。结果：CPT处理膀胱RT4和MGH细胞后,细胞凋亡率明显上升,且CPT诱导的膀胱癌凋亡呈时间依赖性和剂量依赖性。同时还可以观察到凋亡执行蛋白caspase-9和caspase-3的活化。结论：CPT能够诱导膀胱癌细胞RT4和MGH发生凋亡．其中caspase-9启动的凋亡内源性途径即线粒体途径可能参与了CPT诱导的膀胱癌细胞的凋亡。     

水飞蓟素诱导高转移表型肺癌细胞系(Anip973)细胞凋亡的分子机制研究

目的 探讨水飞蓟素诱导高转移表型肺癌细胞系Anip973细胞凋亡的分子机制.方法 采用四甲基偶氮唑蓝(MTT)比色法、倒置显微镜和电子显微镜等形态学检测、流式细胞仪(FCM)技术检测、DNA Ladder分析、PARP的表达分析以及凋亡相关蛋白Bax、Bcl-2、caspase-3和caspase-9表达活性分析.结果 水飞蓟素对人高转移表型肺癌细胞系Anip973细胞的增殖有显著抑制作用.随着浓度的增加,倒置显微镜下可见细胞数目减少,胞体变小、变圆,到高浓度时出现较多的死亡细胞.扫描电镜观察发现,凋亡细胞表现出典型的超微结构特征.流式细胞仪检测显示G1期细胞比例增多,S期细胞明显减少,G2期细胞略有减少,并出现明显的凋亡峰.水飞蓟素作用后的Anip973细胞出现明显的DNA Lad-der和PARP降解增加等凋亡特征.凋亡相关蛋白Bax表达增加、caspase-3和caspase-9酶活性增加,而Bd-2表达降低.结论 水飞蓟素可以在体外抑制人肺腺癌细胞Anip973的增殖作用,并通过激活线粒体依赖的caspase凋亡通路,诱导其凋亡.     

柴苓小方对大鼠肾小球系膜细胞异常增殖的影响

目的：探讨柴苓小方对大鼠肾小球系膜细胞（GMCs）异常增殖的影响及相关机制。方法：采用脂多糖刺激系膜细胞异常增殖。MTT法检测柴苓小方对㈣异常增殖的影响；免疫细胞化学方法检测药物作用后增殖细胞核抗原（PCNA）的表达；乳酸脱氢酶释放实验检测柴苓小方的细胞毒作用。采用RT—PCR检测药物作用不同时间点细胞周期蛋白依赖性激酶抑制剂p21的表达。结果：柴苓小方可以剂量依赖性的抑制系膜细胞异常增殖并显著减少PCNA阳性细胞数而没有明显的细胞毒作用。并且柴苓小方可以显著地上调p21基因的表达（P〈0．05）。结论：柴苓小方可剂量依赖性的抑制GMC8异常增殖．其机制可能与上调p21mRNA表达有关。     

青蒿琥酯对脂多糖诱导的大鼠肾小球系膜细胞凋亡及Bcl-2基因表达的影响

目的：观察青蒿琥酯（Art）对脂多糖（LPS）诱导的大鼠肾小球系膜细胞（glomerular mesangial cells,GMC）凋亡及Bcl-2基因表达的影响,并探讨其可能的作用机制。方法：用不同浓度的青蒿琥酯刺激脂多糖诱导增殖的大鼠肾小球系膜细胞,HE染色观察凋亡细胞形态,DAPI荧光染色观察细胞核凋亡情况,用流式细胞仪检测细胞凋亡率,免疫细胞化学结构和计算机图文分析系统检测Bcl-2的表达。结果：青蒿琥酯对系膜细胞凋亡有明显诱导作用,呈一定剂量依赖性;并能明显地抑制脂多糖诱导上调的Bcl-2基因的表达,且高浓度组的抑制作用强于低浓度组。结论：青蒿琥酯能够剂量依赖性的诱导大鼠系膜细胞凋亡,其机制可能与调控Bcl-2的表达有关。     

还氧合酶-2抑制剂NS398对人前列腺癌细胞系PC-3的增殖抑制作用

目的 观察还氧合酶-2(COX-2)选择性抑制剂NS398对人前列腺癌细胞系PC-3的增殖抑制作用，探讨NS398的抗癌机理。方法 逆转录-聚合酶链反应(RT—PCR)用于检测PC-3的COX-2mRNA的表达情况。采用四甲基偶氮唑蓝(MTT)法观察NS398对PC-3的增殖抑制作用。为进一步探讨NS398对PC-3的增殖抑制机制，通过DNA梯形电泳和透射电镜试验对细胞凋亡的情况进行检测。结果 PC-3的COX-2mRNA表达阳性。NS398对PC-3的增殖抑制作用表现为剂量依赖性和时间依赖性。DNA梯形电泳显示特征性的梯形谱带。透射电镜表现为细胞核膜完整，染色质聚集成块、边集，胞浆内可见明显的“空泡”和“凋亡小体”，细胞膜完整，细胞表面的微绒毛消失。结论 NS398对人前列腺癌细胞系PC-3的增殖具有抑制作用，并能诱导PC-3细胞的凋亡。NS398对PC-3的抑制可能是通过诱导细胞凋亡的途径实现的。     

牛磺酸对系膜细胞增殖及凋亡的影响

目的 观察牛磺酸对系膜细胞增殖及凋亡的影响，并探讨其作用的可能机制。方法 用噻唑蓝MTT法检测细胞增殖，流式细胞仪检测细胞周期及凋亡，吖啶噔活体染色观察细胞凋亡形态，免疫细胞化学结合计算机图文分析系统检测cjn,cfos表达。结果 在5－50mmol/L浓度范围内牛磺酸能剂量依赖性地抑制系膜细胞增殖，可使G0－G1期细胞增多，S期细胞减少，并能明显抑制c-jun,c-fos的表达（P＜0．01），而牛磺酸并不诱导系膜细胞的凋亡。结论 牛磺酸可显著抑制系膜细胞增殖，使系膜细胞阻滞于G0－G1期；牛磺酸对系膜细胞增殖的抑制作用与其抑制c-jun,c-fos的表达有关。牛磺酸对系膜细胞凋亡无诱导作用。     

多沙唑嗪诱导前列腺癌PC-3细胞凋亡机制的实验研究

目的探讨多沙唑嗪(doxazosin)对雄激素非依赖性人前列腺癌PC-3细胞株增殖与凋亡的影响及其机制。方法用MTT法测定不同浓度多沙唑嗪对PC-3细胞生长抑制情况,用荧光显微镜、流式细胞仪观察多沙唑嗪对细胞凋亡的诱导作用,Western-Blot检测caspase-3,caspase-8,caspase-9、FADD和Bid、Bax、Bcl-2的表达水平。流式细胞仪检测不同时段Caspase-9抑制剂对细胞凋亡率的影响。结果多沙唑嗪可显著抑制PC-3细胞的体外生长,其25,50,100μmol/L浓度组的A值与对照组比较差异有统计学意义(P<0.01)。PC-3细胞中FADD、caspase-3,-9,-8和Bid、Bax表达显著增加,Bcl-2无明显变化。PC-3细胞凋亡可显著被caspase-9抑制剂抑制。结论多沙唑嗪能诱导PC-3细胞凋亡,其作用可能通过死亡信号途径激活caspase-8,同时Bid,Bax表达上调,引发线粒体途径激活caspase-9和caspase-3而实现的。     

容量敏感外向整流性氯通道参与氧化应激介导的系膜细胞凋亡

目的 探讨容量敏感外向整流性(VSOR)氯通道在H2O2介导系膜细胞凋亡中的作用和可能的机制。 方法 全细胞膜片钳技术用于检测VSOR氯电流。细胞凋亡通过吖啶橙/溴化乙锭荧光染色、电子显微镜、TUNEL染色和半胱氨酸天冬氨酸蛋白酶(caspase)-3活性确定。 结果 150 μmol/L H2O2激活系膜细胞VSOR氯电流，H2O2激活的氯电流具有典型的VSOR氯电流电生理特性，包括：外向整流性、电压依赖性失活。对细胞外高渗透压敏感及被VSOR氯通道阻断剂抑制。VSOR氯通道阻断剂DIDS(100 μmol/L)，NPPB(10 μmol/L)和尼氟灭酸(10 μmol/L)明显抑制H2O2介导的系膜细胞凋亡。150 μmol/L H2O2处理2 h内，细胞容量明显下降，但这种细胞容量下降被100 μmol/L DIDS，10 μmol/L NPPB和10 μmol/L尼氟灭酸抑制。结论 VSOR氯通道参与H2O2介导的系膜细胞凋亡，其机制与介导凋亡性容量下降有关。     

Effective component from verbena officinalis L.inhibits proliferation and induces apoptosis of human choriocarcinoma JAR cells

<正> Objective:To examine the action of the effective component,4'-methylether-scutellarein,from Verbena officinalis L.(VOL)on the proliferation and apoptosis of human choriocarcinomaJAR cells.Methods:Cell proliferation was measured by MTT[3-(4,5-dimethylthiazol-2-yl)-2,5-di-phenyl tetrasodium bromide,MTT]assay and the incorporation of tritiated thymidine(~3H-TdR).Apoptosis of cell was evaluated by flow cytometry(FCM)and the characteristic apoptoticDNA ladder by agarose gel electrophoresis,and the morphological changes of apoptotic JAR cellswere observed under fluorescence microscopy and electron microscopy(EM).Expressions of ap-optosis proteins,poly(ADP-ribose)polymerase(PARP)and caspase-3,-8,and -9 were deter-mined with Western blot.Results:The effective component from VOL inhibited the proliferation of JAR cells in a dose-and time-dependent manner.The treated cell cycle was arrested in S phase and an apoptotic peakwas found in S phase using FCM analysis.A typical DNA ladder appeared in the treatment groupwhen analyzed by agarose gel electrophoresis.Using fluorescence microscopy,the percentage ofapoptotic cell was 0.9%,6%,and 14% after treatments of 10,20,and 40 mg·L~(-1) of the effec-tive component,respectively,for 48 h.Typical apoptotic changes,such as condensed chromatinand presence of apoptotic bodies,were observed under EM.Treatment with effective componentfor 48 h and 72 h also induced protein expression of PARP and caspase-3,-8,and -9 as seen byWestern blot.Conclusions:The effective component from VOL inhibits cell proliferation and induces apop-tosis in human choriocarcinoma JAR cells.     

辛伐他汀通过PI3K/Akt通路缓解肺泡Ⅱ型细胞缺氧复氧损伤

目的 观察辛伐他汀( SIM)对缺氧复氧损伤肺泡Ⅱ型细胞(ATⅡ)的保护作用并探讨其机制.方法 体外培养人ATⅡ来源的A549细胞株,建立二氯化钴(CoCl2)诱导的缺氧复氧损伤模型;不同浓度辛伐他汀(5～100 μmol/L)处理A549细胞,CCK-8比色法检测细胞增殖率;Hoechst33342染色检测细胞凋亡;Western blot检测ATⅡ特征标志表面蛋白C(SP-C)及PI3K/Akt通路蛋白Akt、P70、mTOR、Caspase-3水平.结果 与对照组比较,缺氧前予低剂量SIM(5～20 μmol/L)预处理30 min,可显著促进A549细胞增殖、抑制其凋亡,并上调SP-C、p-Akt及下游增殖相关蛋白p-P70、mTOR水平,同时下调凋亡蛋白Caspase-3水平,两者差异有统计学意义(P＜0.01).与SIM组比较,给予PI3K抑制剂渥曼青霉素(wortmannin)可拮抗SIM的保护作用,表现为SP-C、p-Akt、mTOR及p-P70蛋白水平下调,Caspase-3蛋白水平上调,两者差异有统计学意义(P＜0.01).结论 辛伐他汀可缓解ATⅡ细胞缺氧复氧损伤,其机制与辛伐他汀活化PI3K/Akt通路有关.     

脂氧合酶抑制剂对乳腺癌细胞MCF-7增殖和凋亡的影响

目的 探讨脂氧合酶抑制剂去甲二氢愈创木酸（NDGA）对乳腺癌细胞MCF-7增殖和凋亡的影响。方法 噻唑蓝（MTT）法绘制细胞生长曲线及检测细胞存活率；流式细胞仪检测细胞周期和凋亡率；倒置相差显微镜和扫描电镜观察细胞凋亡。结果 MTT法显示NDGA对MCF-7细胞生长增殖有明显的抑制作用（P〈0.05），呈剂量和时间依赖效应；流式细胞仪结果显示NDGA阻滞细胞周期于S期，诱导细胞凋亡；在光镜和扫描电镜下，NDGA诱导细胞发生凋亡形态学变化，导致细胞表面超微结构的改变和凋亡小体的形成。结论 NDGA能够抑制MCF-7细胞的增殖，诱导细胞凋亡，为脂氧合酶抑制剂临床应用于乳腺癌提供了科学依据。     

矢车菊素-3-葡萄糖苷诱导人胃癌细胞株SGC7901凋亡及其机制

目的 观察矢车菊素-3-葡萄糖苷(C3G)在体外诱导人胃癌SGC7901细胞凋亡的生物学效应,并初步探讨其分子机制.方法 分别采用不同浓度梯度的C3G处理SGC7901胃癌细胞,MTT比色法检测细胞生长率;激光共聚焦显微镜观察细胞形态改变;TUNEL法分析细胞凋亡率;Western blot法检测凋亡相关基因Bcl-2、Bax、Caspase-3、ICAD蛋白表达的情况.结果 C3G在体外能明显抑制人胃癌SGC7901细胞的生长增殖,且呈浓度、时间依赖性(P＜0.01).激光共聚焦显微镜下可见Hoechst33258荧光染色细胞呈凋亡特征性改变;TUNEL法检测实验组细胞凋亡率均明显高于阴性对照组(P＜0.01);C3G可下调Bcl-2蛋白表达,上调Bax蛋白表达,降低Bcl-2/Bax蛋白的比值,促使Caspase-3酶原蛋白活化,下调ICAD蛋白表达(P＜0.01).结论 C3G具有在体外抑制胃癌细胞增殖的作用,并能诱导其凋亡,其机制可能与Bcl-2/Bax降低导致Caspase-3活化、ICAD蛋白表达下降相关.     

Methylglyoxal induces apoptosis through activation of p38 mitogen-activated protein kinase in rat mesangial cells

BACKGROUND: The formation of methylglyoxal (MG), a highly reactive dicarbonyl compound, is accelerated through several pathways, including the glycation reaction under diabetic conditions, presumably contributing to tissue injury in diabetes. On the other hand, apoptotic cell death of glomerular cells has been suggested to play a role in the development of glomerulosclerosis in various types of glomerular injuries. We therefore examined whether MG was capable of inducing apoptosis in rat mesangial cells to address the possible mechanism by which hyperglycemia-related products accelerated pathologic changes in diabetic glomerulosclerosis. METHODS: Rat mesangial cells were incubated with 0 to 400 micromol/L MG, followed by the detection of apoptosis by both TUNEL method and electrophoretic analysis for DNA fragmentation. In addition, we investigated intracellular mechanisms mediating MG-induced apoptosis, focusing especially on the p38 mitogen-activated protein kinase (MAPK) pathway. RESULTS: MG induced apoptosis in rat mesangial cells in a dose-dependent manner and was accompanied by the activation of p38alpha isoform. Aminoguanidine and N-acetyl-l-cysteine inhibited the MG-induced p38 MAPK activation, as well as apoptosis in rat mesangial cells, suggesting the involvement of oxidative stress in these phenomena. SB203580, a specific inhibitor of p38 MAPK also suppressed the MG-induced apoptosis in rat mesangial cells. CONCLUSIONS: These results suggest a potential role for MG in glomerular injury through p38 MAPK activation under diabetic conditions and may serve as a novel insight into the therapeutic strategies for diabetic nephropathy.     

15-Deoxy-Delta12,14-prostaglandin J2 regulates mesangial cell proliferation and death

BACKGROUND: Proliferation of intrinsic glomerular cells is a common response to renal injury. Acutely, proliferation may be beneficial, but sustained glomerular hypercellularity after injury is associated with progressive renal failure. To identify endogenous factors that may be responsible for regulating glomerular cell number, the effects of J-series cyclopentenone prostaglandins (PGs) on human glomerular mesangial cell proliferation and death were examined. METHODS: Human mesangial cells were grown in the presence or absence of PGJ2 or its metabolite 15-Deoxy-Delta12,14-PGJ2 (15dPGJ2). The number of viable cells was measured by the reduction of the tetrazolium MTS to a colored formazan product. Apoptosis was assessed by caspase-3 activation and DNA fragmentation. RESULTS: PGJ2 at concentrations up to 10 micromol/L caused mesangial proliferation. 15dPGJ2 also caused mesangial proliferation at low concentrations (< or =2.5 micromol/L), but induced mesangial cell death at higher concentrations (>5 micromol/L). Cell death occurred in part through apoptosis, measured as an increase in caspase-3 activity and DNA fragmentation in 15dPGJ2-treated cells. Cell death was associated with a decline in baseline phosphorylation of the survival factor Akt and increased Akt degradation, whereas 15dPGJ2-induced mesangial proliferation was blocked by inhibition of the PI 3-kinase/Akt pathway. 15dPGJ2 is a potent PPARgamma agonist. Like 15dPGJ2, treatment of mesangial cells with thiazolidinedione-type PPARgamma ligands (10 to 20 micromol/L) caused significant cell death, but at lower concentrations also caused a small degree of proliferation. CONCLUSIONS: J-series prostaglandins thus may be involved in the initiation of glomerular hypercellularity through Akt-dependent proliferation, and restoration of normal glomerular architecture through PPARgamma-mediated apoptosis. Manipulation of these prostaglandins may be relevant to the treatment of progressive glomerular disease.     

丝裂霉素C诱导成纤维细胞凋亡的作用及其机制

目的 观察丝裂霉素C(MMC)对成纤维细胞诱导凋亡的作用及机制.方法 体外培养成纤维细胞并传至5～8代,分别用含有0～0.3g/L MMC的MEM培养液处理成纤维细胞12h后CCK-8试剂盒检测不同浓度MMC对成纤维细胞的生长抑制效果;以0～0.2g/L MMC处理细胞爬片后用Hoechest33342细胞核染色,荧光显微镜下观察凋亡细胞核形态并用image pro-plus软件计数凋亡细胞;按不加药培养细胞12 h作对照组、0.2g/L.MMC培养细胞12h、P13K/Akt抑制剂LY294002 100 nmol/L与MEK/ERK抑制剂PD98059 100nmol/L分别处理细胞1h后换新鲜培养基培养11h分为4组,提取胞质蛋白,Western blot法检测Caspase-3、p-ERK1/2、p-Akt表达量.结果 MMC对成纤维细胞的抑制率与药物浓度呈正相关性,0.2g/L MMC对细胞活性抑制率达50.21%;Hoeeheat33342细胞核染色随MMC浓度升高出现凋亡细胞核特征的染色质凝聚,边集,呈亮蓝色核的比率逐渐增高,凋亡细胞计数显示0.05、0.1、0.2g/L MMC处理组凋亡率显著高于其他低浓度组,差异有统计学意义(P<0.05).Western blot检测0.2 g/L MMC组、PD98059组、LY294002组C8spase-3表达均显著高于对照组,p-ERK1/2、p-Akt表达均较对照组降低.结论 一定浓度的MMC可诱导成纤维细胞发生凋亡,其效应途径部分是通过降低ERK1/2、Akt的磷酸化水平使胞质内凋亡执行蛋白Caspase-3的表达水平增加,诱导成纤维细胞由正常增生状态转而发生凋亡减少胶原纤维生成,从而抑制瘢痕形成.     

Effects of triptolide on proliferation and apoptosis in rat mesangial cells induced by transforming growth factor β1 and related mechanisms

Objective To investigate the effects of triptolide on proliferation, apoptosis and the changes of Ski, Smad3, Smad7 and collagen type Ι (ColΙ) in cultured rat mesangial cells induced by transforming growth factor (TGF)?β1. Methods Cultured HBZY?1 rat mesangial cells were divided into 5 groups: (1)normal control group; (2)TGF?β1 group (10 μg/L); (3)-(5)triptolide (0.4, 2, 10 μg/L)+TGF?β1 (10 μg/L) groups. The cell proliferation was detected by MTT. Apoptosis of mesangial cells was detected by TUNEL assay. The expressions of Ski, Smad3, Smad7 mRNA were examined by real?time quantitative PCR. The expressions of Ski, Smad3, Smad7 and ColΙ protein were detected by Western blotting. The localizations of Ski and Smad3 protein were detected by laser confocal fluorescence microscope. Results Compared with the normal control, TGF?β1 (10 μg/L) significantly stimulated mesangial cells proliferation, while decreased apoptosis. The mRNA and protein expressions of Ski, Smad7, Smad3 and ColΙ protein expression in TGF?β1 group were increased (P＞0.05). In comparison with TGF?β1 group, triptolide could significantly inhibit TGF?β1?induced mesangial cells proliferation in dose?dependent manner, and promote the apoptosis of mesangial cells. In TGF?β1 group, mRNA and protein expresscons of Ski and Smad7 were increased (P<0.05), Smad3 mRNA and protein were decreased (P＞0.05), and ColΙ protein was decreased (P<0.01). In comparison with TGF?β1 group, fluorescence intensity of Ski, Smad3 proteins was significantly increased in cytoplasm, while decreased in nucleus. Conclusions Triptolide can inhibit TGF?β1?induced mesangial cells proliferation through regulating the expressions of Ski, Smad7 mRNA and protein, inhibiting Ski. Smad7 translocation to the nucleus, and down?regulating Smad3 mRNA and protein expression. Triptolide can promote apoptosis of mesangial cells.     

Overexpression of decorin induces apoptosis and cell growth arrest in cultured rat mesangial cells in vitro

Background: Decorin (DCN) is a small leucine-rich proteoglycan that plays an important role in the regulation of intercellular contact, cell migration and proliferation. DCN suppresses cell growth and induces apoptosis in various tumour cells. The aim of this study was to investigate whether overexpression of DCN could induce apoptosis and cell growth arrest in mesangial cells (MsCs) in vitro. Methods: PcDNA3.1A-DCN plasmid was transfected into cultured rat MsCs, and positive clones stably expressing DCN (MsC/DCN) were selected. SiRNA was used for blocking DCN expression in MsC/DCN. Apoptosis and cell growth of MsCs were assayed by flow cytometry. Hoechst staining was used for observing apoptotic cells. Expressions of active Caspase-3, epidermal growth factor receptor (EGFR), P21 and transforming growth factor-β (TGF-β1) were analyzed using Western blot. Results: Overexpression of DCN in MsCs induced apoptosis and arrested cells in the G₀/G₁ phase. The protein level of active Caspase-3 was significantly elevated in MsC/DCN (P < 0.01). DCN transfection induced downregulation of EGFR and up-expression of P21. In addition, the expression of TGF-β1 was significantly inhibited. DCN-siRNA transfection remarkably blocked the expression of DCN and reversed the downregulatory effects of DCN on MsC's proliferation. Conclusion: Overexpression of DCN could inhibit MsCs proliferation by inducing apoptosis and cell growth arrest in vitro and it also downregulates expression of TGF-β1. These results suggest novel strategies for regulating the proliferation of MsC in glomerular diseases.