大黄酸及大黄素诱导高糖大鼠肾小球系膜细胞的凋亡

背景：课题组前期实验证实中药复方消可宁能有效防治早期糖尿病肾病。目的：比较大黄酸与大黄素对高糖培养的大鼠肾小球系膜细胞凋亡的影响程度。方法：分别用不同浓度20,40,80μmol/L的大黄酸和大黄素刺激高糖培养的肾小球系膜细胞,苏木精-伊红染色观察凋亡细胞形态,DAPI荧光染色观察细胞核凋亡情况,流式细胞仪观察细胞凋亡率。结果与结论：苏木精-伊红染色及 DAPI 染色结果显示大黄酸对髙糖培养的肾小球系膜细胞凋亡的影响程度强于大黄素的影响程度。细胞凋亡率的对比显示大黄酸对髙糖培养的肾小球系膜细胞的早期凋亡率与晚期凋亡率均强于大黄素。说明低中高浓度的大黄酸、大黄素均可诱导肾小球系膜细胞凋亡,但大黄酸的药效强于大黄素。     

大黄酸可影响高糖培养SD大鼠肾小球系膜细胞的凋亡

背景：含有大黄的复方中药消可宁能有效防治早期糖尿病肾病。目的：观察大黄酸对髙糖培养的SD大鼠肾小球系膜细胞凋亡的影响。方法：以20,40,80μmol/L浓度的大黄酸刺激髙糖培养的肾小球系膜细胞,苏木精-伊红染色观察凋亡细胞形态,DAPI荧光染色观察细胞核凋亡情况,流式细胞仪观察细胞凋亡率。结果与结论：苏木精-伊红染色及DAPI染色结果显示大黄酸可促进髙糖培养的肾小球系膜细胞的凋亡,且与浓度与时间成正相关。各组肾小球系膜细胞的细胞凋亡率差异有显著性意义（P〈0.05）,表明大黄酸对髙糖培养的肾小球系膜细胞凋亡产生影响且与浓度及时间成正相关。     

青蒿琥酯对脂多糖诱导的大鼠肾小球系膜细胞凋亡的影响

目的:观察青蒿琥酯对脂多糖诱导的大鼠肾小球系膜细胞(glomerular mesangial cells,GMC)凋亡的影响。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。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。结论:青蒿琥酯能够剂量依赖性地诱导大鼠系膜细胞凋亡。     

应用4,6-联脒-2-苯基吲哚大体染色法快速观察生物支架材料体外内皮化程度

目的:建立一种新的形态学方法,以简便快捷地观察生物支架材料体外内皮化的程度。方法:实验于2005-07/2006-02在复旦大学上海医学院分子生物学实验室完成。①新鲜的猪心瓣膜经过十二烷基磺酸钠和脱氧胆酸处理,获得去细胞生物支架。②采用消化法获得脐静脉内皮细胞。③将脐静脉内皮细胞(2×105/cm2)种植于支架上,静态培养10d。用0.5mg/L的4,6-联脒-2-苯基吲哚溶液对再内皮化支架进行直接的大体染色及染色后冰冻切片,同时用苏木精-伊红染色,免疫荧光染色和扫描电镜对支架的内皮化情况进行平行观察。结果:①猪心瓣膜的细胞成分完全脱去,胞外基质保存完好。②原代分离的脐静脉内皮细胞,培养5d后融合成扁平单层,梭形或多角形,具有典型的铺路石样形态;CD31免疫荧光染色细胞表面呈现黄绿色荧光。③脐静脉内皮细胞种植10d后,支架经4,6-联脒-2-苯基吲哚大体染色显示内皮化程度达90%以上,观察结果与电镜观察相同;冰冻切片观察显示支架表面有一融合的内皮细胞单层,与免疫荧光染色及常规苏木精-伊红染色结果相吻合。结论:4,6-联脒-2-苯基吲哚大体染色法可直接观察生物支架再内皮化程度,简便、准确直观,是体外构建组织的一种有效的形态学检测方法。     

糖肾汤对大鼠肾小球系膜细胞增殖的影响

背景肾小球系膜细胞是糖尿病肾病致病的主要靶细胞,高糖环境下系膜细胞发生一系列功能改变,导致疾病不断发生发展,应用中药进行干预治疗,对延缓糖尿病肾病的发展具有重要意义.目的探讨肾小球系膜细胞在高糖环境下增殖情况以及糖肾汤含药血清对其的影响.设计对照实验.单位浙江中医学院附属第二医院实验室.材料以2003-08取自上海市第一人民医院肾内科SD大鼠系膜细胞株作为实验对象,2003-10浙江中医学院实验动物中心的清洁级SD大鼠45只,雌雄各半,体质量(180±20)g,作为制备含药血清动物.糖肾汤(由大黄、桃仁、虫、黄芪、地骨皮、黄连等组成),按比例称取后,水煎、过滤、浓缩至含生药3 g/mL.方法将SD大鼠随机分为正常组、糖肾汤组、卡托普利组各15只.将糖肾汤和卡托普利含药血清分为不同的剂量组作用于系膜细胞体系,正常组每日灌胃相同容量蒸馏水,用锥虫蓝染色法和四唑盐法分别检测用药后对系膜细胞毒性及增殖的影响.主要观察指标在含药血清刺激后大鼠肾小球系膜细胞活力,及细胞增殖情况.结果实验大鼠均进入结果分析.①系膜细胞在高糖持续作用下,其增殖受到抑制(与空白组比较P＜0.05),糖肾汤及卡托普利干预后均可不同程度地逆转上述高糖的抑增殖作用(与高糖组比较P＜0.05).②各组含药血清干预大鼠肾小球系膜细胞后,无明显毒性作用(细胞活力均达94%以上,与空白组比较P＞0.05).结论高糖可抑制系膜细胞增殖,而糖肾汤可刺激系膜细胞增殖.     

大鼠脂肪干细胞与细菌纤维素膜的复合培养

背景:国外一些研究表明细菌纤维素作为组织工程支架材料在组织工程方面有很大的应用潜力,但国内在这方面的报道非常少.目的:以大鼠脂肪干细胞作为组织工程种子细胞,考察其与细菌纤维素复合培养的效果.设计、时间及地点:观察性实验,于2007-0912008-08在吉林大学约学院生物工程实验中心和吉林大学第一医院耳鼻咽喉-头颈外科研究所完成.材料:细菌纤维素膜由天津科技大学天津市工业微生物重点实验室制备,材料的平均孔径为0.6-2.8μm,孔隙率为90%.方法:取3周龄Wistar大鼠腹股沟部脂肪垫,以Ⅱ型胶原酶消化获得大鼠脂肪干细胞,采用含有体积分数为0.1新生牛血清的DMEM培养液进行培养.将处十对数生长期的细胞,以3.0×108L-1的密度种植于细菌纤维素膜上,采用含有体积分数为0.1新生牛血清的DMEM培养液继续培养,分别于培养的不同时间取细胞-材料复合物,冰冻切片.采用免疫荧光染色方法对生长于细菌纤维素膜上脂肪干细胞的生物学特征进行测定.主要观察指标:①体外培养大鼠脂肪干细胞形态.②苏木精-伊红染色观察脂肪干细胞在细菌纤维素膜上的生长情况.③免疫荧光染色标记蛋白-CD29在复合物上脂肪干细胞中的表达.结果:体外原代培养72 h的脂肪干细胞以长梭形为主,7 d后梭形细胞逐渐增多融合,细胞对数生长期为第3～9天.细胞-支架复合物在培养第10天经苏木精-伊红染色后可见细胞呈单层生长于细菌纤维素膜表面,细胞与细菌纤维素膜接触紧密,未见深入细菌纤维素膜内部生长的细胞.免疫荧光技术分析结果表明,标记蛋白-CD29在细菌纤维素膜上的脂肪干细胞有特异性表达.结论:生长于细菌纤维素膜上的脂肪干细胞不仅能够增殖,而且仍然保持着干细胞的生物学活性.     

选择性环氧合酶抑制剂NS-398对高糖诱导系膜细胞增殖的影响

目的:研究选择环氧合酶抑制剂NS-398对高糖诱导的系膜细胞抑制作用.方法:空白组,不加任何刺激剂;高糖组,加入30 mmol/L葡萄糖处理;干预组,不同浓度选择性环氧合酶抑制剂NS-398(5、10、20、40μmol/L)加入高糖培养的大鼠肾小球系膜细胞中,分别共同培养0、12、24、48、72 h,用[3H]-亮氨酸掺入法检测细胞蛋白质的从头合成情况,用流式细胞仪分析检测细胞G0-G1周期分布的变化,观察选择性环氧合酶抑制剂NS-398对系膜细胞增殖的影响.结果:高糖刺激的系膜细胞的[3H]-亮氨酸掺入量明显增加,用不同浓度的NS-398共同培养高糖系膜细胞,观察不同时间[3H]-亮氨酸掺入量,选择性环氧合酶抑制剂NS-398在5～40μmol/L浓度范围内,随着时间的延长、浓度的增加,可明显抑制MC[3H]-亮氨酸掺入,且呈浓度和时间依赖性.流式细胞仪检测干预组细胞G0-G1周期百分比较高糖组差异有显著性(P＜0.05或P＜0.01).结论:选择性环氧合酶抑制剂NS-398能抑制高糖诱导系膜细胞增殖,促进细胞凋亡.     

兔骨髓间充质干细胞与羊膜的共培养

背景:近年来利用组织工程技术构建软骨以修复软骨缺损已经取得较大进步。骨髓间充质干细胞在软骨组织工程研究中作为一种常见的种子细胞被广泛应用。目的:以人羊膜作为细胞支架负载兔骨髓间充质干细胞,观察羊膜的生物相容性。方法:将兔骨髓间充质干细胞接种于人脱细胞羊膜上,体外共培养后进行形态学观察,以四甲基偶氮唑盐比色法观察骨髓间充质干细胞增殖活性,评价羊膜的组织相容性。应用4’,6-二脒基-2-苯基吲哚染色法观察细胞存活和凋亡情况。结果与结论:骨髓间充质干细胞在羊膜上生长较好,细胞活力增强,4’,6-二脒基-2-苯基吲哚染色结果示仅有少量细胞凋亡。提示羊膜是一种具有良好生物相容性的支架材料,可以促进骨髓间充质干细胞增殖。     

糖肾汤对大鼠肾小球系膜细胞增殖的影响

背景：肾小球系膜细胞是糖尿病肾病致病的主要靶细胞,高糖环境下系膜细胞发生一系列功能改变,导致疾病不断发生发展,应用中药进行干预治疗,对延缓糖尿病肾病的发展具有重要意义.目的：探讨肾小球系膜细胞在高糖环境下增殖情况以及糖肾汤含药血清对其的影响.设计：对照实验.单位：浙江中医学院附属第二医院实验室.材料：以2003-08取自上海市第一人民医院肾内科SD大鼠系膜细胞株作为实验对象,2003-10浙江中医学院实验动物中心的清洁级SD大鼠45只,雌雄各半,体质量（180&;#177;20）g,作为制备含药血清动物.糖肾汤（由大黄、桃仁、虫、黄芪、地骨皮、黄连等组成）,按比例称取后,水煎、过滤、浓缩至含生药3 g/mL.方法：将SD大鼠随机分为正常组、糖肾汤组、卡托普利组各15只.将糖肾汤和卡托普利含药血清分为不同的剂量组作用于系膜细胞体系,正常组每日灌胃相同容量蒸馏水,用锥虫蓝染色法和四唑盐法分别检测用药后对系膜细胞毒性及增殖的影响.主要观察指标：在含药血清刺激后大鼠肾小球系膜细胞活力,及细胞增殖情况.结果：实验大鼠均进入结果分析.①系膜细胞在高糖持续作用下,其增殖受到抑制（与空白组比较P＜0.05）,糖肾汤及卡托普利干预后均可不同程度地逆转上述高糖的抑增殖作用（与高糖组比较P＜0.05）.②各组含药血清干预大鼠肾小球系膜细胞后,无明显毒性作用（细胞活力均达94%以上,与空白组比较P＞0.05）.结论：高糖可抑制系膜细胞增殖,而糖肾汤可刺激系膜细胞增殖.     

青藤碱对大鼠肾小球系膜细胞增殖和凋亡的影响

目的探讨青藤碱(sinomenine,SIN)对大鼠肾小球系膜细胞(mesangial cell,MC)增殖和凋亡的影响。方法体外培养的大鼠MC,分为SIN分为高、中、低剂量组和空白对照组。采用四甲基偶氮唑蓝(MTT)法检测细胞的增殖情况,碘化丙啶染色后流式细胞术检测凋亡率,吖啶橙/溴乙锭染色后采用荧光显微镜观察细胞凋亡水平。结果MTT实验和流式细胞检测结果显示,与空白对照组比较,SIN高(P<0.01)、中(P<0.05)、低(P<0.05)剂量组MC增殖降低,凋亡增加,吖啶橙/溴乙锭染色结果显示SIN干预的MC出现凋亡的百分率增加。结论青藤碱可抑制大鼠肾小球系膜细胞增殖并促进其凋亡,可能会使肾脏病理性损害减轻。     

Rhein effects on apoptosis of glomerular mesangial cells cultured with high glucose in Sprague Dawley rats

BACKGROUND:Our previous study has already manifested that Chinese medicine Xiaokening containing Rhein can effectively control the development of early diabetic nephropathy. OBJECTIVE:To investigate the effect of Rhein on apoptosis of glomerular mesangial cel s (GMCs) of Sprague Dawley rats cultured with high glucose. METHODS:The GMCs cultured with high glucose were stimulated by different concentrations of Rhein (20,40,80μmol/L). Morphology of GMCs was discriminated by hematoxylin-eosin staining. Karyon apoptosis was examined by fluorescence staining of 4’,6-diamidino-2-phenylindole (DAPI). Cel apoptosis rate was detected by flow cytometry. RESULTS AND CONCLUSION:The outcome of hematoxylin-eosin staining and 4’,6-diamidino-2-phenylindole staining indicated that Rhein could obviously induce apoptosis of GMCs in a dose-and time-dependent manner. The apoptosis rate had significant difference among different groups (P<0.05). This result shows that Rhein can obviously induce apoptosis of GMCs cultured with high glucose and in a dose-and time-dependent manner.     

Sonodynamically Induced Apoptosis by Protoporphyrin IX on Hepatoma-22 cells In Vitro

The synergistic effect of ultrasound and certain chemicals on cells is known as sonodynamic therapy (SDT). It has been reported that the direct sonochemical and subsequent redox reactions induced by SDT treatment can lead to apoptotic cell death. However, the detailed biologic mechanism about it is not well understood until now. In this study the effect of low-intensity ultrasound on Hepatoma-22 cells (H22) in the presence of the sonosensitizing drug protoporphyrin IX (PpIX) was evaluated at different incubation times after sonication. Trypan blue exclusion was used to detect cell viability. The presence of apoptotic cells was identified by 4'-6-diamidino-2-phenylindole (DAPI) nuclear staining and transmission electric microscope (TEM) observation. An inverted confocal laser scanning microscope was used to detect the release of mitochondrial protein cytochrome c (Cyt c) and the redistribution of Bcl-2 family proteins Bid and Bax. Additionally, the generation of intracellular reactive oxygen species (ROS) and the loss of mitochondria membrane potential (MMP) were quantificationally measured by a fluorescence microplate reader. The results indicated that the synergistic cytotoxicity of PpIX and ultrasound increased in a time-dependent manner and the mitochondria damage may be the main factor for sonodynamically induced apoptosis by PpIX in H22 cells. (E-mail: lshaof@snnu.edu.cn)     

超顺磁性氧化铁和DAPI双标记骨髓间充质干细胞

背景：骨髓间充质干细胞目前已经成为重要的组织工程种子细胞,而若想深入研究其在体内外增殖、分化的规律,首先需要解决如何能高效、安全地标记骨髓间充质干细胞。目的：观察超顺磁性氧化铁及DAPI对大鼠骨髓间充质干细胞的双标记效果及其对细胞存活、增殖以及凋亡的影响。方法：分离培养大鼠骨髓间充质干细胞,用超顺磁性氧化铁及DAPI双标记后分别用普鲁士蓝染色法和激光共聚焦显微镜观察其铁标记率及荧光标记率。用锥虫蓝检测细胞活力;MTT法检测标记干细胞增殖力;Calcein-AM/PI以及AO/PI双染细胞,检测细胞存活率以及凋亡率。结果与结论：超顺磁性氧化铁及DAPI在体外双标记大鼠骨髓间充质干细胞效率高,可达100%;锥虫蓝染色显示标记细胞的存活率为97%;MTT法检测发现双标记干细胞组的活力以及增殖力与未标记干细胞组相比差异均无显著性意义（P〉0.05）;Calcein-AM/PI染色,未标记细胞及双标记细胞的存活率分别为96%和95%;AO/PI染色,未标记及双标记细胞的凋亡率均为1%。提示超顺磁性氧化铁和DAPI双标记大鼠骨髓间充质干细胞后,对于干细胞的存活、增殖以及凋亡无影响。     

YC-1 suppresses constitutive nuclear factor-kappaB activation and induces apoptosis in human prostate cancer cells

Although the indazole compound, YC-1, is reported to exert anticancer activities in several cancer cell types, its target and mechanism of action have not been well explored. The objectives of this study were to ascertain whether YC-1 directly induces apoptosis in prostate cancer cells and to explore the mechanism(s) whereby YC-1 causes cell death. Hormone-refractory metastatic human prostate cancer PC-3 cells were selected for this study. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay indicated that YC-1 suppresses growth of PC-3 cells in a concentration-dependent and time-dependent manner. Apoptosis was determined using 4',6-diamidino-2-phenylindole staining, and cell cycle progression was examined by FACScan flow cytometry. YC-1 treatment showed chromatin condensation and increased the percentage of PC-3 cells in the hypodiploid sub-G0-G1 phase, indicative of apoptosis. Additionally, exposure to YC-1 was found to induce activation of caspase-3 and cleavage of poly(ADP-ribose) polymerase. Translocation and activation of nuclear factor-kappaB (NF-kappaB) were determined by immunofluorescent staining and ELISA, respectively. The results showed that YC-1 abolished constitutive nuclear translocation and activation of NF-kappaB/p65. Furthermore, inhibition of inhibitor of kappaBalpha (IkappaBalpha) phosphorylation and accumulation of IkappaBalpha were observed. The antitumor effects of YC-1 were evaluated by measuring the growth of tumor xenografts in YC-1-treated severe combined immunodeficient mice. The volumes of PC-3 tumors produced in severe combined immunodeficient mice were observed to decline significantly after treatment with YC-1 compared with vehicle controls. We concluded that the antitumor effects of YC-1 in PC-3 cells include the induction of apoptosis and the suppression of NF-kappaB activation. Given these unique actions, further investigations of the effects of YC-1 against hormone-refractory prostate cancer are warranted.     

肝星状细胞活化增殖和凋亡及奥曲肽的影响

背景:肝星状细胞的活化增殖在肝硬化的发生发展中起关键作用,临床广泛应用的生长抑素衍生物奥曲肽有多种生物学效应,但它对肝星状细胞的活化增殖及凋亡有何影响尚不清楚。目的:观察奥曲肽对大鼠肝星状细胞增殖和凋亡的影响。方法:实验选用肝星状细胞株HSC-T6的传代细胞。MTT法检测不同浓度(1×10-2,1×10-3,1×10-4,1×10-5,1×10-6,0mmol/L)奥曲肽干预24,48,72h对肝星状细胞增殖的影响;TUNEL凋亡检测试剂盒荧光显微镜方法检测不同浓度(0,1×10-5,1×10-2mmol/L)奥曲肽干预24h对肝星状细胞凋亡的影响。结果与结论:奥曲肽浓度越高、作用时间越长对培养的肝星状细胞增殖抑制越明显,呈现浓度和时间依赖性;奥曲肽对培养的肝星状细胞凋亡随奥曲肽浓度的增加而增加。结果可见奥曲肽对培养的肝星状细胞增殖抑制呈浓度(0～10-2mmol/L)和时间(24,48,72h)依赖性,并能诱发其凋亡。     

大鼠胚胎后肾间充质细胞体外标记的方法

背景:干细胞移植为慢性肾脏病的治疗提供了新的视角,特异性标记干细胞并体内示踪是这一领域研究的基础,目前尚无一种公认的适用于所有细胞的标记方法.目的:观察DAPI及绿色荧光蛋白标记细胞在阿霉素肾病大鼠体内的定位及分化,探讨便捷的大鼠胚胎后肾间充质细胞的体外标记方法.设计、时间及地点:分组对比观察,于2007-04/12在中南大学湘雅二医院完成.材料:清洁级雌性Sprague-Dawley(SD)大鼠60只,体质量180～220 g,用于制备阿霉素肾病大鼠模型.方法:分别将DAPI、DAPI体外标记后肾间充质细胞、绿色荧光蛋白体外标记后肾间充质细胞,通过尾静脉注入阿霉素肾病大鼠体内:于细胞移植后1,3,5周行肾组织冰冻切片,观察DAPI及绿色荧光蛋白荧光分布,并ABC免疫酶染色法检测肾组织中绿色荧光蛋白表达.主要观察指标:比较DAPI及绿色荧光蛋白两种不同方法标记后肾间充质细胞在阿霉素慢性肾病大鼠肾内的定植情况,以及绿色荧光蛋白标记细胞不同时间点移植在肾内的定植改变.结果:①DAPI标记细胞及单纯DAPI尾静脉注射后1周,均可在肾小管上皮细胞中观察到DAPI阳性细胞,至移植后5周仍可观察到,但随时间延长荧光有减弱趋势.②绿色荧光蛋白标记细胞移植后1周,可在肾小管上皮细胞中观察到绿色荧光蛋白阳性细胞,至移植后5周仍可观察到,荧光无明显减弱.③ABC免疫酶染色法检测,显示绿色荧光蛋白阳性细胞主要表达于近端肾小管中,肾小球系膜区亦可观察到少最表达:绿色荧光蛋白阳性产物主要表达于胞浆.绿色荧光蛋白表达半定量评分显示,标记细胞的早期应用阳性率大于晚期应用者,且随观察时间延长,阳性率有升高趋势.结论;后肾间充质细胞的脂质体感染法绿色荧光蛋白标记是一便捷,有效的体外标记、示踪方法,适于移植细胞的短期内示踪、观察.     

黄芩素与顺铂联合用药对人骨肉瘤MG63细胞株细胞凋亡的影响

【目的】研究黄芩素与顺铂联合用药对人骨肉瘤MG63细胞增殖和凋亡的影响。【方法】在骨肉瘤MG63细胞株中,分别采用MTT法检测黄芩素单独用药,与顺铂合用后细胞活性的变化情况;Hochest染色、TuNEL法检测药物单用及合用时细胞凋亡的变化情况。【结果】黄芩素30μM对细胞活性没有明显影响,但在100pM可时间依赖的降低细胞活性。黄芩素（100gM）与顺铂（3肛M）舍用后,能明显抑制细胞活性,具呈时效关系。黄芩素（100μM）可诱导细胞发生凋亡,黄芩素（30～100肛M）与顺铂（3μM）两者合用具有协同诱导细胞凋亡的作用。【结论】黄苓素可抑制MG63细胞活性,诱导凋亡;与顺铂合用具有协同效应。     

Mesangial cell apoptosis induced by stimulation of the adenosine A3 receptor: signaling and apoptotic events.

Mesangial cell apoptosis has been proposed as a means of resolution of glomerular hypercellularity in proliferative forms of glomerular disease. We previously demonstrated that adenosine causes mesangial cell apoptosis by stimulating the A3-type adenosine receptor. This is a G protein-coupled receptor shown to activate kinases involved in apoptotic signaling. In this work, we assessed changes in phosphorylation of the mitogen-activated protein kinase extracellular signal-regulated kinase (ERK)1/2 and in levels of specific pro- and antiapoptotic proteins following exposure of mesangial cells to the A3 adenosine receptor agonist N(6)-(3-iodobenzyl)-adenosine-5'-N-methyluronamide (IB-MECA). Cultured mesangial cells were incubated with IB-MECA for 30 minutes and 6, 24, and 48 hours. IB-MECA was used at a concentration (30 microM) that induces a reproducible degree of mesangial cell apoptosis. Changes in ERK1/2 phosphorylation and in protein levels of Bcl-2, Bax, and caspase 3 were assessed by Western blot analysis. IB-MECA markedly increased phosphorylation of ERK1/2. This effect peaked at 5 minutes, dissipated by 20 minutes, and was abolished by the inhibitor of ERK phosphorylation, compound U0126, in a dose-dependent manner. This inhibitor had no effect on the extent of IB-MECA-induced apoptosis. Bcl-2 levels progressively declined, whereas those of Bax and activated caspase 3 increased. These observations indicate that stimulation of the A3-type adenosine receptor causes mesangial cell apoptosis via mechanisms independent of ERK activation. The observations also point to an imbalance in the expression of antiapoptotic (Bcl-2) and proapoptotic (Bax, caspase 3) proteins as a potential mechanism underlying adenosine-induced mesangial cell apoptosis.