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2025年8月6日 星期三
农业生物技术学报  2019, Vol. 27 Issue (11): 2004-2012    DOI: 10.3969/j.issn.1674-7968.2019.11.011
  研究论文与报告 本期目录 | 过刊浏览 | 高级检索 |
葛根素通过抑制线粒体凋亡通路和调节HSP72表达缓解热应激诱导的LLC-PK1细胞凋亡
王若冰*, 朱雨凝*, 丰艳妮, 李华涛, 丛霞, 曹荣峰, 姜忠玲*, 田文儒*
青岛农业大学 动物医学院 繁殖障碍性疾病研究所,青岛 266109
Puerarin Attenuates Heat Stress-induced Apoptosis in LLC-PK1 Cells by Inhibiting Mitochondrial Apoptotic Pathway and Regulating HSP72 Expression
WANG Ruo-Bing*, ZHU Yu-Ning*, FENG Yan-Ni, LI Hua-Tao, CONG Xia, CAO Rong-Feng, JIANG Zhong-Ling**, TIAN Wen-Ru**
Institute of Reproductive Disorders, College of Veterinary Medicine, Qingdao Agricultural University, Qingdao 266109, China
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摘要 葛根素对动物具有解热作用,但其作用机制尚不清楚。本研究目的在于明确葛根素是否对受热应激猪(Sus scrofa)近端肾小管细胞(pig kidney proximal tubular cells, LLC-PK1)具有保护作用及其潜在机制。本研究将LLC-PK1细胞分为对照(C)组,热应激(HS, 42 ℃)组,葛根素(Pue, 10 µmol/L)组和热应激加葛根素(HS+Pue)组。分别用CCK-8和流式细胞术检测细胞生存率和凋亡率,使用qRT-PCR和Western blot分别检测B细胞淋巴瘤-2 (B-cell lymphoma, Bcl),Bcl-2相关X蛋白(Bcl-2 associated X protein, Bax)、细胞色素c (cytochrome c, Cyto c)和细胞凋亡诱导因子(apoptosis inducing factor, AIF) mRNA和蛋白表达水平,用相应试剂盒检测半胱天冬酶(cysteinyl aspartate specific proteinase, caspase) Caspase-9和Caspase-3活性及热休克蛋白(heat shock proteins, HSPs) HSP72和HSP90的含量。结果发现,葛根素抑制LLC-PK1线粒体Cyto c的释放和AIF mRNA的表达,并降低热应激条件下LLC-PK1细胞中Caspase-9和Caspase-3的活性。同时,热应激加葛根素组细胞中Bcl与Bax的比率以及HSP72的表达增加,HSP90的表达降低。上述结果表明,葛根素能降低热应激诱导的LLC-PK1细胞损伤,其机制是通过抑制线粒体凋亡通路的激活,上调细胞HSP72的表达来实现的。本研究揭示了葛根素抑制热应激诱导的LLC-PK1细胞损伤的部分机制,为临床上使用葛根素作为抗热应激的有效药物提供了基础资料。
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王若冰
朱雨凝
丰艳妮
李华涛
丛霞
曹荣峰
姜忠玲
田文儒
关键词 葛根素猪近端肾小管细胞(LLC-PK1)热应激凋亡热休克蛋白(HSPs)    
Abstract:Puerarin has been proved to have antipyretic effects in animals. However, how puerarin works on the cellular or molecular levels is still poorly understood. The aim of the present study is to determine if puerarin has protective effect on the pig (Sus scrofa) kidney proximal tubular cells (LLC-PK1) subjected to heat stress and its possible underlying mechanisms in vitro. The LLC-PK1 cells were divided into control (C) group, heat stress (HS, 42 ℃) group, puerarin (Pue, 10 µmol/L) group, and heat stress + puerarin (HS+Pue) group. Cell viability was measured by CCK-8; cells apoptosis rate was detected by flow cytometry; mRNA and protein expression levels of B-cell lymphoma-2 (Bcl-2), Bcl-2 associated X protein (Bax), cytochrome c (Cyto c) and apoptosis inducing factor (AIF) were detected by qRT-PCR and Western blot, respectively. Cysteinyl aspartate specific proteinase-9 (Caspase-9) and Caspase-3 activities and total protein content of heat shock protein 72 (HSP72) and HSP90 were detected with the corresponding kits. The results showed that puerarin suppressed Cyto c and AIF mRNA expressions and their proteins released from mitochondria, and decreased both Caspase-9 and Caspase-3 activities of the LLC-PK1 cells under HS. Moreover, puerarin further increased Bcl-2/Bax ratio and HSP72 expressions, and decreased HSP90 expressions of the cells under HS. The results indicated that puerarin attenuates HS-induced apoptosis of the LLC-PK1 cells through inhibiting mitochondrial apoptotic pathway activation, and up-regulating HSP72 expression. This study reveals a part of the mechanism by which puerarin inhibits heat stress-induced damage in LLC-PK1 cells, providing basic materials for the use of puerarin as an effective drug against heat stress in practice.
Key wordsPuerarin    Pig kidney proximal tubular cells (LLC-PK1)    Heat stress    Apoptosis    Heat shock proteins (HSPs)
收稿日期: 2019-03-06     
ZTFLH:  S854.53  
基金资助:国家自然科学基金(No. 31772815; No. 31572590)和山东省重点研发计划(No. 2018GNC110005)
通讯作者: jzhl200666@126.com;wrtian@126.com   
引用本文:   
王若冰, 朱雨凝, 丰艳妮, 李华涛, 丛霞, 曹荣峰, 姜忠玲, 田文儒. 葛根素通过抑制线粒体凋亡通路和调节HSP72表达缓解热应激诱导的LLC-PK1细胞凋亡[J]. 农业生物技术学报, 2019, 27(11): 2004-2012.
WANG Ruo-Bing, ZHU Yu-Ning, FENG Yan-Ni, LI Hua-Tao, CONG Xia, CAO Rong-Feng, JIANG Zhong-Ling, TIAN Wen-Ru. Puerarin Attenuates Heat Stress-induced Apoptosis in LLC-PK1 Cells by Inhibiting Mitochondrial Apoptotic Pathway and Regulating HSP72 Expression. 农业生物技术学报, 2019, 27(11): 2004-2012.
链接本文:  
http://journal05.magtech.org.cn/Jwk_ny/CN/10.3969/j.issn.1674-7968.2019.11.011     或     http://journal05.magtech.org.cn/Jwk_ny/CN/Y2019/V27/I11/2004
 
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