Heat shock protein 90 protects rat mesenchymal stem cells against hypoxia and serum deprivation-induced apoptosis via the PI3K/Akt and ERK1/2 pathways.
Mesenchymal stem cell (MSC) transplantation has shown a therapeutic potential to repair the ischemic and infracted myocardium, but the effects are limited by the apoptosis and loss of donor cells in host cardiac microenvironment. The aim of this study is to explore the cytoprotection of heat shock protein 90 (Hsp90) against hypoxia and serum deprivation-induced apoptosis and the possible mechanisms in rat MSCs. Cell viability was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Apoptosis was assessed by Hoechst 33258 nuclear staining and flow cytometric analysis with annexin V/PI staining. The gene expression of Toll-like receptor-4 (TLR-4) and V-erb-b2 erythroblastic leukemia viral oncogene homolog 2 (ErbB2) was detected by real-time polymerase chain reaction (PCR). The protein levels of cleaved caspase-3, Bcl-2, Bcl-xL, Bax, total-ERK, phospho-ERK, total-Akt, phospho-Akt, and Hsp90 were detected by Western blot. The production of nitric oxide was measured by spectrophotometric assay. Hsp90 improves MSC viability and protects MSCs against apoptosis induced by serum deprivation and hypoxia. The protective role of Hsp90 not only elevates Bcl-2/Bax and Bcl-xL/Bax expression and attenuates cleaved caspase-3 expression via down-regulating membrane TLR-4 and ErbB2 receptors and then activating their downstream PI3K/Akt and ERK1/2 pathways, but also enhances the paracrine effect of MSCs. These findings demonstrated a novel and effective treatment strategy against MSC apoptosis in cell transplantation.
Gao
Feng
F
Department of Cardiology, the Second Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, China.
Hu
Xin-yang
XY
Xie
Xiao-jie
XJ
Xu
Qi-yuan
QY
Wang
Ya-ping
YP
Liu
Xian-bao
XB
Xiang
Mei-xiang
MX
Sun
Yong
Y
Wang
Jian-an
JA
D000818:Animals; D017209:Apoptosis; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism*; D000860:Hypoxia; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000737:chemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013234:Stem Cells / Q000166:cytology; D013778:Tetrazolium Salts / Q000494:pharmacology; D013844:Thiazoles / Q000494:pharmacology
D018841:HSP90 Heat-Shock Proteins; D011994:Recombinant Proteins; D013778:Tetrazolium Salts; D013844:Thiazoles; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D048049:Extracellular Signal-Regulated MAP Kinases; C022616:thiazolyl blue