GAPDH-knockdown reduce rotenone-induced H9C2 cells death via autophagy and anti-oxidative stress pathway.
BACKGROUND
GAPDH, well known for its house-keeping functions, has also been shown to be involved in cell injury, apoptosis and death under conditions of stress such as starvation, chemical injury and oxidative stress. This study examines the effect of GAPDH knockdown on cell injury in response to Rotenone.


METHODS
GAPDH was knocked down in H9C2 cardiomyoblasts using siRNA prior to exposure to rotenone (0 nM, 20 nM, 40 nM and 80 nM). Autophagy was detected by western blot for autophagy proteins (Beclin-1, Atg5, LC-3A/B and p62) and MDC staining for acidic substances. Pro-apoptosis protein and flow cytometry were used to assess cell apoptosis and death and intracellular ATP relative concentration was measured. Oxidant stress was assessed by measuring DCFH-DA, TBARS, GSH and SOD.


RESULTS
In this study, GAPDH-knockdown enhanced autophagy in rotenone-induced H9C2 cells, decreased oxidant stress and increased antioxidant pathways; and reduced cell apoptosis and death. Furthermore, GAPDH-knockdown preserved cell energy.


CONCLUSIONS
siRNA-mediated GAPDH knockdown reduced rotenone-induced H9C2 cell death occurring via autophagy and anti-oxidative stress pathway. This study enriches the understanding of GAPDH pathophysiology role, and provides potential new therapeutic targets for cardiac disease states characterized by oxidative stress.
Department of Cardiology, The Second Affiliated Hospital of Wenzhou Medical University, Wenzhou, Zhejiang Province, China. Electronic address: shaoliang021224@gmail.com.
Shao
S
Liang
North Shore Heart Research Group, Kolling Institute of Medical Research, University of Sydney, Sydney, Australia; Department of Cardiology, Royal North Shore Hospital, Sydney, Australia.
Gemma
G
Figtree
Department of Cardiovascular Medicine, First Affiliated Hospital of Medical College of Xi'an Jiaotong University, Xi'an, Shaanxi Province, China.
Ma
M
Aiqun
Department of Geriatrics & Neurology, The Second Affiliated Hospital, Wenzhou Medical University, Wenzhou, Zhejiang, China. Electronic address: zhangkiki520@126.com.
Zhang
Z
Ping
D000818:Animals; D000975:Antioxidants; D017209:Apoptosis / Q000187:drug effects; D001343:Autophagy / Q000187:drug effects*; D016923:Cell Death / Q000187:drug effects*; D002460:Cell Line; D055785:Gene Knockdown Techniques; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics / Q000502:physiology*; D032383:Myocytes, Cardiac / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D018384:Oxidative Stress / Q000187:drug effects*; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D012402:Rotenone / Q000494:pharmacology*
D000975:Antioxidants; D017382:Reactive Oxygen Species; D012402:Rotenone; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases
Autophagy; Cell injury; GAPDH; H9C2 cell; Oxidative stress; Rotenone