Des-acyl ghrelin protects microvascular endothelial cells from oxidative stress-induced apoptosis through sirtuin 1 signaling pathway.
OBJECTIVE
Ghrelin is a stomach-derived hormone. Acylation of ghrelin has been essential for its biological activities such as stimulating appetite. On the other hand, the function of des-acyl ghrelin (Des-G) has not been fully elucidated. The aim of the present study is to examine the anti-apoptotic effect of Des-G on endothelial cells.


METHODS
After human retinal microvascular endothelial cells (RMECs) were pretreated with or without 100nmol/L Des-G, apoptosis was induced with 0.1mmol/L hydrogen peroxide (H2O2). For pharmacological inhibition of surtuin 1 (SIRT1) catalytic activity, the cells were treated with 10μmol/L Ex-527. Inhibition of SIRT1 with siRNA was also performed. The quantitative estimation of DNA fragmentation was used as a marker of apoptosis. Furthermore, total SIRT activity in nuclear extracts, mRNA and protein levels of SIRT1, manganese superoxide dismutase (MnSOD) and catalase were determined.


RESULTS
Des-G pretreatment protected RMECs from oxidative stress-induced apoptosis and increased SIRTs deacetylase activity in nuclear extracts. On the other hand, both pharmacological and siRNA mediated inhibition of SIRT1 attenuated the anti-apoptotic effect of Des-G. Moreover, Des-G increased mRNA and protein levels of SIRT1 and antioxidant enzymes such as MnSOD and CAT, which are downstream targets of SIRT1. Although the treatment of Ex-527 did not alter mRNA expression levels of SIRT1, it decreased mRNA expression levels of antioxidant enzymes in the cells with Des-G pretreatment.


CONCLUSIONS
Our results suggest that SIRT1 signaling pathway contributes to protective effect of Des-G against oxidative stress-induced apoptosis.
The First Department of Medicine, Wakayama Medical University, Wakayama, Japan.
Takeshi
T
Shimada
The First Department of Medicine, Wakayama Medical University, Wakayama, Japan. Electronic address: hfuruta@wakayama-med.ac.jp.
Hiroto
H
Furuta
The First Department of Medicine, Wakayama Medical University, Wakayama, Japan.
Asako
A
Doi
The First Department of Medicine, Wakayama Medical University, Wakayama, Japan.
Hiroyuki
H
Ariyasu
The First Department of Medicine, Wakayama Medical University, Wakayama, Japan.
Hiromichi
H
Kawashima
The First Department of Medicine, Wakayama Medical University, Wakayama, Japan.
Hisao
H
Wakasaki
The First Department of Medicine, Wakayama Medical University, Wakayama, Japan.
Masahiro
M
Nishi
The First Department of Medicine, Wakayama Medical University, Wakayama, Japan.
Hideyuki
H
Sasaki
The First Department of Medicine, Wakayama Medical University, Wakayama, Japan.
Takashi
T
Akamizu
D017209:Apoptosis; D015153:Blotting, Western; D002374:Catalase / Q000378:metabolism; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology* / Q000201:enzymology / Q000378:metabolism; D054439:Ghrelin / Q000502:physiology*; D006801:Humans; D055806:Microvessels / Q000166:cytology* / Q000201:enzymology / Q000378:metabolism; D018384:Oxidative Stress; D034741:RNA, Small Interfering / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D012171:Retinal Vessels / Q000166:cytology / Q000201:enzymology / Q000378:metabolism; D015398:Signal Transduction; D056564:Sirtuin 1 / Q000235:genetics / Q000378:metabolism*; D013482:Superoxide Dismutase / Q000378:metabolism
D054439:Ghrelin; D034741:RNA, Small Interfering; C422650:ghrelin, des-n-octanoyl; D002374:Catalase; D013482:Superoxide Dismutase; D056564:Sirtuin 1
Catalase; Ghrelin; Manganese superoxide dismutase; Sirtuin