title: Prevention of hyperglycemia-induced myocardial apoptosis by gene silencing of Toll-like receptor-4.
abstract: BACKGROUND
  Apoptosis is an early event involved in cardiomyopathy associated with diabetes mellitus. Toll-like receptor (TLR) signaling triggers cell apoptosis through multiple mechanisms. Up-regulation of TLR4 expression has been shown in diabetic mice. This study aimed to delineate the role of TLR4 in myocardial apoptosis, and to block this process through gene silencing of TLR4 in the myocardia of diabetic mice.
  
  
  METHODS
  Diabetes was induced in C57/BL6 mice by the injection of streptozotocin. Diabetic mice were treated with 50 μg of TLR4 siRNA or scrambled siRNA as control. Myocardial apoptosis was determined by TUNEL assay.
  
  
  RESULTS
  After 7 days of hyperglycemia, the level of TLR4 mRNA in myocardial tissue was significantly elevated. Treatment of TLR4 siRNA knocked down gene expression as well as diminished its elevation in diabetic mice. Apoptosis was evident in cardiac tissues of diabetic mice as detected by a TUNEL assay. In contrast, treatment with TLR4 siRNA minimized apoptosis in myocardial tissues. Mechanistically, caspase-3 activation was significantly inhibited in mice that were treated with TLR4 siRNA, but not in mice treated with control siRNA. Additionally, gene silencing of TLR4 resulted in suppression of apoptotic cascades, such as Fas and caspase-3 gene expression. TLR4 deficiency resulted in inhibition of reactive oxygen species (ROS) production and NADPH oxidase activity, suggesting suppression of hyperglycemia-induced apoptosis by TLR4 is associated with attenuation of oxidative stress to the cardiomyocytes.
  
  
  CONCLUSIONS
  In summary, we present novel evidence that TLR4 plays a critical role in cardiac apoptosis. This is the first demonstration of the prevention of cardiac apoptosis in diabetic mice through silencing of the TLR4 gene.
authors.affiliation: Department of Endocrinology, West China Hospital of Sichuan University, Chengdu, China. tongnanwei@yahoo.com.cn
authors.forename: Yuwei
authors.initials: Y
authors.lastname: Zhang
authors.forename: Tianqing
authors.initials: T
authors.lastname: Peng
authors.forename: Huaqing
authors.initials: H
authors.lastname: Zhu
authors.forename: Xiufen
authors.initials: X
authors.lastname: Zheng
authors.forename: Xusheng
authors.initials: X
authors.lastname: Zhang
authors.forename: Nan
authors.initials: N
authors.lastname: Jiang
authors.forename: Xiaoshu
authors.initials: X
authors.lastname: Cheng
authors.forename: Xiaoyan
authors.initials: X
authors.lastname: Lai
authors.forename: Aminah
authors.initials: A
authors.lastname: Shunnar
authors.forename: Manpreet
authors.initials: M
authors.lastname: Singh
authors.forename: Neil
authors.initials: N
authors.lastname: Riordan
authors.forename: Vladimir
authors.initials: V
authors.lastname: Bogin
authors.forename: Nanwei
authors.initials: N
authors.lastname: Tong
authors.forename: Wei-Ping
authors.initials: WP
authors.lastname: Min
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000235:genetics*; D053148:Caspase 3 / Q000378:metabolism; D061945:Caspase Inhibitors; D003921:Diabetes Mellitus, Experimental / Q000201:enzymology / Q000473:pathology; D020868:Gene Silencing; D006943:Hyperglycemia / Q000473:pathology / Q000517:prevention & control*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009206:Myocardium / Q000201:enzymology / Q000473:pathology*; D019255:NADPH Oxidases / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D051197:Toll-Like Receptor 4 / Q000235:genetics* / Q000378:metabolism; D015854:Up-Regulation / Q000235:genetics
chemical_list: D061945:Caspase Inhibitors; D017382:Reactive Oxygen Species; C493487:Tlr4 protein, mouse; D051197:Toll-Like Receptor 4; D019255:NADPH Oxidases; D053148:Caspase 3