ABSTRACT
The Src homology domain 2 (SH2)-containing tyrosine phosphatase-1 (SHP-1) plays a key role in apoptosis and decreases phosphorylation of Akt. Apoptosis of cardiomyocytes is thought to contribute to the increased area of acute myocardial infarction (AMI), and Akt activation exerts a powerful cardioprotective effect after ischemia. Thus, a therapeutic strategy designed to inhibit expression of SHP-1 would be beneficial in AMI. Here we report that siRNA targeting SHP-1 reduced infarct size in a rat model of AMI. Upon injection into the ischemic left ventricular wall, the vector-based siRNA significantly suppressed the increase in the SHP-1 mRNA and the SHP-1 protein levels. The siRNA vector also significantly reduced the SHP-1 that bound to Fas-R. The SHP-1 siRNA vector increased phospho-Akt and reduced DNA fragmentation and caspase activity compared with the scramble siRNA vector. Finally, the area of myocardial infarction was significantly smaller with the SHP-1 siRNA vector than with the scramble siRNA vector at 2 days after LCA ligation. In conclusion, SHP-1 in the heart increased from the early stage of AMI, and this increase was thought to contribute to the increased area of myocardial infarction. Suppression of SHP-1 with the SHP-1 siRNA vector markedly reduced the infarct size in AMI.
Subject(s)
Intracellular Signaling Peptides and Proteins/physiology , Myocardial Infarction/metabolism , Myocardium/pathology , Myocytes, Cardiac/pathology , Protein Tyrosine Phosphatases/physiology , RNA Interference , Animals , Apoptosis , Blotting, Northern , Blotting, Western , Caspases/metabolism , Coronary Vessels/pathology , DNA Fragmentation , Echocardiography , Genetic Therapy , Genetic Vectors , Heart Ventricles , Immunoprecipitation , Male , Models, Biological , Myocardial Ischemia , Protein Phosphatase 1 , Protein Tyrosine Phosphatase, Non-Receptor Type 6 , RNA, Messenger/metabolism , RNA, Small Interfering/metabolism , Rats , Rats, Wistar , Time Factors , fas Receptor/metabolism , src Homology DomainsABSTRACT
Increased circulating and cardiac TNF-alpha levels during myocardial ischemia have been found in both experimental animals and patients with ischemic heart disease and advanced heart failure. Soluble TNF-alpha receptor 1 (sTNFR1) is an antagonist to TNF-alpha. In the present study, we examined whether sTNFR1 improves cardiac function in rats after myocardial infarction. Male Wistar rats were subjected to left coronary artery (LCA) ligation. Immediately after the ligation, a total of 200 microg of either the sTNFR1 or LacZ plasmid was injected into three different sites in the left ventricular wall. From 1 to 21 days after LCA ligation, TNF-alpha bioactivity in the heart was higher in rats receiving LacZ plasmid than in sham-operated rats, whereas sTNFR1 plasmid significantly suppressed the increase. The LV diastolic dimension was significantly lower, and the fractional shortening was significantly higher in rats treated with the sTNFR1 plasmid than in those treated with the LacZ plasmid. At 21 days after LCA ligation, the LV end-diastolic pressure was also significantly lower in the rats treated with the sTNFR1 plasmid. In addition, the sTNFR1 expression plasmid had significantly reduced the infarct size. In conclusion, TNF-alpha bioactivity in the heart increased during the early stage of infarction and remained elevated. This elevation seemed partially responsible for the impairment of LV function and the increased infarct size. Suppression of TNF-alpha bioactivity from the early stage of infarction with the sTNFR1 plasmid improved cardiac function and reduced infarct size.
Subject(s)
Antigens, CD/physiology , Genetic Therapy , Genetic Vectors/therapeutic use , Myocardial Infarction/therapy , Receptors, Tumor Necrosis Factor/physiology , Tumor Necrosis Factor-alpha/antagonists & inhibitors , Animals , Antigens, CD/genetics , Apoptosis , Coronary Vessels , Drug Evaluation, Preclinical , Genetic Vectors/administration & dosage , Heart Ventricles/diagnostic imaging , Injections, Intralesional , Ligation , Male , Myocardial Infarction/diagnostic imaging , Myocardial Infarction/pathology , Myocardial Infarction/physiopathology , Myocardium/pathology , Protein Structure, Tertiary , RNA, Messenger/biosynthesis , Rats , Rats, Wistar , Receptors, Tumor Necrosis Factor/genetics , Receptors, Tumor Necrosis Factor, Type I , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/physiology , Reverse Transcriptase Polymerase Chain Reaction , Solubility , Transfection , Ultrasonography , Ventricular Function, LeftABSTRACT
Apoptosis in the myocardium is linked to ischemia/reperfusion injury, and TNF-alpha induces apoptosis in cardiomyocytes. A significant amount of TNF-alpha is detected after ischemia and reperfusion. Soluble TNF-alpha receptor 1 (sTNFR1) is an extracellular domain of TNF-alpha receptor 1 and is an antagonist to TNF-alpha. In the present study, we examined the effects of sTNFR1 on infarct size in acute myocardial infarction (AMI) following ischemia/reperfusion. Male Wistar rats were subjected to left coronary artery (LCA) ligation. After 30 min of LCA occlusion, the temporary ligature on the LCA was released and blood flow was restored. Immediately after reperfusion, a total of 200 microg of sTNFR1 or LacZ plasmid was injected into three different sites of the left ventricular wall. At 6 h, 1 and 2 days after reperfusion, the TNF-alpha bioactivity in the myocardium was significantly higher in rats receiving LacZ plasmid than in sham-operated rats, whereas sTNFR1 plasmid significantly suppressed the increase in the TNF-alpha bioactivity. The sTNFR1 plasmid significantly reduced DNA fragmentation and caspase activity compared to the LacZ plasmid. Finally, the sTNFR1 expression-plasmid treatment significantly reduced the area of myocardial infarction at 2 days after ischemia/reperfusion compared to LacZ plasmid. In conclusion, the TNF-alpha bioactivity in the heart increased from the early stage of ischemia/reperfusion, and this increase was thought to contribute in part to the increased area of myocardial infarction. Suppression of TNF-alpha bioactivity with the sTNFR1 plasmid reduced the infarct size in AMI following ischemia and reperfusion.
Subject(s)
Apoptosis , Genetic Therapy , Myocardial Infarction/therapy , Myocardial Reperfusion Injury/therapy , Plasmids/administration & dosage , Receptors, Tumor Necrosis Factor, Type I/genetics , Animals , Caspases/metabolism , DNA Fragmentation , Male , Myocardial Infarction/metabolism , Myocardial Infarction/pathology , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/pathology , Plasmids/genetics , Protein Structure, Tertiary/genetics , Rats , Rats, Wistar , Tumor Necrosis Factor-alpha/antagonists & inhibitors , Tumor Necrosis Factor-alpha/metabolismABSTRACT
Apoptosis is the major independent form of cardiomyocyte cell death in acute myocardial infarction (AMI). TNF-alpha release early in the course of AMI contributes to myocardial injury, and TNF-alpha induces apoptosis in cardiomyocytes. Soluble TNF-alpha receptor 1 (sTNFR1) is an antagonist to TNF-alpha. However, the effect of sTNFR1 on AMI remains unclear. Here we report that direct injection of an sTNFR1 expression plasmid DNA to the myocardium reduces infarct size in experimental rat AMI. Treatment with sTNFR1 expression plasmid DNA reduced the TNF-alpha bioactivity in the myocardium and the apoptosis of cardiomyocytes. These findings suggest that the anti-TNF-alpha therapy by sTNFR1 can be a new strategy for treatment of AMI.