Effect Of Profilin-1on Myocardial Lesions In Rats Induced By AGEs

BackgroundDiabetic cardiomyopathy (DCM) is a major complication of diabetes, leading death rate of diabetes. Now that the occurrence and development of DM is mainly closely related to increased production of AGEs;AGEs can cause oxidative stress endothelial dysfunction, tissue and reconstruction through direct or indirect pathways;AGEs/RAGE signaling pathway is one of the most important signaling pathways, which suggest that it may be an important therapeutic target for prevention and treatment of DCM. Profilin-1, a key regulator of actin protein, expressing relatively specific in Cardiovascular tissues,can directly affect the cytoskeleton; nevertheless changes in the cytoskeleton may be a key contributing factor of myocardial hypertrophy and myocardial injury. Rho/ROCK/profilin-1mediates actin polymerization, resulting in cytoskeletal remodeling, suggesting that this pathway may play a role in DCM.Related research about profilin-1and DCM can not be found currently. This study intends to establish a rat model of myocardial injury induced by AGEs to observe changes in cardiac morphology and function, myocardial fibrosis, myocardial damage and changes in the expression of various cytokines and protein profilin-1; application shRNA adenoviral PROFILIN-1gene silenced technology for observing the impact of the intervention in cardiomyopathy induced by AGEs is carried out to directly demonstrate the pivotal role of profilin-1in AGEs-induced myocardial injury. This research aims to explore new targets to the prevention and treatment of diabetic cardiomyopathy and provide a theoretical basis for development of new drugs.MethodPreparation of AGEs; building up PROFILIN-1gene silenced shRNA adenovirus vector. SD rats were randomly divided into:normal control group (control group):continuously intravenous injection of saline; AGEs groups:continuous intravenous injection of AGEs (25mg/kg); AGEs+shProfilin-1group:intermittently apical injection of PFN1shRNA and continuous intravenous injection of AGEs-BSA (25mg/kg); empty virus group (empty virus control group):intermittently apical injection of adenovirus vectors without gene silenced. ECG is underwent after8weeks; rapid test method is used to measure fasting glucose; rats were sacrificed for measurement of serum ICAM-1、 ADMA、MDA levels; myocardium is observed by HE staining; myocardial immunohistochemistry is utilized to test expression of protein profilin-1; myocardial protein is extracted to detect protein expression of profilin-1、Rho、NF-κB by Western Blot; myocardial tissue RNA is extracted to detect relative expression of profilin-1mRNA、MMP-2mRNA、MMP-9mRNA by RT-PCR.Results(1) The validation results of cell infected by adenovirus interfered by PFN1shRNA show the efficiency of the adenoviral vector; The PFN1shRNA adenoviral vector was constructed successfully.(2) electrocardiogram T-wave decline is noteblely at AGEs group contrasting to control group and there was statistically significant, P<0.05.(3) myocardium HE staining display (400x):Compared to the control group AGEs group myocyte disarray, cytoplasmic turbid; nucleus increases, nuclear stained portion of a nuclear cardiomyocytes edge set; myocardial interstitial fibers increased, disorganized, cell gap filling, showing inflammatory cell infiltration. contrasting to AGEs group, shProfilin-1group with cells arranged more regularly, no significant edema, the nucleus is clear, no significant myocardial interstitial hyperplasia and inflammatory cell infiltration.(4) myocardial immunohistochemistry (400x) shows AGEs group compared with the control group, profilin-1highly expression,P<0.01; shProfilin-1group compared with AGEs group profilin-1was significantly lower,P<0.01(5) AGEs group compared with the control group, a significantly increase in the expression of protein Profilin-1, Rho and NF-κB,P<0.05; shProfilin-1group compared with AGEs group, expression of protein Profilin-1, Rho and NF-κB was significantly decrease,P<0.05(6) AGEs group compared with the control group, the expression of Profilin-1mRNA and MMP-2mRNA in myocardium was significantly increased, respectively P<0.01, P<0.05; shProfilin-1group compared with AGEs group, a significant reduction in the expression of Profilin-1mRNA and MMP-2mRNA in myocardium; respectively, P<0.01, P<0.05; no significant difference among the expression of MMP-9mRNA in each group, P>0.05(7) AGEs group compared to the control group, the level of serum ICAM-1、ADMA、 MDA increase significantly,P<0.05; shProfilin-1group compared with control group, the level of serum ICAM-1、ADMA、MDA significantly reduced,P<0.05.Conclusion(1) Profilin-1mediated AGEs-induced myocardial injury and remodeling; its mechanism involve oxidative myocardial cell injury, inflammation and endothelial dysfunction.(2) Rho/ROCK signaling pathway may involve in Profilin-1mediated heart disease.