Effects Of Inducible Nitric Oxide Synthase Expression On Rat Cardiac Fibroblasts Proliferation And Collagen Synthesis And Its Possible Signal Transduction Pathway

Tangdu Hospital, the Fourth Military Medical University Xi'an 710038, ChinaThe heart is composed of myocytes and stroma. The interstitium collagen is a multilayer continuum with complex three dimensional spacial structure, and myocytes are packed in it. Myocytes have contractility, automaticity and conductivity, which are all necessary for heart to maintain blood circulation as a muscle pump. For many years the cardiac interstitium is been recognized only be responsible for scaffolding that supports muscle cells and blood vessels. So the importance of cardiac interstitium was less considerded. But in recent years, studies about cardiac interstitium found: (l)Adjoining myocytes are connected to one another with interstitium collagen, and collagen fibers insert into Z band of contractile protein. These connections not only prevent myocytes slippage, ensure myocytes length concordance during the cardiac cycle; but also serve to transduction of force between myocyets, coordinate cardiac function. (2)Interstitial collagen contents represents 3%~5% in normal myocardium, when it increases to 8%~12% ventricular diastolic dysfunction with impaired relaxation and increased resistance to filling appears. Left ventricular maximum filling velocity decreases and left ventricular filling pressure increases; Ventricular systolic dysfunction come forth when collagen contents go up to20%. Because myocytes were wrapped and sealed by excessive collagen eject fraction and cardiac output decline. (3)Interstitial fibroblasts proliferation and excessive fibrillar collagen synthesis participate in replacement scarring of necrotic myocardium after myocardial infarction and hypertensive left ventricular hypertrophy; Adverse accumulation and inappropriate degradation of fibrillar collagen get involved in idiopathic dilated cardiomyopathy, infarct expansion, ventricular rupture and aortic aneurysm. Therefore, matrix remodeling is pathologic basis of myocardial remodeling. We should study matrix remodeling carefully and take effective measures to inhibit or promote matrix remodeling, they may give us a possible way to improve cardiovascular function.In order to elucidate the relation of nitric oxide(NO) to matrix remodeling, the study investigate the effects of arginine vasopressin(AVP) on nitric oxide synthase-nitric oxide(NOS-NO) system activity in interstitial fibroblasts and its possible signal transduction pathway so as to provide valuable insight into mechanisms of the etiology and find a new way to treatment of matrix remodeling.Isolated and cultured cardiac fibroblasts(CFs) of neonatal Sprague-Dawley(SD) rats were used as experiment model. (1) Nitric acid reductase method were used to detect NO contents, spectrophotometry were used to determine NOS activity, Western-blotting were used to measure iNOS protein expression and reverse transduction-polymerase chain reaction(RT-PCR) were used to detect the iNOS mRNA expression(NOS-NO system activity) respectively with or without AW, and the intervening effects of IL-1 > TNF- a and Y -IFN on these aspects in CFs induced by AVP. (2) The effects of NOS-NO system activity in CFs on proliferation and collagen synthesis were measured by MTT technique and 3H-proline incorporation respectively. (3) Immunofluorescence-interactive laser cytometer techniques and western-blotting were adopted to estimated the activation and expression of NF- K B in CFs with or without AVP. Whether AVP receptors, protein kinase C and extracellular signal-regulated kinases are involved in enhancement of NOS-NO system activity were evaluated at the same time.The results shows: (1) NO contents, NOS activity, iNQS protein expressions and iNOS mRNA expressions all increase in a time-dependent manner without AVP. Moreover, NO contents(41.73 ?5.28 u mol/L), NOS activity(37.18?.52U/ml), iNOS protein expressions (1.47?.25) and iNOS mRNA expressions(0.29?.02) of 36h group were all higher than those of 6h group(14.11 ?.49 u mol/L, 10.26+4.18 U/ml, 0.8?.2, 0.1 ?.04) and those of 12h group(21.01?umoI/L,...