Effects Of Atorvastatin On The Expression Of Cx40 In Atrial Myocytes And MMP-9/TIMP-1 In Cardiac Fibroblasts

Objective:1.To explore the Primary Cultured method for atrial myocytes and cardiac fibroblasts;2. To investigate the effect of atorvastatin on AngiotensinⅡ-induced atrial myocytes hypertrophy and proliferation of cardiac fibroblasts and thecorrelation studies of MMP-9/TIMP-1;3.To investigate the mechanism of preventing and curing AF and the correlation studies of connexin 40(Cx40).Method:1. Methods atrial myocytes and cardiac fibroblasts of neonate rats culture and identification:A total of 18 Wistar rats of half gender,weighing from 35 to 40 grams and aged about one week were used for culturing primary atrial myocytes and cardiac fibroblasts by differential adhesion separation technology and mixed enzyme digestion methods,while atrial myocytes specific expressed a-sarcomeric actin and cardiac fibroblasts specific expressed protein vimentin was detected with immunofluorescence histochemistry;2. Atorvastatin on the hypertrophy of atrial myocytes and proliferation of cardiac fibroblasts induced by AngⅡand the effects of MMP-9/TIMP-1,Cx40 expression: The cells were devided into six groups:normal control, AngⅡ(10-6μmol/L), AngⅡ+ dimethyl sulfoxide (DMSO) and AngⅡ+ atorvastatin (0.1μmol/L, 1.0μmol/L, 10μmol/L).After 72h, to detection cell proliferation by MTT assay; MMP-9 activity was measured by gelatinase zymograph, the synthetic rate of protein in atrial myocytes was detected by [3H]-leucine incorporation.MRNA expression of natriuretic peptide precursor B (Nppb) which is the precursor of brain natriuretic peptide (BNP) and MMP-9/TIMP-1,transforming growth factor-β1 (TGF-β1),Cx40 was measured by reverse transcription-polymerase chain reaction (RT-PCR) MMP-9 and TIMP-1 mRNA expressions were detected by RT-PCR.Result:Compared with normal control group, AngⅡcan induce the proliferation of CF(p<0.01). It also showed AngⅡsignificantly increased MMP-9 activities. The mRNA expressions of MMP-9 was also significantly increased (P<0.01)while decreasing the expression of TIMP-1,3H-leucine incorporation and mRNA expression of Nppb (P<0.05) increased significantly in AngⅡgroup, and mRNA expression of TGF-β1increased while mRNA expression of Cx40 decreased. Atorvastatin can revers thoses changges,and the efect of 10μmol/L group was the most obvious (P<0. 05), but DMSO which as the solvent has no effects (P>0.05).Conclusions:1. we succeeded to obtain primary atrial myocytes and cardiac fibroblasts;2. Atorvastatin has the action of inhibition the proliferation of CF, and consequently inhabit the activity and mRNA expression of MMP-9 and enhance the one of TIMP-1, and adjust the balance of MMP-9/TIMP-1; Atorvastatin inhibits hypertrophy of atrial myocytes induced by angiotensinⅡby inhibiting the increase of mRNA expression of Nppb, and inhibiting the increase of mRNA expression of TGF-β1 to decrease atrial fribosis,while maybe reverse the remoding of connexin by increase of mRNA expression of Cx40.