Effects Of Probucol On Atrial Remodeling And Inducibility Of Atrial Fibrillation In Alloxan-induced Diabetic Rabbits

Objective Diabetes mellitus (DM) is an independent risk factor for atrial fibrillation (AF) and has been recognized as a low-grade inflammation disease. Inflammation has also been shown to have a direct role in the initiation, maintenance, and recurrence of AF. On the other hand, oxidative stress play critical role in the pathophysiology of DM and the development of vascular complications, which may also relate to the development of AF. So we hypotheses that inflammation and oxidative stress may be important mechanisms for the increased propensity for AF in DM. The purpose of this study was to elucidate the underlying mechanisms of AF and to evaluate the effects of probucol on atrial electrophysiological changes and AF promotion in alloxan-induced diabetic rabbits.Methods The experiment was divided into three parts. In each part,40Japanese rabbits were randomly assigned to a normal control group (C, n=10), a alloxan-induced diabetic group (DM, n=10), probucol-treated group (CPR, n=10) and probucol-treated diabetic group (DPR, n=10). In the DM and DPR group, alloxan monohydrate was immediately administered intravenously via the marginal ear vein. Rabbits in the DPR and CPR groups were orally administered Probucol (1000mg/day) for8weeks. The animals in four groups were housed in cages and a standard laboratory pellet diet for8weeks. In part one, isolated Langendorff perfused rabbit hearts were prepared to evaluate atrial refractory effective period (AERP) and its dispersion (AERPD), interatrial conduction time (IACT) and vulnerability to AF. Plasma malonaldehyde (MDA), superoxide dismutase (SOD), TNF-a, myeloperoxidase(MPO) and catalase (CAT) levels were measured by chemical colourimetric methods. Sirius-Red staining was used to evaluate atrial fibrosis. In part two, using whole-cell voltage-clamp techniques,Ica,L,INa and action potential(AP) were measured in left atrial myocytes isolated from the rabbits of four groups. In part three, the expression levels of Cavl.2, NF-κB, ERK, P-ERK, TGF-β and HSP70in left atrial tissue were analysed by western blot, the expression levels of TNF-a and TLR4were analysed by RT-PCR methods.Results IACT and AERPD were prolonged in diabetic rabbits compared with controls. Inducibility of AF in diabetic group was significant higher than controls (8/10vs1/10, P<0.05). Extensive interstitial fibrosis was observed in the DM group (P<0.05). Plasma MDA and TNF-a were increased significantly in the DM group (P<0.05). The density of Ica,L current in diabetic myocytes was significantly higher than that in the control, the density of INA current was significantly decreased in diabetic myocytes. APD90and APD50were also prolonged in diabetic myocytes. NF-κB, ERK, P-ERK, TGF-β and HSP70protein and the levels of Cavl.2, TNF-a mRNA and TLR4mRNA expression were significantly upregulated in DM group. The DPR rabbits exhibited significant alleviation of oxidative stress displayed as decreased plasma MDA and TNF-a compared with diabetic rabbits (P<0.05), probucol administration increases stability of vulnerable atrial fibrillation in diabetic rabbits (P<0.05). Histological analysis revealed suppression of DM-related histological changes (interstitial fibrosis) by probucol. The density of iCa,L current was significantly decreased and the density of INa current was significantly increased in DPR myocytes. Probucol significantly downregulated atrial NF-κB, TGF-β and HSP70protein expression and TNF-a mRNA and TLR4mRNA expression in DPR left atrial tissue.Conclusion There is an increased NF-κB activity in diabetes mellitus. Some of target cytokines NF-κB controls and stimulus of NF-κB like TGF-β, TLR4are responsible for inflammation, atrial fibrosis and hypertrophy of atrial tissue, which may serve as an important substrate for the development of AF. Alterations of atrial Ica,L and INa were due to the increased oxidative stress, which lead to the unstablity of electrical activity and subsequent occurrence of AF. Probucol through its powerful anti-inflammatory and antioxidant activity preserves the structural integrity of atrial tissues and increases stability of vulnerable atrial fibrillation in alloxan-induced diabetic rabbits.