The Role Of G-CSF Mediated CXCR4 And Non-CXCR4 Axis In Post-ischemic Arrhythmogenesis

Background Myocardial infarction is the leading cause of morbidity and mortality in cardiovascular diseases.It can cause arrhythmogenic remodeling,such as mechanical remodeling and electrical remodeling.Recently,stem cells mobilization by cytokines has been involved in myocardium repair.Granulocyte colony-stimulating factor(G-CSF),as an agent of bone marrow stem cells mobilization,can improve cardiac function through mobilization and recruitment of bone marrow CXCR4+ stem cells.Moreover,researchers have found G-CSF can exert non-genomic effects and anti-apoptosis effects that associated with long-term gene expression on cardiac function,via binding with G-CSF receptor(G-CSFR)of cardiomyocyte membrane.Taken together,the specific mechanisms of G-CSF can be concluded as effects of bone marrow mobilization(CXCR4 pathway)and direct function on targeted cells(non-CXCR4 pathway).Up to date,how G-CSF changes the post-ischemic ventricular myocytes' electrophysiological characteristics and reduces arrhythmia via the non-CXCR4 pathway,is not reported.Objective In the present study,the effects of acute G-CSF of various concentrations intervention on ventricular myocytes' ion channels in ischemic condition were observed.G-CSF alone or combined with antagnist of CXCR4 receptor(AMD3100)intervention on ischemic models in Wistar rats were investigate to elucidate Whether G-CSF can improve the electrophysiological characteristics in acute phase of myocardial ischemia,even the beneficial effects exist persistently in chronic phase.Methods We used cellular etectrophysiology and integrity electrophysiology methods to perform the experiments.Wistar rats ventricular myocytes were obtained by enzymolysis.Whole-cell patch clamp was used to gain theⅠ-Ⅴcurve,availability curve and activity curve of L-type calcium channel current(I_Ca.L)and voltage-dependent sodium channel current (I_Na),with or without G-CSF in various concentrations.Wistar rat MI models were established by undergoing left anterior descending artery(LAD)ligation.In the I/R heart models,the hearts were subjected to 30 min global ischemia followed by 120 min reperfusion.Operation and no ligation of rat models were in Sham group.The rat models were randomly divided into G-CSF group,G-CSF+AMD3100 group and saline group,each group contained two categories:7 days and 3 months.Keeping beating with continuous perfusion with a Working heart system,the effects of G-CSF in various concentrations on the sinus cardiac cycle(SCL),effective refractory period(ERP),monophasic action potential(MAP)of the ventricle and threshold value of ventricular fibrillation(VF) were recorded by a multi-channel electrophysiolograph.Results1.TheⅠ-Ⅴcurves of I_Ca.Lwere changed by acute G-CSF intervention in a dose-dependent way.G-CSF did not change the activity curve or availability curve of I_Ca.Lexcept the former in an 300μg/kg dose,that indicated the effects of G-CSF on I_Ca.Lwere in non-voltage-dependent way.TheⅠ-Ⅴcurves of I_Ca.Lin ischemic condition were gradually approaching the levels of the control group with the higher of G-CSF dose.The effects of 300μg/kg G-CSF on I_Ca.Lsimilar to 100μg/kg G-CSF indicated that 100μg/kg is the optimal dose of G-CSF for the acute intervention.2.Acute G-CSF intervention of maximum dose(300μg/kg)did not change theⅠ-Ⅴcurve,activity curve,availability curve or resting availability curve of I_Na.3.MI Group:The difference of electrophysiological parameters between instant G-CSF intervention group and MI-saline group was significant(P＜0.05),indicated that instant G-CSF intervention could improve the electrophysiological characteristics of MI models.The electrophysiological parameters of delay G-CSF intervention group,however,were similar to that of MI-saline group with no statistical significance.Compared with delay G-CSF intervention group,the electrophysiological parameters of instant G-CSF intervention group were improved significantly(P＜0.05).The electrophysiological parameters of instant G-CSF+AMD3100 group and delay G-CSF+AMD3100 group were better than that of MI-saline group,whereas both the former two group compared with Sham group were worse.It had a significant difference among the groups(P＜0.05).The electrophysiological parameters in MI group were improved persistently from the 7 days to 3 months,but all were not approching the levels of Sham group with statistical significant difference(P＜0.05).I/R Group:The electrophysiological parameters of instant G-CSF intervention group and delay G-CSF intervention group were better than that of I/R-saline group with significant difference(P＜0.05).The former two groups did not reach the levels of Sham group,however,it had a significant difference among groups(P＜0.05). Compared with the Sham Group,instant G-CSF+AMD3100 group and delay G-CSF+AMD3100 group had the similar results with no significant difference.The electrophysiological parameters in I/R group were improved persistently from the 7 days to 3 months,but all were not approching the levels of Sham group with statistical significant difference(P＜0.05).Conclusions1.L type calcium channel current of ischemic ventricular myocytes in rats can be affected by acute G-CSF intervention in a dose-dependent way when the dose ranging from 0μg/kg to 300μg/kg.Compared to the dose of 100μg/kg,the 300μg/kg G-CSF intervention has the similar effects on L type calcium channel current, indicates that 100μg/kg is the optimal dose for acute C-CSF intervention.2.The parameters of Na channel current are not changed compared with control group in the condition of maximum dose(300μg/kg)G-CSF intervention,indicates that Na channel current can not be affected by acute G-CSF intervention.From the view of integrity electrophysiology,the upstroke of action potential is not changed.3.The electrophysiological parameters can be improved by most groups of intervention in MI models and I/R models.The results of instant intervention group are better than delay intervention group,whereas a few of delay intervention groups has no beneficial effects,even has a worse potential.The beneficial effects on the electrophysiological parameters exist persistently from the early stage to late stage of ischemic myocardium.Combined G-CSF with AMD3100 has a tendency to be better effects than G-CSF alone.