| Atrial fibrillation(AF)is the common type of arrhythmias.During the past decades,its incidence rate has increased by 33%and the number of patients has reached 43.6 million worldwide.Worse of all,these upward trends are still expected to grow in the next 30 years.Currently,the treatment of AF concentrates on symptomatic treatment,including drugs and surgery to control heart rate and antithrombosis.The in-depth study on the pathological mechanism of AF is of great significance for improving clinical prognosis of AF.Current studies believe that the atrial remodeling is the main pathogenesis of AF,which is characterized by atrial enlargement,deformation,and fibrosis.Atrial fibrosis is the major pathological factor of AF,as well as the basic pathological feature of atrial remodeling.On the one hand,continuous fibrous changes in atrial tissue can lead to changes in local conduction velocity and cell refractory,resulting in conduction block and promoting the formation of micro reversion and unstable waves.On the other hand,AF facilitates the formation and development of atrial fibrosis due to the tissue hypoxia,inflammation,oxidative stress,and mechanical stress.Atrial fibrosis has been found to associate with thromboembolic diseases and mortality.Therefore,atrial fibrosis is an important target for the treatment of AF,and anti-fibrosis should be emphasized in the future.AF belongs to the category of heart palpitation in traditional Chinese medicine(TCM),which occurs in the heart.The pathogenesis of AF relates to the blood stasis.Fibrosis is similar to "stasis" and "mass" in TCM.Activating blood and removing stasis is an important method in the TCM treatment of fibrosis in AF.Leech is a traditional Chinese medicine,which has the function of promoting blood circulation and removing blood stasis.It is especially good at treating blood stasis caused by long illness without hurting qi.Modern pharmacological studies have proved that leeches exert the fuction of inhibiting inflammation,fighting thrombosis and fibrosis.Mai Xue Kang capsule and Mai Xue Kang enteric-coated tablet,commonly used leech preparations in clinical practice,are Chinese patent medicines that made from leech by freezing,drying and extraction.Previous studies have found that leeches can resist fibrosis of liver,renal and pulmonary,but there is no evidence of atrial fibrosis.Our previous meta-analysis showed that Mai xue kang combined with conventional treatment significantly reduce the frequency of AF attacks and decrease thrombosis and bleeding events.However,it is unclear whether the mechanism of its treatment of AF is related to the inhibition of atrial fibrosis.Based on this,we put forward a hypothesis:"blood stasis syndrome is the main TCM pathological factor for the occurrence and development of AF,and leech preparation can prevent atrial fibrosis of AF".According to this hypothesis,we performed two studies:(1)by reviewing the medical records of AF patients hospitalized in Xiyuan Hospital in recent five years,we conducted telephone follow-up of clinical outcomes to analyze the risk factors affecting the long-term prognosis of AF patients;(2)Mai xue kang enteric coated tablet was used to intervene AF rats.The effect and mechanism of Mai xue kang enteric coated tablet on atrial fibrosis in AF rats were studied by proteomics.Part 1 A retrospective study on the prognostic factors of 282 patients with non-valvular atrial fibrillationObjective:To analyze the factors affecting the poor prognosis of all-cause mortality and stroke in patients with non-valvular AF retrospectively to provide evidence for the diagnosis and treatment of AF.Methods:The medical records of patients with non-valvular AF hospitalized at Xiyuan Hospital from November 1,2016 to November 1,2021 were analyzed retrospectively and followed up by telephone.The morbidity,risk factors,treatment and adverse events(all-cause mortality and stroke)of the patients were analyzed.Risk factors for all-cause mortality and stroke events in patients during follow-up were analyzed by logistic regression,COX regression,and Kaplan Miere survival analysis.Results:A total of 282 patients with NVAF were included in this study,and the Median followup time was 45 months.The results are as follows:1.During the follow-up period,a total of 44 patients(15.6%)experienced all-cause death and stroke compound events.All-cause death occurred in 30 patients(10.6%)and stroke in 14 patients(5.0%).2.Patients with increased left atrial diameter(LAD)(HR:1.095,95%CI:1.034-1.160),blood stasis syndrome(HR:2.834,95%CI:1.135-7.074),and high-sensitivity C-reactive protein level(hs-CRP)(HR:1.014,95%CI:1.000-1.027)showed significantly higher incidence of all-cause death and stroke composite endpoint events during patient follow-up.3.Heart failure(HR:3.571,95%CI:1.354-9.419),increased LAD(HR:1.085,95%CI:1.024-1.148),and blood stasis(HR:3.072,95%CI:1.1358.311)significantly increased the incidence of all-cause mortality endpoint events.4.Patient with previous stroke(HR:7.306,95%CI:1.769-30.176),larger LAD(HR:1.179,95%CI:1.037-1.340),and increased of Fbg(HR:1.494,95%CI:1.172-1.905)demonstrated higher incidence of stroke during follow-up.5.Patients treated with anticoagulants had a significantly lower incidence of all-cause mortality and stroke composite endpoint events than patients that were not treated.(Log Rank=10.915,P=0.004).6.Female(OR:0.360,95%CI:0.168-0.774)was protective factor for enlarged LAD,elevated pro-BNP level(OR:1.001,95%CI:1.0001.002)and blood stasis syndrome(OR:3.991,95%CI:1.982-8.040)were the risk factors for increased LAD.Conclusions:Enlarged LAD,blood stasis and hs-CRP levels significantly increased the incidence of all-cause mortality and stroke composite endpoint during follow-up.Heart failure,LAD enlargement and blood stasis significantly increased the incidence of all-cause mortality endpoint events during follow-up.History of stroke,increased LAD,and increased Fbg levels showed significantly higher stroke endpoint events during follow-up.Female was protective factor for enlarged LAD,elevated pro-BNP level and blood stasis syndrome were the risk factors for the increase of LAD.Part 2 Effect and mechanism of Mai Xue Kang enteric coated tablet on atrial fibrosis in rats with atrial fibrillationObjective:To study the effects and mechanisms of Mai Xue Kang Enteric-coated Tablet on atrial fibrosis in atrial fibrillation model rats.Methods:AF rats were randomly divided into AF model group(Model),Mai Xue Kang Enteric-coated Tablet group(MXK),and dabigatran capsule group(DBJQ),20 rats in each group.Control group of 20 animals was also set up.After 24 h of modeling,each group was given the corresponding drugs by gavage,MXK group was given Mai Xue Kang Enteric-coated Tablet(378 mg/kg/d)for 6 weeks;DBJQ group was given dabigatran(27 mg/kg/d)for 6 weeks;Model and Control groups were given distilled water gavage for 6 weeks.Atrial tissue fibrosis was evaluated by HE staining,MASSON staining,Sirius red staining and immunofluorescence.Inflammatory factors,coagulation function and clotting factors were measured by ELISA in each group of rats,and fibrotic proteins were measured by Western Blot in each group.Quantitative proteomics was used to identify the differentially expressed proteins in the atrial tissues of rats in Model group,MXK group and Control group,and the differential proteins were analyzed combined with bioinformatics methods.qRT-PCR.Western Blot and ELISA were used to verify the results of proteomics research.Results:Compared with the Control group,the LAD of the rats in the Model group was significantly increased(P<0.05);compared with the model group,the LAD of the rats in the MXK group and DBJQ group was significantly decreased(P<0.01,P<0.05).Compared with the control group,the left ventricular end diastolic diameter(LVEDD),left ventricular endsystolic(LVESD),left ventricular diastolic volume(LVVd),and left ventricular systolic volume(LVVS)of rats in the model group increased(P<0.05).Compared with the model group,the LVEDD,LVVd and LVVS of the MXK group were significantly lower than those of the model group(P<0.01),and the LVESD of the MXK group was smaller than that of the model group(P<0.05).Compared with the control group,the atrial collagen volume index of the rats in the model group was significantly increased(P<0.05),and the contents of fibrosis-related proteins COL-Ⅰ,COL-Ⅲ,αSMA and TGFβ1 were significantly increased(P<0.01).Compared with the model group,the collagen volume index and the level of fibrosis-related proteins COL-I,COL-Ⅲ,αSMA and TGFβ1 in the MXK and DBJQ group were significantly decreased(P<0.01).MXK group exerted better effect than DBJQ group(P<0.05).Compared with the control group,the inflammatory factors interleukin-6(IL-6),high-sensitivity C-reactive protein(hsCRP),and monocyte chemoattractant protein-1(MCP-1)levels in the model group were significantly increased(P<0.01).Compared with the Model group,MXK reduced the levels of IL-6,hs-CRP and MCP-1(P<0.05).Compared with the control group,the prothrombin time(PT)and activated partial thromboplastin time(APTT)of the rats in the model group were shortened,and the level of fibrinogen(FIB)was increased(P<0.05),APTT and PT of the rats in the MXK group and DBJQ group were significantly prolonged and the content of FIB was decreased(P<0.05).Compared with DBJQ group,MXK group significantly prolonged the PT and APTT(P<0.05),but there was no significant difference in reducing FIB content(P>0.05).Compared with the control group,the levels of prothrombin(TM)and factor Xa(FXa)in the model group were significantly increased(P<0.01),and the intervention of MXK and DBJQ reduced the level of AF in the rats.TM and FXa levels(P<0.05).There was no significant difference between the MXK group and the DBJQ group in reducing the levels of TM and FXa(P>0.05).Atrial tissue proteomics identified 4766 differential proteins in model group,MXK group and Control group.Bioinformatics analysis of the differential proteins showed that.compared with the Control group,IP3R,NLRP3,ASC,Capase-1,pro-Capase-1,IL-18,IL-1βof the NOD signaling pathway in model group were significantly up-regulated(P<0.05);compared with the model group,the MXK group down-regulated the expression of the proteins mentioned above in the NOD signaling pathway(P<0.05).The IP3R/NLRP3 signaling pathway proteins were verified by Western Blot,ELISA,and qRT-PCR methods,and the results showed that compared with the Control group,the level of IP3R,NLRP3,ASC,capase-1,pro capase-1,IL-18 and IL-1β in the atrial tissue of Model group was up-regulated(P<0.01),and MXK decreased the expression of the above proteins(P<0.01).Conclusion:Compared with the Model group,MXK significantly ameliorated atrial tissue fibrosis and atrial remodeling in AF rats and inhibited the expression of inflammatory response and thrombosis-related factors.The underlying mechanism of MXK in inhibiting atrial fibrosis in AF rats may related to regulating the IP3R/NLRP3 inflammatory response signaling pathway. |
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