| Ventricular remodeling is a common pathway for the development of various organic heart disease to heart failure.Ventricular remodeling is a progressive pathophysiological process that continues throughout the development of heart failure.The characteristic change is abnormal heart geometry,myocardial hypertrophy accompanied by dysfunction,decreased number,increased cardiac interstitial cells,accompanied by decreased cardiac function.Hypertension,coronary heart disease,cardiomyopathy,valvular disease,stress response,and chemicals can cause ventricular remodeling.Among them,hypertension and coronary heart disease are the most common causes.Ventricular remodeling is an important factor in the progression of hypertension and coronary heart disease.Inhibition of ventricular remodeling is a key factor in delaying the progression of heart failure and improving the prognosis of patients.There are many basic and clinical studies on ventricular remodeling,but the mechanism is still not completely clear.Although there are many treatments,there is still no specific treatment that can inhibit the reversal of ventricular remodeling.It can be seen that this pathophysiological process itself is extremely complicated,and various factors participate in its development,including: ischemia,pressure compliance,inflammatory response,renin-angiotensin-aldosterone system,immune system,cytokines,apoptosis,and Cell signaling pathway regulation,etc.How to rationally use drugs to inhibit ventricular remodeling,improve heart function,reduce the incidence of heart failure and mortality,and maximize the prognosis,has significant clinical and social benefits.Aquaporins(AQPs)have been found for 30 years since their discovery.A total of 13 AQPs(AQP0~AQP12)have been expressed on mammalian cell membranes.Their role is to transport water molecules across membranes under osmotic pressure gradients.Coordinate water translocation across cells.The study found that AQP1 is most expressed in the AQPS family in human and mouse myocardial tissues.Moreover,AQP1 in myocardial tissue is mainly expressed on vascular endothelial cells and myocardial cell membranes,and under physiological action,it plays a role in coordinating the transmembrane transport of water molecules,and maintains the balance of water between tissue cells.Under pathological conditions,after MI/RI,AQP1 also plays an important role in myocardial cell edema and microvascular permeability after myocardial ischemia.When MI/RI occurs,the blood supply to myocardial tissue in the lesion area decreases or disappears.In addition,it causes insufficient oxygen supply to the heart,insufficient myocardial energy acquisition,and increased release of toxic metabolites,resulting in abnormal function of myocardial cells.In the early stage of ischemia-reperfusion,due to the release of a large number of cytokines and inflammatory mediators caused by abnormal ischemia and hypoxia metabolism,this will lead to an increase in vascular permeability and myocardial interstitial edema.Due to the abnormal metabolism of oxygen supply,local lactic acid accumulation causes cytotoxic side effects,which affect cell membrane permeability and cause myocardial cell edema.A study on burn-induced myocardial edema in rats found that burns caused significant edema in rat myocardial tissue.In addition,the expression of AQP1 protein in rat myocardial tissue was also significantly increased.The degree of myocardial edema and AQP1 protein were found.The expression was positively correlated,indicating that AQP1 may be involved in the pathogenesis of myocardial edema induced by ischemia and hypoxia in myocardial tissue after burn.Myocardial edema itself is an important pathophysiological process.It will aggravate tissue ischemia and hypoxia in the early stage of ischemia.The two interact to form a vicious circle.In addition,it will aggravate the rigidity of the left ventricle and reduce its compliance.Sex,it has been reported that only 3.5% increase in intramyocardial water will reduce cardiac output by 40%.Uneven distribution of intraventricular edema can lead to repolarization,triggering malignant arrhythmia and increasing mortality.Increased myocardial interstitial edema can compress capillaries and aggravate myocardial ischemia and hypoxia.When the duration of myocardial edema is further prolonged,it will cause myocardial structural changes.The most important thing is to cause myocardial fibrosis and accelerate left ventricular remodeling,which may lead to cardiac dysfunction and affect the long-term prognosis and quality of life of patients.In addition,studies have shown that Hg2+ reduces cell edema by inhibiting AQPs,reduces mitochondrial membrane permeability and inhibits apoptosis,indicating that AQPs play a role in apoptosis.In addition,several studies have found that down-regulation of AQP1 expression can reduce cell edema and reduce apoptosis,but no report of AQP1 on cardiomyocyte apoptosis.Therefore,under the condition of ischemia and pressure overload,the expression of AQP1 in myocardial tissue and its influencing factors have important clinical significance,which provides a new treatment idea for inhibiting ventricular remodeling.The key point of this study is to find a practical AQP1 inhibitor.A large number of studies have reported that AQPs inhibitors,blockers,various hormones,physical and chemical factors and active protein molecules have a regulatory effect on AQP1 protein expression.The influencing factors involved in the study include hypertonic environment,hypoxic state,hormone level,phosphorylation induction,mercury ion,acetazolamide,RNA interference factor,etc.Many of the above factors are difficult to operate or have toxic side effects.For clinical treatment,Acetazolamide(AZA),as a widely used drug,has potential promotion and application value in the treatment of ischemia-reperfusion injury.Acetazolamide is a carbonic anhydrase inhibitor that has been studied in kidney tissue and found to inhibit water transport in AQP1 and reduce renal tubular water permeability.In vitro cell culture studies also found that the addition of AZA significantly reduced the expression of AQP1 mRNA and protein in Schwann cells induced by hypoxia,indicating that AZA can significantly inhibit the expression of AQP1.A study on AZA has shown that it can reversibly inhibit the permeability of AQP1 in Xenopus oocytes and reduce the expression of AQP1 in tumor tissues to reduce tumor invasion and metastasis.However,the effect of AZA on the expression of AQP1 in cardiomyocytes has not been reported.Previous studies on inflammatory factors have found that many heart diseases cause increased expression of inflammatory cytokines such as tumor necrosis factor(TNF-?),interleukin-6(IL-6),and these factors in disease.It plays an important role in the pathological process of the outcome,and the inflammatory response can accelerate the ventricular remodeling caused by cardiac hypertrophy.Cardiomyocytes intervened by IL-6 can cause increased protein synthesis in cardiomyocytes,and some protein factors that cause cardiac hypertrophy will also increase,such as p-myosin heavy chain(P-MHC).Overexpression of TNF-? transgenic mice found that elevated TNF-? not only leads to cardiomyocyte hypertrophy,but also has a high heart rate and mortality in long-term observation.Studies have shown that cardiomyocytes can synthesize TNF-? and have TNF-? Receptor on the cell membrane.Numerous studies have found that cardiomyocytes act as target cells for TNF-? action,and high concentrations of TNF-? can cause ventricular remodeling due to cardiomyocyte hypertrophy,and TNF-? levels are closely related to the degree of heart failure.Studies have shown that IL-6 and TNF-? also affect myocardial systolic and diastolic function,leading to decreased cardiac function and involvement in the development of ventricular remodeling.There is no research on the relationship between AQP1 and inflammatory factors.P38 MAPK is an important inflammation in the body.Signal-mediated pathway,involved in a variety of inflammatory responses and promotes apoptosis,which is involved in the ischemia,reperfusion injury of the heart,brain and kidney,in addition to mediating the inflammatory response,Studies have shown that hypoxia can up-regulate AQP1 in prostate cancer cells in a P38MAPK-dependent manner.Other studies have found that in hypoxic human pleural mesothelial cells,peptidoglycan,which inhibits p38 MAPK,can be used.The expression of AQP1 was down-regulated.It is speculated that P38 MAPK may be an important way to up-regulate the expression of AQP1.However,there is no report on the relationship between AQP1 expression and P38 MAPK in cardiomyocytes.Although there are many studies on AQP1,there are few reports on its expression in the myocardium and its effect on ventricular remodeling.In this study,we observed the expression of AQP1 in myocardium under ischemia and pressure overload,and selected AZA as an intervention to study its effect on AQP1 expression in myocardial tissue.Observe this treatment for heart water content,heart weight,and cells apoptosis and cardiac ultrasound indicators,in order to delay the inhibition of ventricular remodeling to find new treatments.In addition,the levels of IL-6,TNF-? and P-P38 MAPK in myocardial tissue were observed at the same time,in order to further study the pathological process of early ventricular remodeling,and to explore the interaction between these three in ventricular remodeling.The study was divided into three parts: the first part is AQP1 expression and role in myocardial ischemia-reperfusion injury in mice.The second part is AZA inhibits early ventricular remodeling induced by myocardial ischemiareperfusion injury in mice by reducing AQP1 transcription and expression.The third part is AZA inhibited early ventricular remodeling induced by pressure overload in mice by reducing AQP1 transcription and expression.Part one AQP1 expression and its role in myocardial ischemia reperfusion injury in miceObjective: To investigate the relationship between AQP1 overexpression and the occurrence and persistence of myocardial edema after ischemiareperfusion,the relationship between inflammatory factors IL-6,TNF? and p38-MAPK activation,the relationship between p-p38 MAPK and AQP1 expression.Furthermore,it provides research directions on how to prevent myocardial edema after ischemia-reperfusion and protect damaged myocardium.Methods: To detect the expression levels and regularity of IL-6,TNF?,p-p38 MAPK and AQP1 in mice after MI/RI,80 mice were randomly divided into the following 16 groups.Group A: sham operation group,according to MI / RI modeling method for thoracotomy,only threading before ligation.Group B was ligated according to the MI/RI modeling method and the anterior descending coronary artery was ligated for 30 minutes.Groups A and B were divided into 8 groups according to the time of material collection.At 4 hours,12 hours,24 hours,48 hours,72 hours,5 days,7 days,and 14 days,the experimental animals were taken at the corresponding time points.The mice were sacrificed by cervical dislocation,the thoracic cavity was quickly opened,and the heart was removed and washed by 4 ° C Pre-cooled saline,made into tissue homogenate to detect IL-6 and TNF? levels in myocardial tissue.Western Blot was used to detect P-P38 MAPK and AQP1 relative protein expression,and dry and wet weight method was used to measure myocardial water content.Results: Ischemia-related changes were observed after the mice were treated with the surgery.The shape of the heart was observed at each time point.After 48 hours,the heart of the MI/RI group was significantly larger than that of the non-surgical group,and the possibility of ventricular remodeling due to myocardial ischemia-reperfusion was considered to be large.The levels of IL-6 and TNF-? in the myocardium of MI/RI group were significantly higher than those in the control group at all time points,which increased from 4 hours after operation and peaked at 24 hours.After that,the concentration gradually decreased,but It was still at a high level.The expression of P-P38 MAPK protein in the MI/RI group was significantly higher than that in the sham operation group at all time points.It increased from 4 hours after surgery and peaked at 48 hours.After that,the protein expression decreased gradually.The relative expression of P-P38 MAPK protein was correlated with TNF-?.The expression of AQP1 protein in MI/RI mice was significantly higher than that in the sham operation group,and it showed a slight downward trend from 4 hours to 24 hours after operation.The expression level increased sharply from 24 hours to 1 week after operation.It reached a peak at 1 week after surgery.There was a correlation between AQP1 and P-P38 MAPK expression.The left ventricular myocardium water content in the MI/RI group was higher than that in the control group at each time point.It increased from 4hours to 12 hours after after surgery,and then decreased to 48 hours after the peak,and then the myocardial content increased again 48 hours after the operation,reached the peak at 5 days,and remained at a high level until 1 week,and then showed a downward trend.Left ventricular myocardial water content was associated with il-6 and TNF-? at various time points in 48 hours.The left ventricular myocardial water content was closely related to AQP1 after 48 hours.Conclusion: Ischemia-reperfusion injury can lead to myocardial edema,ventricular remodeling,and changes in the shape of the heart,resulting in a significant increase in ventricular volume.In the early stage of ischemiareperfusion injury,IL-6 and TNF-? were significantly increased in myocardial tissue,and were closely related to early myocardial edema.IL-6 and TNF-? were one of the important factors mediating ventricular remodeling.The level of TNF-? is correlated with the expression of P-P38 MAPK.Phosphorylation of P38 MAPK is closely related to the expression of AQP1 protein in mouse myocardium.The expression of AQP1 was closely related to the water content of myocardial tissue in mice,especially myocardial edema after 48 hours.Part two AZA inhibited early ventricular remodeling induced by myo-cardial ischemia-reperfusion injury in mice by reducing AQP1 expressionObjective: To investigate whether AZA administration can improve early ventricular remodeling caused by myocardial ischemia-reperfusion and explore the ways in which AZA administration affects early ventricular remodeling.Methods: Forty C57 BL / 6 mice were randomly divided into four groups,10 in each group: sham operation group(sham group),sham operation + acetazolamide group(sham + AZA group),ischemia reperfusion injury group(MI/RI group): ischemia-reperfusion injury + acetazolamide group(MI/RI+ AZA group).One week after the operation,the mice were anesthetized with ether and echocardiographically detected the cardiac structure and cardiac function.Then,the mice were sacrificed by cervical dislocation,the heart was taken out,and the heart weight/body weight ratio(HW / BW)was calculated.The myocardial tissue was sectioned and stained,and the morphology and apoptosis of the cells were observed.The myocardial tissue was homogenized and the expression of inflammatory factors was detected.The protein expression of P-P38 MAPK was detected by Western blot.and AQP1 mRNA and protein expression were detected by RT-PCR and Western blot,dry and wet weight method was used to calculate myocardial water content.Results: Compared with the control group,the heart volume of the MI/RI group was significantly increased and the lumen was enlarged.HE staining,myocardial cell in MI/RI group was edema,structural destruction,myocardial fiber disorder,gap widening,and a large number of inflammatory cell infiltration were observed.AZA administration can reduce heart enlargement,reduce myocardial cell edema,structural damage,and inflammatory cell infiltration.TUNEI staining showed that the apoptotic rate of myocardial cells in MI/RI group was significantly higher than that in the control group.AZA administration could significantly reduce the apoptosis induced by MI/RI.The levels of IL-6 and TNF-? in myocardial tissue of MI/RI group were significantly higher than those of control group.The administration of AZA could decrease the levels of IL-6 and TNF-? in myocardial tissue of MI/RI mice.The relative expression of AQP1 protein and the relative expression of mRNA in the MI / RI group were significantly higher than those in the control group.AZA administration can reduce the expression of AQP-1 protein and mRNA in myocardial tissue of MI/RI mice.The relative expression of P-P38 MAPK protein in MI/RI group was significantly higher than that in the control group.AZA administration can reduce the relative expression of P-P38 MAPK protein in myocardial tissue of MI/RI mice.The myocardial water content and heart weight/body weight(HW/BW)ratio in the MI/RI group were significantly higher than those in the control group.AZA administration can reduce the water content and HW/BW of myocardial tissue in MI/RI mice.IVSD,LVAWD,and IVSS were significantly thicker in the MI/RI group than in the control group.LVAWS was thinner than the control group,LVEDD and LVESD were significantly greater than the control group,LVEF were lower than the control group.AZA administration improved MI/RI-induced LVEF decreased,and MI/RI caused LVEDD,LVESD increased,and LVAWD thickened,but had no significant effect on IVSD,IVSS,and LVAWS changes.Conclusion: Ischemia-reperfusion injury can lead to myocardial cell edema,structural destruction,apoptosis,increased heart volume,ventricular remodeling,result in increased transcription and expression of AQP1,which in turn causes myocardial cell edema,increased apoptosis,myocardial tissue water content and HW/BW ratio increase,and the left ventricle volume enlarged,the heart function decreased,and then ventricular remodeling occuring.AZA administration can significantly reduce the transcription and expression of AQP1,reduce myocardial cell edema,apoptosis,myocardial water content,cardiac volume,and improve cardiac structure and function,inhibit ventricular remodeling,but have no significant effect on cardiac hypertrophy.AZA administration can reduce the expression of P-P38 MAPK protein induced by MI/RI,suggesting that AZA may play a role in reducing AQP1 transcription and expression by inhibiting P38 MAPK signaling pathway.AZA administration also reduced the levels of IL-6 and TNF-? in myocardial tissue induced by MI/RI,which may reduce the apoptosis and delay the ventricular remodeling.Part three The expression and function of AQP-1 in the pressure overload modelsObjective to observe the expression levels of il-6,TNF-,p-p38 MAPK and AQP1 in mice with pressure overload at different time points,as well as the myocardial water content at different time points,so as to explore the correlation between the overexpression of AQP1 and myocardial edema under the condition of myocardial pressure overload,as well as the correlation between the levels of inflammatory cytokines il-6,TNF-,and P38 MAPK phosphorylation.Methods 80 mices were randomly divided into two groups as follows: group A: sham group : thoracotomy was performed according to the method of model making,but the aortic arch was not ligated by thread.In group B,aortic arch constriction group(TAC group)was modeled by partially ligated according to aortic arch constriction.In the two groups,respectively,in 4 hours,12 hours,24 hours,48 hours,72 hours,five days,7 days and 14 days time line based on experimental animals,with cervical dislocation executed mice,quickly open the chest,take out the heart,with 4 ? cold saline flushing,tissue homogenate detection of myocardial tissue IL-6,TNF alpha level,Western Blot test P-P38 lightning MAPK,AQP1 protein expression,The content of water in myocardial tissue was measured by dry and wet gravimetric method.Results myocardial pressure overload led to myocardial injury and ventricular remodeling.The levels of il-6 and TNF-in myocardial tissue of the TAC group were significantly higher than those of the control group at all time points.The expression of AQP1 in sham group was constant at each time point,and there was no statistically significant difference among the time points.The expression level of AQP1 protein in mice in the TAC group increased significantly from 4 hours after surgery,reached a peak at 72 hours after surgery,then decreased,and remained at a high level until 14 days after surgery.Compared with sham group,the expression levels of AQP1 protein at each time point in TAC group were significantly increased.The expression level of p-p38 MAPK protein in mice in the TAC group increased significantly from 4 hours after the surgery,reached a peak 48 hours after the surgery,then decreased,and remained at a high level until 14 days after the surgery.Compared with sham group,the expression levels of p-p38 MAPK protein in TAC group were significantly increased at each time point.The content of myocardial water in the left ventricle of the sham group was constant at each time point,and there was no statistically significant difference in the content of water between the time points.In the TAC group,the myocardial water content in the left ventricle increased from 4 hours after surgery to 5 days after surgery,and then decreased slightly to 14 days after surgery.Conclusion pressure overload can lead to myocardial edema,ventricular remodeling,change of cardiac geometry,and increase cardiac volume.Pressure overload significantly increased the expression level of AQP1,which was closely related to the water content of the mouse cardiac tissue.Stress overload significantly increased il-6 and TNF-in myocardial tissue and was associated with myocardial edema.The phosphorylation level of P38 MAPK was correlated with the expression of AQP1 in mouse myocardium.Part four AZA inhibits early ventricular remodeling induced by pressure overload in mice by reducing AQP1 expressionObjectives: To investigate the effects of pressure overload on myocardial hypertrophy,edema,apoptosis and ventricular remodeling in mice,and to analyze the role of AQP1 in this progress,whether AZA administration can improve the above indicators.Methods: 40 C57 BL / 6 mice were randomly divided into four groups,10 in each group: sham operation group,sham operation + acetazolamide group,aortic arch constriction(TAC)group,aortic arch constriction + acetazolamide(TAC+AZA)group.At 2 weeks after surgery,mice were anesthetized with ether and echocardiography was used to detect cardiac structure and cardiac function.The mice were sacrificed by cervical dislocation,the heart was taken out,and the heart weight/body weight ratio(HW / BW)was calculated.Some ischemic tissue was taken for sectioning and staining to observe the morphology and apoptosis of the cells.Another part of myocardial tissue was taken for homogenization to detect the inflammatory factors.the expression of P-P38 KMAP protein was detected by Western blot.The mRNA and protein expression of AQP1 were detected by RT-PCR and Western blot,respectively.The water content of myocardium was calculated by dry and wet weight method.Results: Compared with the control group,the TAC model group had increased heart volume,geometric shape,and left ventricular cavity enlargement;HE staining showed myocytes edema,hypertrophy,irregular shape,and increased intercellular space;AZA administration can reduce myocardial cells Edema,hypertrophy,structure damage,inhibit heart enlargement;TUNEI staining showed that the apoptosis rate of cardiomyocytes in TAC group was significantly higher than that of control group;AZA administration could significantly reduce TAC-induced apoptosis;IL-6 and TNF-? in myocardial tissue of TAC group was significantly increased.AZA administration could decrease the expression of IL-6 and TNF-? induced by TAC.The relative expression of AQP1 mRNA and protein in TAC group was significantly increased,AZA administration could reduce the AQP1 expression.The relative expression of P-P38 MAPK protein in TAC group was significantly increased,AZA administration could reduce the expression of P-P38 MAPK.TAC group heart weight / body weight(HW / BW)and water content of myocardial tissue was significantly increased.AZA administration could reduce the water content and HW/BW radio.Compared with the control group,IVSD,IVSS and LVPWD were significantly thickened in the TAC group,while LVEDD,LVEF and LVFS were significantly decreased.AZA administration could significantly improve LVEF and LVFS in TAC group.However,in AZA administration group there was no significant impact on IVSD,IVSS,LVPWD and LVEDD changes induced by TAC.Conclusion: Pressure overload can lead to myocardial cell edema,hypertrophy,structural damage,increased heart volume,ventricular cavity geometry changes,leading to ventricular remodeling;result in increased transcription and expression of AQP1,which in turn causes myocardial cell edema and increased apoptosis,increased myocardial tissue water content and HW/BW ratio,enlarged left ventricular volume and decreased cardiac function.AZA administration can significantly reduce AQP1 transcription and expression,myocardial cell edema,apoptosis,myocardial water content,and heart Volumetric quality,then improve heart function.AZA administration can decrease the expression of P-P38 MAPK protein,suggesting that AZA may play the role in reducing AQP1 transcription and expression by inhibiting P38 MAPK signaling pathway.In addition,inhibition of the P-P38 MAPK signaling pathway may reduce IL-6 and TNF-? levels.AZA administration also reduced the levels of IL-6 and TNF-? in myocardial tissue caused by pressure overload,which may reduce apoptosis and further improve cardiac function.In addition,it is speculated that myocardial hypertrophy caused by pressure overload may be more correlated with elevated levels of IL-6 and TNF-?. |
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