| Part 1:Construction of minipig model of right heart failure induced by pressure and volume overload and screening of specific markers of right heart failure based on untargeted metabolomicsObjective:Right heart failure refers to the clinical syndrome that occurs when the systolic and/or diastolic dysfunction of the right ventricle caused by any cause is insufficient to provide the required cardiac output of the body.Pressure and volume overload are the most common causes of right heart failure.However,the mechanism of right ventricular failure and the metabolic changes in right ventricular myocardial tissue are still unclear,and there are no effective and specific biomarkers for right heart failure.In this study,Bama minipig models of right heart failure caused by pressure and volume overload was constructed,and metabolomics technology was used to analyze the changes of metabolites in the myocardium and plasma of minipigs caused by right heart failure,so as to find the biomarkers and underlying mechanism of right heart failure.Methods:In this research,13 6-month-old Bama miniature pigs were divided into three groups randomly,including pulmonary artery binding(PAB)group(n=5),pulmonary valve regurgitation(PVR)group(n=5),and SHAM group(n=3).In the PAB group,the operation resulted in right ventricular pressure overload,while in the PVR group,the operation resulted in right ventricular volume overload due to the removal of two pulmonary leaflets.Through echocardiography,the structural and functional parameters of the right heart were recorded before surgery and in 1,2,and 3 months after surgery,and cardiac MRI was conducted before tissue collection.Peripheral blood and right ventricular myocardium in anterior wall were collected.Masson staining was used to analyze the fibrosis of the right ventricular myocardium,and transmission electron microscopy was used to observe the myocardial structural changes.Differential metabolites and metabolic pathways were screened by and untargeted metabolomics sequencing based on LC-MS technology.Results:3 months after the modeling operation,transthoracic ultrasound measurements showed that the thickness of the anterior wall and free wall of the right ventricle in the PAB group increased significantly compared with the baseline.Thickness of right ventricular anterior wall increased by 62.0%in the basal segment,61.5%in the middle segment,and 54.8%in the apical segment.Free wall thickness increased by 49.7%in the basal segment,49.8%in the middle segment,and 47.0%in the apical segment.Similar to the PAB group,the thickness of the anterior wall and free wall of the right ventricle of the PVR group also showed a trend of increasing over time.The basal segment of anterior wall increased by 41.5%,the middle segment of anterior wall increased by 37.1%,and the apical segment of anterior wall increased by 49.2%.The basal segment of free wall increased by 39.7%,the middle segment of free wall increased by 48.3%,and the apical segment of free wall increased by 52.6%.Size of the right ventricular chamber in the two operation groups also showed an increasing trend.The right ventricular function of the two operation groups was significantly lower than that at the baseline.The RVEF of the PAB group decreased from(60.0±4.86)%at the baseline to(45.2±5.70)%(p=0.016),and the RVEF of the PVR group decreased from(58.8±8.03)%to(42.4±9.81)%(p=0.007).The LVEF in the PAB group remained stable,while the LVEF in the PVR group decreased from(64.2±4.50)%to(49.0±5.30)%(p=0.022).Compared the two operation groups,PAB group had more obvious wall thickness increase,and PVR group had more obvious heart chamber enlargement.Cardiac MRI revealed that in the two operation groups,the anterior wall thickness of the right ventricle was about twice as thick as that in the SHAM group and the volume of the right ventricle increased in two operation groups.The end-diastolic volume in the PVR group was as about 1.9 times as that of the SHAM group[(100.83±14.44)ml vs.(53.08±2.09)ml,p=0.006],and the end-systolic volume was about as 3.0 times as that of the SHAM group[(62.04±17.30)ml vs.(20.67±1.43)ml,p=0.003].Masson staining found that the area of fibrosis in the SHAM group was(1.52±0.18)%,the area of fibrosis in the right ventricle of the two operation groups was(5.19±1.20)%in the PAB group and(5.04±1.52)%in the PVR group,respectively.The fibrosis area increased significantly in the operation groups(p=0.011 in PAB group andp=0.014 in PVR group).The transmission electron microscope found that the myocardium z-line became shallower and the mitochondria were swollen and broken in the two operation groups.According to OPLS-DA analysis,the myocardial and plasma metabolic profiles of animals in SHAM group,PAB group and PVR group were significantly different.In the myocardium,a total of 44 metabolites were identified by PAB vs.SHAM,and 21 metabolites were identified by PVR vs.SHAM.Among them,11 were common differential metabolites under pressure and volume overload.In plasma,a total of 34 metabolites were screened for PAB vs.SHAM,and 24 metabolites were screened for PVR vs.SHAM.Among them,8 were in common.Compared with SHAM group,the different metabolic pathways in PAB group were Histidine metabolism,Aginine and proline metabolism,DGlutamine and D-glutamate metabolism,etc.Compared with SHAM group,the different metabolic pathways in PVR group were Histidine metabolism,β-Alanine metabolism,Amino sugar and nucleotide sugar metabolism.In plasma,the different metabolic pathways compared SHAM group and PAB group including Vitamin B6 metabolism,Phenylalanine metabolism and Arginine biosynthesis.Compared with SHAM group,Arginine biosynthesis,Pentose and glucuronate interconversions,Pentose and glucuronate interconversions and pentose phosphate pathways were different in PVR group.Conclusion:(1)Both pressure and volume overload can cause typical right heart failure phenotypes such as right ventricular wall thickening,right ventricular chamber enlargement,and right heart function reduction in minipigs;(2)Compared with volume overload,pressure overload causes more obvious thickening of the right ventricular wall,and volume overload has a greater impact on the size of the right ventricular chamber and left ventricular function;(3)Untargeted metabolomics sequencing of myocardial tissue found that compared with the SHAM group,amino acid such as histidine metabolic pathways in the PAB group and PVR group were more vulnerable,suggesting that amino acid metabolism may be involved in the occurrence and development of right heart failure;(4)In the plasma of minipigs in PAB group and PVR group,arginine biosynthesis was a common differential metabolic pathway and arginine was a common differential metabolite compared with SHAM group,suggesting the potential of arginine as a biomarker of right heart failure.Part 2:Single-cell atlas of the human normal and regurgitant tricuspid valveObjective:The occurrence and severity of tricuspid regurgitation often indicate a poor prognosis for patients.However,the mechanism of TR is currently poorly understood,and the cellular composition,molecular characteristics,and cellular heterogeneity in the regurgitant TR remain largely unknown.In this study,by separating the tricuspid valve tissue of patients with tricuspid valve replacement and heart transplantation,the single-cell map of normal and regurgitant tricuspid valve was constructed by single-cell sequencing technology,hoping to provide some clues for the occurrence and development mechanism of tricuspid regurgitation.Methods:In this study,10 tricuspid valves from patients with tricuspid valve replacement surgery and heart transplantation were divided into non diseased control(NC)group and regurgitation(TR)group according to the ultrasonic examination.After separating single cells from the valve,cell types were identified based on classic marker genes and cell subtypes were further identified.Joint comparative analysis was conducted to systematically describe the patterns of abnormal gene expression in the samples.Perform computer analysis was used to record dysregulated intercellular interactions and validation was done through Opal multi-color IHC staining and Western Blot.Results:The study evaluated the transcriptional profiles of 84102 cells and identified 6 main cell types and 25 cell clusters in NC and TR group specimens.Valve interstitial cell(VIC)is the most abundant cell type in leaflet,followed by myeloid,lymphoid,valve endothelial cells(VEC),myofibroblast-like valve interstitial cell(myoVIC)and mast cell.Further analysis showed that in TR group,the number of cells characterized by high expression of COMP in VIC was more,and these cells may associated with valve fibrosis and extracellular matrix remodeling.The enhanced interaction of COMP-CD47 in TR may explain the mechanism of extracellular matrix remodeling of the regurgitant tricuspid valve.Conclusion:(1)For the first time,single-cell transcriptome sequencing technology was used to analyze the gene expression and regulatory characteristics of cell types and cell clusters in human tricuspid valve tissue;(2)The VIC cluster characterized by high expression of COMP and the interaction between COMP and CD47 may play a certain role in the remodeling of valve extracellular matrix in the TR group;(3)This study advanced our understanding of cellular heterogeneity in tricuspid valve tissue and may shed light on mechanisms elucidating of tricuspid regurgitation in the future.Part 3:Concomitant repair of moderate or less tricuspid regurgitation during mitral-valve surgery improves early and midterm prognosis of tricuspid valve and right heart functionObjective:Valve disease of the left heart system,especially the mitral valve,is the most common cause of functional tricuspid regurgitation.It was previously believed that tricuspid regurgitation can be alleviated after left heart valve surgery alone,and the structure and function of the right heart can also be restored.However,the surgical timing and surgical methods for functional tricuspid regurgitation are still controversial.In this study,the clinical data of patients undergoing mitral valve surgery were collected,and the effect of concomitant surgical repair of less than moderate tricuspid regurgitation on the prognosis of tricuspid valve and right heart was analyzed,in order to shed a light on the diagnosis and treatment of the dysfunction of the right heart system.Methods:In this study,a retrospective study of 461 patients with mitral valve disease requiring cardiac surgery combined with less-than moderate tricuspid regurgitation in Fuwai hospital from 2011 to 2014 was done.According to whether they received tricuspid valve repair(including Kay’s annuloplasty,Devega’s annuloplasty or annular ring implantation),the patients(of which 152 were female,the median age was 53 years)were divided into mitral valve surgery only group(nTAP group)and concomitant tricuspid valve repair group(TAP group).The degree of tricuspid regurgitation and right ventricular diameter were recorded before and after operation.At the same time,43 patients whose tricuspid valve annulus diameter was less than 40mm in the TAP group were analyzed in subgroups.Results:The median follow-up duration was 3.0 years(range from 0.1 to 9.3 years).There was no perioperative death,and tricuspid valve surgery did not increase perioperative complication rates.Three months after surgery,the anteroposterior diameter of the right ventricle in the TAP group was significantly improved compared with that in the nTAP group(-1(-3,1)mm vs.0(-0.2,2)mm,p=0.048).Three years after surgery,the improvement of right ventricular anteroposterior diameter in the TAP group was still significant compared with the nTAP group(-1(-2.75,2)mm vs.2(-0.75,4)mm,p=0.014),and the patients in the TAP group were less likely to develop moderate or more tricuspid regurgitation(3.64%vs.35.64%,p<0.001).Annuloplasty ring implantation was more effective in preventing regurgitation progression(p=0.044).For patients with a tricuspid annulus diameter of less than 40 mm,concomitant tricuspid valve repair was still effective in improving the anteroposterior diameter of the right ventricle in the early follow-up(p=0.036).Conclusion:(1)Concomitant tricuspid valve repair for patients with less-than-moderate tricuspid regurgitation during mitral valve surgery can effectively improve the tricuspid valve and right heart function in the early and mid-term after surgery;(2)Annuloplasty ring implantation was more effective in preventing regurgitation progression than suture plasty;(3)Patients with preoperative annulus less than 40 mm can also obtain improvement in right ventricular structure from less than moderate tricuspid regurgitation intervention during mitral valve surgery. |
PDF Full Text Request