A New Method For Assessing Pathological Remodeling Of Pulmonary Hypertension And A Database Of BMPR2 Mutations Were Established

Objective Right heart catheterization is the gold-standard for measurement of right ventricular hemodynamics.However,it is difficult to perform in mice.We thus aimed to establish a new open chest method to record and analyze the right ventricular hemodynamic parameters in intubated mice connected to a respirator under anesthesia.Methods Eight-week old male C57BL/6 mice were divided into the control group,and the hypoxia group using completely random method.Mice in the hypoxia group were exposed in low pressure oxygen chamber,which simulated the environment of plateau(5500 m altitude,10%oxygen)for 3 weeks.Mice in the control group were exposed in normal pressure and oxygen environment.The mice were intratracheally intubated and connected to a respirator,chest was opened,right ventricular hemodynamics were measured with a needle(0.7 mm×19 mm)punctured into right ventricle(5 mm in death),which was connected to the measuring equipmentResults The open chest procedure could be finished within 5-10 minutes.The successful rate was 100%.There was no significant difference in heart rate between control group and hypoxia group under anesthetia((306.4±11.5)bpm vs.(320.4 ± 16.0)bpm,p>0.05).Compared with the control group,both the right ventricular systolic pressure(RVSP)and the mean right ventricular pressure(mRVP)were increased in hypoxia group((17.1±1.0)mmHg vs.(22.6±1.0)nmHg,p<0.01;(9.6±0.8)mmHg vs.(12.4± 0.3)mmHg,p<0.01).Moreover,the absolute value of the maximal rate of increase in right ventricular pressure(dp/dtmax)and the maximal rate of decrease in right ventricular pressure(dp/dtmin)were higher in the hypoxia group(421.3± 30.6)mmHg/s vs.(639.7 ± 47.7)mmHg/s,p<0.01;(-324.3±24.0)mmHg/s vs.(-496.5±40.6)mmHg/s,p<0.01),indicating the pathological status after hypoxia treatment.Conclusions Right ventricular hemodynamics measurement with this open chest technique is easy to handle and can accurately reflect the right ventricular pressure in mice under physiological and pathological conditions.Background:Right ventricular failure(RVF)is a common cause of death among patients with pulmonary arterial hypertension(PAH).Early detection of RVF is critical for clinical management of PAH.The two-dimensional speckle tracking echocardiography(2DSTE)is a novel non-invasive method to assess RVF.Here,we investigate whether 2DSTE could detect early right ventricular remodeling in PAH rat model.Method:PAH and subsequent RVF were induced in male Sprague-Dawley rats by a single i.p.injection of monocrotaline(MCT,55mg/kg).Hemodynamic changes were measured by right heart catheter(RHC).Right ventricular(RV)structure and function were assessed by 2DSTE and conventional echocardiography(CE)simultaneously on the 0th,7th,14th,28th,and 42th day after MCT injection.Results:Mean PAP was significantly increased on the 14 day post MCT injection,assessed by RHC(mPAP,day 0 v.s.day 14 21.63±1.08 mmHg v.s.31.55±1.80mmHg,p<0.001).No changes were detected in the RV and vessel using conventional echocardiography,including the right ventricular anterior wall thickness at diastolic(RVAW,d),interventricular septal thickness at diastole(IVSd),pulmonary vessel diam(PV diam).In addition,no observable changes were detected in the RV and vessel function through conventional echocardiography,such as ejection fraction(EF),tricuspid annular plane systolic excursion(TAPSE),pulmonary artery acceleration time-to-ejection time ratio(PAAT/PAET),pulmonary vessel peak vel(PV peak vel).Interestingly,average radial strain(RS)and strain rate(RSR)of the anterior,middle and posterior part of right ventricular wall,showed marked decrease as early as day 14(RS,dayO v.s.day14 35.54 ± 4.21%v.s.22.14±3.61%,p<0.05)(RSR,dayO v.s.dayl46.58±0.67 1/S v.s.4.19±0.51 1/S,p<0.01)after MCT injection.Moreover,Kaplan-Meier curve according to the 14 day shows that higher ARS(p = 0.08)and ARSR(p = 0.04)with better survival rate.Conclusions:Our study indicate that 2DSTE serves as a more sensitive approach for early detection of right ventricular dysfunction than CE and RHC,it can predict the outcome of PAH rat.Aim:Hereditary Pulmonary Artery hypertension(HPAH)and Idiopathic Pulmonary Artery hypertension(IPAH)are rare disease with high mortality,the mechanism of it still unclear.BMPR2 mutation is the main virulence gene,but there still have no ideal animal model.By review the literature,we identified all BMPR2 mutations and pick out high frequency mutants,we want to develop a new BMPR2 mutation animal modle.Method:Firstly,searching informations on the Pubmed,filting out useful informations.Secondly,using Excel software building a data base,which including all the BMPR2 mutations.Thirdly,finding out mutations which frequency greater than or equal to 3times in patients.And using SIFT,Polyphen2,MutationTaster to blast the conservatism of gene sequence and animo acid sequence.to making sure the mutations are pathogenic,then developing an rat animal model.Results:We finally find out 434 BMPR2 gene independent mutations and we located the absolute location of them by Vector NTI and UCSC website.We find out most of the mutation only have been found out one time(78%),the mutation widely located all the regions of the gene,but most of them in the kinase domain.Secondly,filting the data base again by the times(≥3)they have been found in the patients,we get 26 different mutations.Most of them are nonsense mutation,next is the missense mutation.making sure the pathogenic information from blast the sequence from SIFT,Polyphen2,MutationTaster finally we get 20 gene mutations.There totally have 4 mutations which frequency greater than 10 times.Mutation c.2695C>T(p.R899X)has been reported 11 times and have been successfully established animal modle.Mutation c.C1471T(p.R491W)had been reported 11 times.The SIFT score is 0(<0.05 means deletorious),Polyphen 2 score and Mutation Taster score are 1(>0.95 means deletorious).It’s widely distributed in patients compare to other mutarion according to the disease type.And our group have been reported this mutation in 2004 in HPAH.Conclusions:Mutation c.C1471T(p.R491W)is the best choice to establish a new Primary Pulmonary Artery Hypertension animal modle.