| Objectives:We intended to explore the natural process of MCT induced PAH in rats using a dosage of 60mg/kg,and investigate whether the model is consistent with the natural process of human PAH.Methods:24 SPF male SD rats were randomly divided into 3 groups with 8 rats in each group:group MCT induced PAH for 3 weeks(group MCT-3wks-PAH),group MCT induced PAH for 4 weeks(group MCT-4wks-PAH)and group control(group Ctrl).Rats in group-PAH were given a single intraperitoneal injection of MCT(60mg/kg)to induce the development of PAH,and group Ctrl received intraperitoneal injection of normal saline with a same volume.On day 21 and 28,the following data of all rats were tested:1)Survival rate.2)weight and weight increment(WI).3)Hemodynamics:Right ventricle pressure(RVP)and pulmonary arterial pressure(PAP),including systolic blood pressure,diastolic blood pressure and mean blood pressure.4)Right ventricular hypertrophy index(RVHI).5)Lung tissue for HE stainingResults:1)The survival rate of groups MCT-3wks,MCT-4wks and Ctrl was 75%(6/8),50%(4/8)and 100%(8/8),respectively.Survival rate of group Ctrl was significantly higher than that of group PAH(P<0.05).2)The weight in group Ctrl continued to grow,while in group PAH showed slower grow up curve and began to decrease since day 21.On day 21,the WI of groups MCT-3wks and MCT 4wks was significantly lower than that of group Ctrl(102.4±13.12,109.8 ±7.57 vs.172.1±7.70g,P all<0.0001);On day 28,the WI of group MCT-4wks was also significantly lower than that of group Ctrl(54.38±32.04 vs.216.4±10.54g,P all<0.0001),and also significantly lower than its own WI on day 21(54.38±32.04 vs.109.8±7.57g,P all<0.0001).3)Compared with group Ctrl,the PAP and RVP in groups MCT-3wks and MCT-4wks was significantly increased:SPAP(49.15±5.31,39.65±4.59 vs.20.23±2.20 mmHg,P all<0.0001),DPAP(42.09±7.11,27.37±6.67 vs.15.60±2.58 mmHg,P all<0.0001)and MPAP(45.97±6.18,33.29±4.35 vs.18.36±2.15 mmHg,P all<0.0001);SRVP(60.51±7.20,47.37±4.84 vs.29.37±5.32 mmHg,P all<0.0001),DRVP(7.17±2.44,13.22±4.94 vs.3.74±1.90 mmHg,P<0.05 and P<0.0001)and MRVP(32.67±4.59,31.00±4.37 vs.15.84±2.55 mmHg,P all<0.0001).The PAP of group MCT-4wks was significantly lower than that of group MCT-3wks:SPAP(39.65±4.59 vs.49.15±5.31 mmHg,P<0.01),DPAP(27.37±6.67 vs.42.09±7.11 mmHg,P<0.01)and MPAP(33.29±4.35 vs.45.97±6.18 mmHg,P<0.01).As for RVP,SRVP of group MCT-4wks was significantly lower than that of group MCT-3wks(47.37±4.84 vs.60.51 ±7.20 mmHg,P<0.01),on the contrary,DRVP was significantly higher than group MCT-3wks(13.22±4.94 vs.7.17±2.44 mmHg,P<0.05),however,MRVP showed no significant difference(31.00±4.37 vs.32.67±4.59 mmHg,ns).4)The RVHI of group MCT-3wks and MCT-4wks was higher than that of group Ctrl(0.612±0.075,0.73 1±0.124 vs.0.285±0.036,P all<0.0001),and RVHI of group MCT-4wks was significantly higher than that of group MCT-3wks(0.731±0.124 vs.0.612±0.075,P<0.05).5)compared with group Ctrl,group MCT-PAH had obvious pulmonary vascular remodeling.And for the pulmonary arteries with a diameter<100?m,pulmonary vascular remodeling is more obvious in group MCT-4wks,which even existed pulmonary vascular occlusion,than group MCT-3wksConclusions:The natural process of MCT(60mg/kg)intraperitoneal injection induced PAH in rats is consistent to the natural process of human PAH.The model is a suitable and stable animal model that can be used for the research on the pulmonary vascular remodeling of PAH.Objectives:We tried to screen the differentially expressed miRNA in the pulmonary arteries between MCT induced PAH rats(MCT-3wks)and Control rats(Ctrl)using miRNA microarray technique,and find out the key miRNA involved in the regulation of pulmonary vascular remodeling in PAH,which could provide a new perspective for the in-depth mechanism study of pulmonary vascular remodeling in PAH and create a new target for the prevention and treatment of PAH.Methods:miRNA expression profiles of the pulmonary arteries samples from 3 MCT-3wks rats and 3 Ctrl rats were analyzed by miRNA microarray,and the differentially expressed miRNA were screened out.The expression level of miR-125a-5p was verified by RT-qPCR in 6 MCT-wks rats and 6 Ctrl rats.Then we used bioinformatics analysis methods to predict the target gene of miR-125a-5p,and their Gene Ontology(GO)enrichment analysis was also performedResults:1)miRNA expression profiles showed there were 79 differentially expressed miRNAs in pulmonary arteries between MCT-3wks rats and Ctrl rats with 27 up-regulated and 52 down-regulated.2)RT-qPCR results showed the expression of miR-125a-5p in the pulmonary arteries of MCT-3wks rats was down-regulated when compared with group Ctrl(P<0.01).3)35 target genes of miR-125a-5p were predicted from the TargetScan,miRnada and miRDB databases.And their GO analysis showed miR-125a-5p is mainly involved in the regulation of mitochondrial function,in which the target gene signal transducer and activator of transcription 3(STAT3)plays a leading roleConclusions:miR-125a-5p may play an important role in the regulation of mitochondrial function in the development of PAH,the specific mechanism is worth further studiesObjectives:To investigate the effect of miR-125a-5p on the function of PASMCs and its specific mechanismMethods:The expression of miR-125a-5p,as well as its tissue specialty,in PASMCs and MCT-PASMCs was detected by RT-qPCR.miR-125a-5p agomir and antagomir were transfected into PASMCs to mimic its overexpression or loss of function,CCK8 and AnnexinV/PI were used for the analysis of proliferation and apoptosis,and the expression of TGF-?1,IL-6,STAT3 and Bcl-2,as well as the expression of miR-125a-5p after stimulation by TGF-?1and IL-6were detected by RT-qPCRResults:The expression of miR-125a-5p in vascular tissues(pulmonary artery and aorta)and enriched vascular tissues(brain and kidney)was significantly higher than that in other tissues.The expression of miR-125a-5p in MCT-PASMCs was significantly lower than that in PASMCs(P<0.001).The transfection of miR-125a-5p agomir can inhibit the proliferation and promote the apoptosis of PASMCs,while miR-125a-5p antagomir can promote the proliferation and inhibit the apoptosis of PASMCs.miR-125a-5p agomir can inhibit the expression of TGF-?1and IL-6,while miR-125a-5p antagomir can stimulate the expression of TGF-?1 and IL-6.Moreover,TGF-?1 can significantly stimulate the expression of miR-125a-5p,while IL-6 can not.In addition,agomir of miR-125a-5p can significantly inhibit the activation of IL-6/STAT3/Bcl-2 pathway,while antagomir of miR-125a-5p can significantly stimulate the activation of IL-6/STAT3/Bcl-2 pathwayConclusions:miR-125a-5p can inhibit the proliferation and promote the apoptosis of PASMCs through the TGF-?1 and IL-6 singal pathways.There may be a negative feedback regulation between miR-125a-5p and TGF-?1. |
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