Protective Effect Of Estrogen On Hypoxic Pulmonary Hypertension And Effects Of Different Phenotypes Of Macrophages

Objective: The pathogenesis of pulmonary hypertension is complex and involves many cellular pathways.However,the role of pulmonary macrophages in the pathogenesis of pulmonary hypertension is still unclear.This study mainly explored the protective effect of estrogen on hypoxic pulmonary hypertension and the changes of macrophage phenotype during this process.Methods:Thirty-six female Sprague-Dawley rats were randomly divided into 6 groups according to random number table(n=6 in each group): sham operation + normoxia group,sham operation + hypoxia group,castration + normoxia group,castration + hypoxia group,castration + normoxia + 17β estradiol(E2)group,castration + hypoxia + E2 group.The ovariectomized group was surgically removed from the ovary;the non-castrated group(sham-operated group)opened the abdominal cavity to find the ovary and did not treat it,and directly sutured it.The hypoxic group was placed in a hypoxic chamber,and the normox group was given normal air.The castration + normoxia + E2 group and the castration + hypoxia + E2 group were given subcutaneous injection of E2 [20ug/(kg·d)],and the other groups were injected subcutaneously with normal saline(0.1ml/d).A model of hypoxic pulmonary hypertension was established for 8 weeks.The mean pulmonary artery pressure(mPAP)of each group of animals was measured by right heart catheterization and the left and right ventricles and ventricular septum were weighed to calculate the right ventricular hypertrophy index(RVHI).The expression of M1 and M2 macrophages was detected by immunofluorescence.The expression levels of iNOS and CD206 in M1 and M2 macrophages were detected by RT-PCR and Western blot.The expressions of iNOS and CD206 mRNA and protein were detected by RT-PCR and Western blot.Results:1.0 successfully established the HPAH rat model:Compared with the sham operation + normoxia group,the sham operation + hypoxia group and the castration + hypoxia group were slow to respond with prolonged hypoxic time,dark coat color,rapid breathing,and gradually reduced water intake.Status,no death specimens.The growth rate of castration + hypoxia + E2 group also slowed down.The activity was slightly better than that of sham operation + hypoxia group and castration + hypoxia group.Compared with sham operation group,castration + normoxia group,go The change of potential + normoxia + E2 group was not obvious.2.mPAP and RVHI for each group of samples:Compared with sham operation + normoxia group,castration + normoxia group,castration + normoxia + E2,mPAP and RVHI were increased in sham operation + hypoxia group and castration + hypoxia group(P<0.05),and in line with the pressure level of pulmonary hypertension,mPAP and RVHI in the castration + hypoxia + E2 group were also relatively elevated(P<0.05),but lower than castration + hypoxia group and sham operation + hypoxia group,suggesting that estrogen can improve hypoxic pulmonary hypertension.3.Morphological changes in lung tissue of each group:Compared with sham operation + normoxia group,castration + normoxia group,castration + normoxia + E2,sham operation + hypoxia group and castration + hypoxia group pulmonary artery wall thickening,arterial lumen is narrow,inflammation The cell infiltration was severe,which was consistent with the morphological changes of pulmonary hypertension.In the castration + hypoxia + E2 group,the castration + hypoxia group and the sham operation + hypoxia group showed different degrees of reversal of the pulmonary artery lumen,and the inflammatory cell infiltration was relatively reduced.4.Immunofluorescence staining to identify the expression of MI-type macrophages and M2 macrophages in lung tissue of each group:The relative expression of iNOS protein in M1 macrophage surface markers in ovariectomized + hypoxia group was significantly higher than that in sham operation + normoxia group(P<0.05).The expression of iNOS protein in sham operation + hypoxia group was lower than that in castration + hypoxia group(P<0.05),and the expression of iNOS protein in castration + hypoxia + E2 group was lower than that in castration + hypoxia group(P<0.05).The relative expression of surface marker CD206 protein in M2 macrophages in castration + hypoxia group was higher than that in sham operation + normoxia group(P<0.05).The relative expression of CD206 protein in castration+ hypoxia+E2 group was higher than that in castration+ hypoxia group(P<0.05).5.RT-PCR and Western blot were used to detect the expression levels of iNOS and CD206 mRNA and protein in lung tissue of each group:Compared with the normoxic group,the iNOS mRNA and protein levels in the hypoxic group were significantly increased.After the E2 intervention,the iNOS mRNA and protein levels were lower in the oxygen + castration group(P<0.05),but still higher than In the normoxic group(P<0.05),the expression of CD206 mRNA and protein in lung tissue of hypoxic group was also higher than that in normoxic group.After E2 intervention,CD206 mRNA and protein levels were lower in oxygen + castration group(P<0.05).Conclusions:Long-term chronic hypoxia can induce the occurrence of HPAH.During this process,the expression of M1 macrophage and M2 macrophage increased,and estrogen may improve by decreasing the expression of M1 macrophages and promoting the polarization of M2 cells.pulmonary hypertension.