| Background: Pulmonary hypertension(PH)is a severe lethal disease characterized by pulmonary vascular remodeling,which causes a sustained increase in pulmonary arterial pressure,leading to right ventricular failure and death.Although the current targeted drug therapy for pulmonary hypertension has made great progress,it is only dedicated to relaxing blood vessels,failing to reverse PH,and has many side effects and high cost in application.New treatments for the vascular remodeling pathway and new targeted drugs are imperative.In all types of pulmonary hypertension,The incidence of the third type of PH caused by respiratory diseases and hypoxia is increasing year by year.The latest epidemiological data show that the total incidence of COPD in China is 8.6%,of which 30% to 70% of patients with COPD have PH.In patients with obstructive sleep apnea syndrome,the prevalence of pulmonary hypertension can reach 17% to 42%.Such patients with pulmonary hypertension mean high mortality and poor prognosis.Under hypoxic conditions,autophagy can be activated to support cell survival.Microtubule-associated protein light chain 3(LC3)was used as a marker to induce autophagy.However,it is still an unsolved problem of whether autophagy is adaptive or not in the development of PH.At the same time,it has shown that autophagy also plays a key role in macrophage polarization in vitro studies.The formation of hypoxic pulmonary hypertension(HPH)has persistent inflammation,which are mainly caused by macrophages.Markers of macrophages include CD68,CD80,etc.When macrophages are polarized,the markers of cells of the M2 subtype become CD163.The natural hormone estradiol(E2)in the body is considered to be the main physiological estrogen.In recent years,the protective effect of estrogen in pulmonary hypertension has been increasing,and its anti-inflammatory effect can improve pulmonary vascular remodeling and vasodilation,but its mechanism has not been fully elucidated.Objective: 1.We explore the effects of estrogen on autophagy and macrophage polarization in ovariectomized hypoxic pulmonary hypertension rats by the model of hypoxic pulmonary hypertension in ovariectomized rats,providing experimental basis for the use of estrogen as a new,effective,and inexpensive.Anti-pulmonary hypertension drugs with few side effects;2.By a case of POPH treated with sildenafil,we explore the safety and efficacy of various targeted drugs and are further to explore the role of estrogen.Methods: The first part: 1.Animal grouping: After 24 female SD rats were ovariectomized,they were randomly divided into 4 groups: ovariectomized normoxia group,ovariectomized hypoxia group E2,ovariectomized hypoxia group,and castration hypoxia E2 group,with 6 rats in each group.Rats in two E2 intervention groups were injected subcutaneously with E2 20?g/kg daily,and the rats in the other group were injected with the same amount of normal saline.Two hypoxic rats were placed in a hypoxic environment animal test chamber with oxygen concentration(10.0 ± 0.5)%.Sodium lime and anhydrous calcium chloride were used to absorb excess CO2 and water,and the CO2 concentration was controlled to <0.5%.The hypoxia was continued for 24 hours every day;the two normoxic groups were kept in normal air,and the remaining feeding conditions were the same as those in the hypoxic group.2.The general condition of each of groups: After 8 weeks,the rats were weighed;the pulmonary artery pressure was measured by right heart catheterization,then the mean pulmonary artery pressure(m PAP)was calculated;the Right ventricule hypertrophy index(RVHI)was calculated by weighing method.Hypoxic pulmonary artery remodeling(HPSR)was observed by HE staining.3.Determination of LC3,CD68 and CD163 m RNA levels in lung tissue by RT-PCR.4.Determination of LC3,CD68 and CD163 protein levels in lung tissue by Western blot and immunofluorescence.The second part: Through the analysis of the diagnosis and treatment of a case of sildenafil in patients with Caroli disease and portal hypertension in the Second Department of Respiratory Medicine of the Second Hospital of Hebei Medical University,the relevant literature was used to discuss the safety and efficacy of various targeted drugs in clinical practice;and then to further explore the role of estrogen in it.Results: The first part: 1.General condition of the rat: Rats fed under hypoxic conditions gradually showed a decrease in activity and decreased eating,slow response,dull coat,rapid breathing,and weight loss,which were statistically significant(P<0.05).No death occurred during the 8 weeks of hypoxia.2.m PAP,pulmonary small blood vessels and right ventricular morphological changes in rats: Compared with Ovariectomy normoxic?Ovariectomy normoxic+E2,the pulmonary artery remodeling in the Ovariectomy hypoxia and the Ovariectomy hypoxia+E2 group showed thickening of the wall,stenosis of the lumen,and obvious inflammatory cell infiltration around.m PAP,RVHI,WT%,and WA% were all increased(P<0.05);and the above indexes of Ovariectomy hypoxia+E2 group were lower than those in Ovariectomy hypoxia(P<0.05).3.Changes of expression levels of LC3,CD68 and CD163 m RNA in rat lung tissue: The expression of LC3 m RNA was detected by RT-PCR: in contrast with the Ovariectomy normoxic group,the LC3 m RNA was increased in the Ovariectomy hypoxia groups(P<0.05),and the Ovariectomy hypoxia+E2 intervention group also increased,which was higher than Ovariectomy hypoxia(P<0.05),there was no significant change in the Ovariectomy normoxic+E2 intervention group(P>0.05).The expression level of CD68 was significantly higher in the Ovariectomy hypoxia group and the Ovariectomy hypoxic+E2 intervention group than in the Ovariectomy normoxic group(P<0.05),and there was no significant change in the Ovariectomy normoxic E2 intervention group(P>0.05).The expression level of CD163 m RNA was significantly higher in the Ovariectomy hypoxia group than in the Ovariectomy normoxic group(P<0.05),and increased in the Ovariectomy hypoxic+E2 intervention group,which was higher than in the Ovariectomy hypoxia group(P<0.05);there was no significant change in the Ovariectomy normoxia+E2 intervention group(P>0.05).4.Changes of LC3,CD68 and CD163 protein levels in rat lung tissue 4.1 the result of Western blot: In contrast with the Ovariectomy normoxicgroup,the LC3(LC3 ?/?)protein was increased in the Ovariectomy hypoxia groups(P<0.05),and the Ovariectomy hypoxia+E2 intervention group also increased,which was higher than Ovariectomy hypoxia(P<0.05),there was no significant change in the Ovariectomy normoxic+E2 intervention group(P>0.05).The expression level of CD68 was significantly higher in the Ovariectomy hypoxia group and the Ovariectomy hypoxic+E2 intervention group than in the Ovariectomy normoxic group(P<0.05),and there was no significant change in the Ovariectomy normoxic E2 intervention group(P>0.05).The expression level of CD163 protein was significantly higher in the Ovariectomy hypoxia group than in the Ovariectomy normoxic group(P<0.05),and increased in the Ovariectomy hypoxic+E2 intervention group,which was higher than in the Ovariectomy hypoxia group(P<0.05);there was no significant change in the Ovariectomy normoxia+E2 intervention group(P>0.05).4.2 the results of Immunofluorescence: 4.2.1 The expression level of CD68 was significantly higher in the Ovariectomy hypoxia group and the Ovariectomy hypoxic+E2 intervention group than in the Ovariectomy normoxic group(P<0.05),and there was no significant change in the Ovariectomy normoxic E2 intervention group (P>0.05).The changes of LC3 in the CD68-positive group were analyzed.It was found that the expression level of LC3 was higher in the Ovariectomy hypoxia group than in the Ovariectomy normoxic group(P<0.05),and the Ovariectomy hypoxia+E2 intervention group was also increased,which was higher than the Ovariectomy hypoxia group(both P<0.05).4.2.2 The expression level of CD163 protein was significantly higher in the Ovariectomy hypoxia group than in the Ovariectomy normoxic group(P<0.05),and increased in the Ovariectomy hypoxic+E2 intervention group,which was higher than in the Ovariectomy hypoxia group(P<0.05);there was no significant change in the Ovariectomy normoxia+E2 intervention group(P>0.05).The changes of LC3 in the CD68-positive group were analyzed.In contrast with the Ovariectomy normoxic group,the LC3 protein was increased in the Ovariectomy hypoxia groups(P<0.05),and the Ovariectomy hypoxia+E2 intervention group also increased,which was higher than Ovariectomy hypoxia(P<0.05).The second part: After the diagnosis of portal hypertension,the patient was treated with sildenafil.Although the pulmonary artery pressure was not significantly reduced,the clinical symptoms and functional status were relatively stable,and no adverse reactions were observed during the treatment.The decline of estrogen levels may be possible to increase pulmonary artery pressure,but its effects still remain to be further studied.Conclusions: 1.E2 may up-regulate the expression of autophagy in the lung tissue of hypoxic pulmonary hypertension rats,causing macrophage to polarize to M2 type,thereby alleviating pulmonary hypertension.2.Sildenafil is an oral drug with selective activation of pulmonary vasculature and no hepatotoxicity compared with other drugs,and its price is relatively cheap and has few side effects,but its clinical application and long-term application to patients' blood flow still require more data support.3.Decreased estrogen levels may aggravate pulmonary hypertension,but further research is needed. |
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