Research On The Role Of The HIF-1α And Its Target Genes In The Pathogenesis Of Neonatal Hypoxic Pulmonary Hypertension

Objective:1. To monitor the changes the serum HIF-la, ET-1, ADM and the expression of HIF-1a, ET-1, ADM’s mRNA and protein in lung tissue of neonatal rats with HPH and to infer the role of HIF-1a and its target genes in the pathogenesis of neonatal HPH.2. To pinpoint the watershed of vascular remodeling as to provide theoretical basis for early clinical treatment of HPH by observing the changes of the mPAP, HPVR index and RVHI at different hypoxic time.3.To monitor the changes of serum HIF-la, ET-1and ADM, the changes of the PASP, RVEF, E/A and right ventricular Tei index in hypoxic and normal neonates by using echocardiography methods, in order to reveal the pathogenesis of hypoxic pulmonary hypertension in neonate, so providing the preliminary theoretical basis for clinical treatment.Methods:120newborn rat were randomly divided into hypoxic group (hypoxia exposure) and normoxic group. The mean pulmonary arteria pressure (mPAP), concentrations of in the serum and expression levels in the lung tissue of HIF-1a, ET-1, ADM were measured in these neonatal rats after3,5,7,10,14and21days, including hypoxic group and normoxic group respectively.117cases of newborns were included in the present study, they were chosen completely at random.85cases of newborns were hypoxic pulmonary hypertension (HPH) group, while32cases of newborns without symptoms of hypoxia were control group. Group of newborns with HPH will be divided into three subgroups, including mild HPH group of38cases (40mmHg≤PASP<50mmHg), moderate HPH group of27cases (50mmHg <PASP<70mmHg), severe HPH group of20cases (PASP>70mmHg).1. Building the model of neonatal rat with HPH, the serum HIF-1α, ET-1,ADM concentrations were measured by the ELISA method and mPAP were determinate by direct manometry aiming in understanding the relationship between three factors and the mPAP of neonatal rat with HPH.2. The intima-media thickness/external diameter ratio (MT%) and the medial wall cross-sectional area/vessel total cross-sectional area (MA%) were calculated to evaluate the thickening of the walls of the pulmonary arterioles by optical microscope and electron microscope. By semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) method, immunohistochemical staining and western blotting to detect HIF-1α, ET-1,ADM mRNA and protein expression in the lung tissue, which compared HIF-1a, ET-1,ADM mRNA and protein expression of the similarities and differences between the hypoxic group and the control group.3.117cases of newborns of the HPH three groups and the control group were conducted echocardiography of PASP, RVE, E/A and Tei-index respectively. While they were determined concentrations of HIF-1a, ET-1,ADM with ELASA, understanding the relevance of HIF-1a, ET-1, ADM and PASP in HPH neonates.Results:1. Compared with the normoxic group, the mPAP increased with the prolonged hypoxic time in all of the hypoxic group(P<0.05).2. Concentrations of HIF-1a in the serum were higher than that of the normoxic group at3,5,7,10,14days(P<0.05), ET-1elevated significantly in all of the hypoxic group, and the serum concentration of ADM in the hypoxic group was higher than that of the normoxic group at3and5days but lower than the normoxic group after14and21days of hypoxia. Expression levels in the lung tissue of HIF-la mRNA, ET-1mRNA and ADM mRNA increased only at3and5days of hypoxia. Protein expression levels in the lung tissue of HIF-1a elevated significantly in all of the hypoxic group, Protein expression levels of ET-1were higher than that of the normoxic group at3,5,7,10,14days(P<0.05), and ADM protein levels were alike with ADM mRNA.3. Newborn rats of Hypoxia group at7days of hypoxia began to appear pulmonary vascular morphology, ultrastructure changes and right ventricular hypertrophy.4. PASP of three case groups after1day are higher than in the normoxic group. To mild to moderate group with the first3day after birth, hypoxia pointer return to normal, PASP resume in7day after birth. To severe group, hypoxia pointer restore to normal in7day after birth, PASP is still higher than normal. RVEF and E/A of right ventricular reduced at severe HPH group. Tei index of the right ventricle showed that Tei index were higher in case groups than the control group, PASP and Tei index of right ventricular was positively related.5. HIF-1α、ET-1levels of three case group increased significantly1day after birth, and aggravated with the increased level of PASP. The serum concentration of HIF-la and ADM in severe HPH, and ET-1in moderate and severe HPH were higher than control group in3day. In7day after birth, only ET-1in severe HPH increased. Correlation coefficient of three indicators in the HIF-1a and PASP was biggest. Relative to HIF-1a and ET-1, change of ADM level was weaker. Conclusion:1.Animal experiment hints hypoxia activate HIF-1a expression in the activation of the transcription level ET and ADM of gene expression, which participates in pathogenesis of pulmonary hypertension.2. Hypoxia-induced pulmonary vascular spasm happened in the early time, which can resume by treatment timely, but if the pulmonary artery pressure was still high in7days after birth (especially in the stage PPHN), was difficulty in treatment. Echocardiography can be used convenient, timely and accurate to diagnosis of neonatal HPH and PPHN.3. HPH was correlated to HIF-1a, ET-1, ADM level in serum. We can speculate that hypoxia can improve a expression of HIF-1a, causing increase of ET-1and ADM, HPH occurred, so it is involved in the pathogenesis of neonatal HPH.4. RVEF and E/A of Newborn were broadened only in severe HPH and Tei index can be sensitive to find mild, moderate and severe HPH with right ventricular dysfunction.