| Objective To investigate an important cell source for the muscularization of non-muscular pulmonary arterioles in hypoxia-induced pulmonary hypertension(HPH). To investigate the dynamic expression of hypoxia inducible factor1α (HIF-1α) and inducible nitric oxide synthase(iNOS) and transforming growth factor-β1 (TGF-β1) and myofibroblast formation in hypoxic pulmonary vascular remodeling of rats. To investigate cultured human embryonic lung fibroblasts phenotype transforming in hypoxic group and hypoxia+ TGF- β1 group. To investigate the myofibroblast in pulmonary artery of chronic obstructive pulmonary disease (COPD) patients, clarify the roles of myofibroblast in HPH development and provide theoretical basis of cell mechanisms.Methods The study consisted of two parts. 1) Models of chronic HPH rats were duplicated by normobaric anoxia (respired mixted gases containing 10% O2, 8 hours per day for 21 days intermittently). After anoxia for 3d, 7d, 14d amd 21d; mean pulmonary artery pressure (mPAP), was measured by right-heart catheterization; right ventricular hypertrophy index (RVHI) was calculated by the ratio of right ventricle to the left ventricle plus septum, and hypoxic pulmonary vascular remodeling (HPSR) was observed with morphmetric analysis by microscope. Ultrastructural characteristics of adventitial myofibroblast and cell phenotype in alveolar wall vessels were observed by electron microscopy.In situ hybridization, immunohistochemistry and Western blot were used to measure mRNA and protein levels of HIF-1α,TGF-β1 and iNOS in pulmonary artery walls and lung tissue respectively. Furthermore, linear correlation relationshipbetween expression of the genes and mPAP, HPSR, RVHI were analyzed, cultured human embryonic lung fibroblasts(KMB17) phenotype were grouped into: normoxic oxygen group N(20%);hypoxic groupH(l%O2±5%CO2±equilibrium nitrogen);hypoxia± TGF-β1 groupH±T(1% O2±5%CO2±equilibrium nitrogen,the terminal concentration of TGF- β1 was 5ng/mL); KMB17 phenotype was identified by the expression of α-SMA by immunohistochemistry.Results (1) The level of mPAP (18.41 ±0.37) mmHg, the ratio of vascular wall area to the total area(WA%) (52.2 ± 0.8) % and the ratio of vascular lumen area to the total area(LA%) (47.8 ± 0.8) % were significantly higher in the hypoxia 7d group than those in the control group [ (14.02 ±0.41) mmHg, (64.5 ±1.3) %, (35.5 ±1.3) % respectively](P<0.05). These parameters reached a high-level stable phase on hypoxia 14d; RVHI was significantly higher[ (25.0±1.8)%] on hypoxia 14d than that in the control group [ (23.6 ±0.5) %] (P<0.05); (2) The distribution of nonmuscular, partially muscular, and muscular vessels(39%,46%, 15% ) is significantly different (Px2 <0.005) from hypoxia 7d than that in the control group (60%, 35%, 5%). (3) The intra-acinar pulmonary arteries with cells expressing a -SMA increased most with hypoxic time by immunocytochemistry. (4) Thick-walled vessel in late-stage wall remodeling (Day 21) was observed by electron microscopy. Myofibroblast phenotype is organized between elastic laminae. Aligning fibroblasts are associated with the wall.(5) HIF-1α mRNA staining was poorly positive in control, hypoxia for 3 days and hypoxia for 7 days, but began to enhance significantly after 14-day of hypoxia (0.203±0.02, P<0.05), then remained stable. Expression of HIF-1α protein in control was poorly positive , but was up-regulated in pulmonary arterial tunica intima of all hypoxic rats. In pulmonary arterial tunica media, the levels of HIF-1α protein was markedly up-regulated after 3-day (0.198±0.02, p<0.05), reaching its peak after 7-day of hypoxia (0.221±0.02, P<0.05), then tended to decline after 14-day and 21-day of hypoxia. Expression of iNOS protein in control group was poorly positive in pulmonary arterial tunica media, the level of iNOS protein was markedly up-regulated in H3 group (0.225±0.030, p<0.01),...
|
PDF Full Text Request