内质网应激促进肺动脉平滑肌细胞的表型转化

目的:探讨内质网应激(ERS)对肺动脉平滑肌细胞表型转化的影响。方法:采用胶原酶Ⅰ消化法培养原代大鼠肺动脉平滑肌细胞(PASMCs),用衣霉素(TM)或4-苯基丁酸(4-PBA)诱导或抑制内ERS,MTS法评价细胞增殖情况,western blot和定量RT-PCR检测蛋白和mRNA表达情况。结果:TM呈浓度依赖性诱导内质网应激标志物GRP78和XBP1 mRNA表达;较低浓度的TM促进PASMCs增殖,高浓度(5μg/mol)使细胞凋亡;TM使PASMCs表达SM22 alpha减少,分泌Ⅰ型胶原增加;4-PBA预处理可逆转TM诱导PASMCs的SM22 alpha减少和Ⅰ型胶原分泌增加。结论:内质网应激促进肺动脉平滑肌细胞表型转化,可能是内质网应激参与肺动脉高压的机制之一。     

Glc-6-PD and PKG contribute to hypoxia-induced decrease in smooth muscle cell contractile phenotype proteins in pulmonary artery

Persistent hypoxic pulmonary vasoconstriction (HPV) plays a significant role in the pathogenesis of pulmonary hypertension, which is an emerging clinical problem around the world. We recently showed that hypoxia-induced activation of glucose-6-phosphate dehydrogenase (Glc-6-PD) in pulmonary artery smooth muscle links metabolic changes within smooth muscle cells to HPV and that inhibition of Glc-6PD reduces acute HPV. Here, we demonstrate that exposing pulmonary arterial rings to hypoxia (20-30 Torr) for 12 h in vitro significantly (P < 0.05) reduces (by 30-50%) SM22α and smooth muscle myosin heavy chain expression and evokes HPV. Glc-6-PD activity was also elevated in hypoxic pulmonary arteries. Inhibition of Glc-6-PD activity prevented the hypoxia-induced reduction in SM22α expression and inhibited HPV by 80-90% (P < 0.05). Furthermore, Glc-6-PD and protein kinase G (PKG) formed a complex in pulmonary artery, and Glc-6-PD inhibition increased PKG-mediated phosphorylation of VASP (p-VASP). In turn, increasing PKG activity upregulated SM22α expression and attenuated HPV evoked by Glc-6-PD inhibition. Increasing passive tension (from 0.8 to 3.0 g) in hypoxic arteries for 12 h reduced Glc-6-PD, increased p-VASP and SM22α levels, and inhibited HPV. The present findings indicate that increases in Glc-6-PD activity influence PKG activity and smooth muscle cell phenotype proteins, all of which affect pulmonary artery contractility and remodeling.     

Over-expression of PKGIα inhibits hypoxia-induced proliferation, Akt activation, and phenotype modulation of human PASMCs: the role of phenotype modulation of PASMCs in pulmonary vascular remodeling

The proliferation of pulmonary artery smooth muscle cells (PASMCs) plays a role in pulmonary vascular remodeling (PVR). Recently, it was shown that vascular smooth muscular cell phenotype modulation is important for their proliferation in other diseases. However, little is known about the role of human PASMC phenotype modulation in the proliferation induced by hypoxia and its molecular mechanism during PVR. In this study, we found using primary cultured human PASMCs that hypoxia suppressed the expression of endogenous PKGIα, which was reversed by transfection with a recombinant adenovirus containing the full-length cDNA of PKGIα (Ad-PKGIα). Ad-PKGIα transfection significantly attenuated the hypoxia-induced downregulation of the expression of smooth muscle α-actin (SM-α-actin), myosin heavy chain (MHC) and calponin in PASMCs, indicating that hypoxia-induced phenotype modulation was blocked. Furthermore, flow cytometry and ³H-TdR incorporation demonstrated that hypoxia-induced PASMC proliferation was suppressed by upregulation of PKGIα. These results suggest that enhanced PKGIα expression inhibited hypoxia-induced PASMC phenotype modulation and that it could reverse the proliferation of PASMCs significantly. Moreover, our previous work has demonstrated that Akt protein is activated in the process of hypoxia-induced proliferation of human PASMCs. Interestingly, we found that Akt was not activated by hypoxia when PASMC phenotype modulation was blocked by Ad-PKGIα. This result suggests that blocking phenotype modulation might be a key up-stream regulatory target.     

Atrial natriuretic peptide in hypoxia

A growing number of mammalian genes whose expression is inducible by hypoxia have been identified. Among them, atrial natriuretic peptide (ANP) synthesis and secretion is increased during hypoxic exposure and plays an important role in the normal adaptation to hypoxia and in the pathogenesis of cardiopulmonary diseases, including chronic hypoxia-induced pulmonary hypertension and vascular remodeling, and right ventricular hypertrophy and right heart failure. This review discusses the roles of ANP and its receptors in hypoxia-induced pulmonary hypertension. We and other investigators have demonstrated that ANP gene expression is enhanced by exposure to hypoxia and that the ANP so generated protects against the development of hypoxic pulmonary hypertension. Results also show that hypoxia directly stimulates ANP gene expression and ANP release in cardiac myocytes in vitro. Several cis-responsive elements of the ANP promoter are involved in the response to changes in oxygen tension. Further, the ANP clearance receptor NPR-C, but not the biological active NPR-A and NPR-B receptors, is downregulated in hypoxia adapted lung. Hypoxia-sensitive tyrosine kinase receptor-associated growth factors, including fibroblast growth factor (FGF) and platelet derived growth factor (PDGF)-BB, but not hypoxia per se, inhibit NPR-C gene expression in pulmonary arterial smooth muscle cells in vitro. The reductions in NPR-C in the hypoxic lung retard the clearance of ANP and allow more ANP to bind to biological active NPR-A and NPR-B in the pulmonary circulation, relaxing preconstricted pulmonary vessels, reducing pulmonary arterial pressure, and attenuating the development of hypoxia-induced pulmonary hypertension and vascular remodeling.     

内质网应激在大鼠低氧高二氧化碳性肺动脉高压中的作用

目的：观察低氧高二氧化碳性肺动脉高压大鼠的肺血管重塑并探讨内质网应激（ERS）在肺动脉高压中的作用。方法：将40只SD大鼠随机分为四组：常氧对照组（N）、低氧高二氧化碳组（HH）、ERS通路抑制剂4-苯基丁酸（4-phenylbutyric acid）组（4-PBA）、ERS通路激动剂衣霉素（tunicamycin）组（TM）,n=10。测量各组大鼠的肺动脉平均压（mPAP）、颈动脉平均压以及右心室肥大指数,免疫荧光α-SMA标记法鉴定各组肺中小动脉平滑肌细胞,电镜观察肺组织及肺中小动脉形态学变化,原位末端标记法（TUNEL）检测各组肺动脉平滑肌细胞的凋亡指数,采用RT-PCR和Western blot分别检测各组大鼠葡萄糖调节蛋白78（GRP78）、C/EBP同源蛋白（CHOP）、c-Jun氨基末端激酶（JNK）、天冬氨酸特异性半胱氨酸蛋白酶-12（caspase-12）mRNA及蛋白质表达。结果：①与N组相比,HH组、4-PBA组、TM组mPAP、右心室游离壁重量/左心室加心室间隔重量[RV/（LV+S）]、肺动脉管壁面积/管总面积（WA/TA）比值增加（P<0.0 1）,肺动脉管腔面积/管总面积（LA/TA）比值减小（P<0.01）,细胞凋亡指数降低（P <0.05或P<0.01）。ERS相关蛋白质及mRNA的表达量升高,各差异均有统计学意义。②与HH组相比,4-PB A组mPAP和[RV/（LV+S）]、WA/TA值减小（P<0.01）,LA/TA值和细胞凋亡指数上升（P<0.05或P<0.01）,ERS相关蛋白质和mRNA的表达量均下调（P<0.05或P<0.01）;③与HH组相比,TM组mPAP、[RV/（LV+S）]、WA/TA值升高（P<0.05或P<0.01）;肺动脉中膜层增厚,LA/TA值和细胞凋亡指数降低（P<0.01）。ERS相关蛋白质及mRNA的表达量均升高,除GRP78蛋白质表达量无明显变化外,其余各差异均有统计学意义。结论：低氧高二氧化碳诱导的肺动脉高压大鼠肺血管重塑可能与肺动脉平滑肌细胞增殖过度及凋亡过少有关;ERS相关因子（JNK、caspase-12和CHOP）参与低氧高二氧化碳性肺动脉高压的调控。     

A HIF-1alpha-related gene involved in cell protection from hypoxia by suppression of mitochondrial function

Recently, we reported that acetylcholine-induced hypoxia-inducible factor-1alpha protects cardiomyocytes from hypoxia; however, the downstream factors reducing hypoxic stress are unknown. We identified apoptosis inhibitor (AI) gene as being differentially expressed between von Hippel Lindau (VHL) protein-positive cells with high levels of GRP78 expression and VHL-negative cells with lower GRP levels, using cDNA subtraction. AI decreased GRP78 level, suppressed mitochondrial function, reduced oxygen consumption and, ultimately, suppressed hypoxia-induced apoptosis. By contrast, knockdown of the AI gene increased mitochondrial function. Hypoxic cardiomyocytes and ischemic myocardium showed increased AI mRNA expression. These findings suggest that AI is involved in suppressing mitochondrial function, thereby leading to cellular stress eradication and consequently to protection during hypoxia.     

Down-regulation of lncRNA MEG3 promotes hypoxia-induced human pulmonary artery smooth muscle cell proliferation and migration via repressing PTEN by sponging miR-21

Hypoxia-induced pulmonary hypertension is a life-threatening disease arising from a progressive increase in pulmonary vascular resistance, irreversible pulmonary vascular remodeling and resulting in right ventricular failure. Recent studies suggested that pulmonary artery smooth muscle cell proliferation and migration played an important role in the pathogenesis of hypoxia-induced pulmonary hypertension. However, the mechanisms of hypoxia-induced pulmonary hypertension are complicated and largely unclear. In this study, we discovered that lncRNA MEG3 was down-regulated in human pulmonary artery smooth muscle cell in hypoxia, and inhibition of MEG3 promoted the cell proliferation and cell migration in both normal and hypoxia condition. Further study demonstrated that MEG3 exerted its function via regulation of miR-21 expression in both normal and hypoxia condition. In addition, we displayed the modulation of PTEN by miR-21 and their role in hypoxia. Ultimately, our study illustrated that MEG3 exerts its role via miR-21/PTEN axis in human pulmonary artery smooth muscle cell under both normal and hypoxia conditions.     

Regulation of S100A4 expression via the JAK2-STAT3 pathway in rhomboid-phenotype pulmonary arterial smooth muscle cells exposure to hypoxia

To investigate the effect of JAKs-STATs signal pathway on expression of S100A4 in pulmonary arterial smooth muscle cells (PASMCs), the action of S100A4 and hypoxia induced factor 1 (HIF-1) on the proliferation of hypoxic PASMCs. The results showed that S100A4 immunostaining was localized in the cytoplasm and nuclei of PASMCs exposure to hypoxia and it was predominantly expressed in rhomboid cells (R-SMCs). The mRNA and protein levels of S100A4 expression increased in PASMCs after hypoxic stimulus for 4, 8, 16 h. The immunofluorescence intensity and protein levels of S100A4 were suppressed, and the number of R-SMCs was reduced, when pretreatment with HIF-1α siRNA, STAT3 siRNA, S100A4 siRNA, and S100A4 inhibitor NSC 95397. Pretreatment with HIF-1α siRNA and anti-IL-6 antibodies, the levels of phospho-JAK2, -STAT3, and S100A4 were decreased, while HIF-1α kept stable in hypoxic cells. Importantly, pretreatment with HIF-1α siRNA, anti-IL-6 antibodies, STAT3 siRNA, and S100A4 siRNA, significantly attenuated the proliferation of PASMCs exposure to hypoxia. These data demonstrate that S100A4 is predominantly expressed in hypoxic R-SMCs, and regulated by the activation of JAK2-STAT3 signal pathway, which is dependent on hypoxia-induced HIF-1α expression. These results suggest that JAK2-STAT3 and HIF-1α could serve as targets for the regulation of phenotype modulation of PASMCs during the process of pulmonary vessel lesions.     

Hypoxia induces Kv channel current inhibition by increased NADPH oxidase-derived reactive oxygen species

There is current discussion whether reactive oxygen species are up- or downregulated in the pulmonary circulation during hypoxia, from which sources (i.e., mitochondria or NADPH oxidases) they are derived, and what the downstream targets of ROS are. We recently showed that the NADPH oxidase homolog NOX4 is upregulated in hypoxia-induced pulmonary hypertension in mice and contributes to the vascular remodeling in pulmonary hypertension. We here tested the hypothesis that NOX4 regulates K(v) channels via an increased ROS formation after prolonged hypoxia. We showed that (1) NOX4 is upregulated in hypoxia-induced pulmonary hypertension in rats and isolated rat pulmonary arterial smooth muscle cells (PASMC) after 3days of hypoxia, and (2) that NOX4 is a major contributor to increased reactive oxygen species (ROS) after hypoxia. Our data indicate colocalization of K(v)1.5 and NOX4 in isolated PASMC. The NADPH oxidase inhibitor and ROS scavenger apocynin as well as NOX4 siRNA reversed the hypoxia-induced decrease in K(v) current density whereas the protein levels of the channels remain unaffected by siNOX4 treatment. Determination of cysteine oxidation revealed increased NOX4-mediated K(v)1.5 channel oxidation. We conclude that sustained hypoxia decreases K(v) channel currents by a direct effect of a NOX4-derived increase in ROS.     

Differential effects of TGF-beta1 and BMP-4 on the hypoxic induction of cyclooxygenase-2 in human pulmonary artery smooth muscle cells

Chronic hypoxia-induced pulmonary hypertension results partly from proliferation of smooth muscle cells in small peripheral pulmonary arteries. Previously, we demonstrated that hypoxia modulates the proliferation of human peripheral pulmonary artery smooth muscle cells (PASMCs) by induction of cyclooxygenase-2 (COX-2) and production of antiproliferative prostaglandins. The transforming growth factor (TGF)-beta superfamily plays a critical role in the regulation of pulmonary vascular remodeling, although to date an interaction with hypoxia has not been examined. We therefore investigated the pathways involved in the hypoxic induction of COX-2 in peripheral PASMCs and the contribution of TGF-beta1 and bone morphogenetic protein (BMP)-4 in this response. In the present study, we demonstrate that hypoxia induces activation of p38MAPK, ERK1/2, and Akt in PASMCs and that these pathways are involved in the hypoxic regulation of COX-2. Whereas inhibition of p38(MAPK) or ERK1/2 activity suppressed hypoxic induction of COX-2, inhibition of the phosphoinositide 3-kinase pathway enhanced hypoxic induction of COX-2. Furthermore, exogenous TGF-beta1 induced COX-2 mRNA and protein expression, and our findings demonstrate that release of TGF-beta1 by PASMCs during hypoxia contributes to the hypoxic induction of COX-2 via the p38MAPK pathway. In contrast, BMP-4 inhibited the hypoxic induction of COX-2 by an MAPK-independent pathway. Together, these findings suggest that the TGF-beta superfamily is part of an autocrine/paracrine system involved in the regulation of COX-2 expression in the distal pulmonary circulation, and this modulates hypoxia-induced pulmonary vascular cell proliferation.     

Calumenin has a role in the alleviation of ER stress in neonatal rat cardiomyocytes

Disturbance of endoplasmic reticulum (ER) homeostasis causes ER stress (ERS), and triggers the unfolded protein response (UPR) that consequently reduces accumulation of unfolded proteins by increasing the quantity of ER chaperones. Calumenin, a Ca²⁺-binding protein with multiple EF hand motifs, which is located in the ER/SR, is highly expressed during the early developmental stage of the heart, similar to other ER-resident chaperones. The aim of this study was to investigate the functional role of calumenin during ERS in the heart. Like other chaperones (e.g., GRP94 and GRP78), calumenin expression was highly upregulated during ERS induced by 10 μg/ml tunicamycin, but attenuated in the presence of 500 μM PBA, the chemical chaperone in neonatal rat ventricular cardiomyocytes (NRVCs). Upon 7.5-fold overexpression of calumenin using a recombinant adenovirus system, the expression levels of ERS markers (GRP78, p-PERK, and p-elF2α) and ER-initiated apoptosis markers (CHOP and p-JNK) were reduced, whereas the survival protein BCL-2 was upregulated during ERS compared to the control. Evaluation of cell viability by TUNEL assay showed that apoptosis was also significantly reduced by calumenin overexpression in ERS-induced cells. Taken together, our results suggest that calumenin plays an essential role in the alleviation of ERS in neonatal rat cardiomyocytes.     

Modulation of pulmonary vascular smooth muscle cell phenotype in hypoxia: role of cGMP-dependent protein kinase

Chronic hypoxia triggers pulmonary vascular remodeling, which is associated with a modulation of the vascular smooth muscle cell (SMC) phenotype from a contractile, differentiated to a synthetic, dedifferentiated state. We previously reported that acute hypoxia represses cGMP-dependent protein kinase (PKG) expression in ovine fetal pulmonary venous SMCs (FPVSMCs). Therefore, we tested if altered expression of PKG could explain SMC phenotype modulation after exposure to hypoxia. Hypoxia-induced reduction in PKG protein expression strongly correlated with the repressed expression of SMC phenotype markers, myosin heavy chain (MHC), calponin, vimentin, alpha-smooth muscle actin (alphaSMA), and thrombospondin (TSP), indicating that hypoxic exposure of SMC induced phenotype modulation to dedifferentiated state, and PKG may be involved in SMC phenotype modulation. PKG-specific small interfering RNA (siRNA) transfection in FPVSMCs significantly attenuated calponin, vimentin, and MHC expression, with no effect on alphaSMA and TSP. Treatment with 30 microM Drosophila Antennapedia (DT-3), a membrane-permeable peptide inhibitor of PKG, attenuated the expression of TSP, MHC, alphaSMA, vimentin, and calponin. The results from PKG siRNA and DT-3 studies indicate that hypoxia-induced reduction in protein expression was also similarly impacted by PKG inhibition. Overexpression of PKG in FPVSMCs by transfection with a full-length PKG construct tagged with green fluorescent fusion protein (PKG-GFP) reversed the effect of hypoxia on the expression of SMC phenotype marker proteins. These results suggest that PKG could be one of the determinants for the expression of SMC phenotype marker proteins and may be involved in the maintenance of the differentiated phenotype in pulmonary vascular SMCs in hypoxia.     

Hypoxia-induced migration in pulmonary arterial smooth muscle cells requires calcium-dependent upregulation of aquaporin 1

Pulmonary arterial smooth muscle cell (PASMC) migration is a key component of the vascular remodeling that occurs during the development of hypoxic pulmonary hypertension, although the mechanisms governing this phenomenon remain poorly understood. Aquaporin-1 (AQP1), an integral membrane water channel protein, has recently been shown to aid in migration of endothelial cells. Since AQP1 is expressed in certain types of vascular smooth muscle, we hypothesized that AQP1 would be expressed in PASMCs and would be required for migration in response to hypoxia. Using PCR and immunoblot techniques, we determined the expression of AQPs in pulmonary vascular smooth muscle and the effect of hypoxia on AQP levels, and we examined the role of AQP1 in hypoxia-induced migration in rat PASMCs using Transwell filter assays. Moreover, since the cytoplasmic tail of AQP1 contains a putative calcium binding site and an increase in intracellular calcium concentration ([Ca(2+)](i)) is a hallmark of hypoxic exposure in PASMCs, we also determined whether the responses were Ca(2+) dependent. Results were compared with those obtained in aortic smooth muscle cells (AoSMCs). We found that although AQP1 was abundant in both PASMCs and AoSMCs, hypoxia selectively increased AQP1 protein levels, [Ca(2+)](i), and migration in PASMCs. Blockade of Ca(2+) entry through voltage-dependent Ca(2+) or nonselective cation channels prevented the hypoxia-induced increase in PASMC [Ca(2+)](i), AQP1 levels, and migration. Silencing AQP1 via siRNA also prevented hypoxia-induced migration of PASMCs. Our results suggest that hypoxia induces a PASMC-specific increase in [Ca(2+)](i) that results in increased AQP1 protein levels and cell migration.     

Targeting mitochondria-associated membranes as a potential therapy against endothelial injury induced by hypoxia

Mitochondrial dysfunction plays a principal role in hypoxia-induced endothelial injury, which is involved in hypoxic pulmonary hypertension and ischemic cardiovascular diseases. Recent studies have identified mitochondria-associated membranes (MAMs) that modulate mitochondrial function under a variety of pathophysiological conditions such as high-fat diet-mediated insulin resistance, hypoxia reoxygenation-induced myocardial death, and hypoxia-evoked vascular smooth muscle cell proliferation. However, the role of MAMs in hypoxia-induced endothelial injury remains unclear. To explore this further, human umbilical vein endothelial cells and human pulmonary artery endothelial cells were exposed to hypoxia (1% O₂) for 24 hours. An increase in MAM formation was uncovered by immunoblotting and immunofluorescence. Then, we performed small interfering RNA transfection targeted to MAM constitutive proteins and explored the biological effects. Knockdown of MAM constitutive proteins attenuated hypoxia-induced elevation of mitochondrial Ca²⁺ and repressed mitochondrial impairment, leading to an increase in mitochondrial membrane potential and ATP production and a decline in reactive oxygen species. Then, we found that MAM disruption mitigated cell apoptosis and promoted cell survival. Next, other protective effects, such as those pertaining to the repression of inflammatory response and the promotion of NO synthesis, were investigated. With the disruption of MAMs under hypoxia, inflammatory molecule expression was repressed, and the eNOS-NO pathway was enhanced. This study demonstrates that the disruption of MAMs might be of therapeutic value for treating endothelial injury under hypoxia, suggesting a novel strategy for preventing hypoxic pulmonary hypertension and ischemic injuries.     

The hypoxia-induced microRNA-130a controls pulmonary smooth muscle cell proliferation by directly targeting CDKN1A

Excessive proliferation of human pulmonary artery smooth muscle cells (HPASMC) is one of the major factors that trigger vascular remodeling in hypoxia-induced pulmonary hypertension. Several studies have implicated that hypoxia inhibits the tumor suppressor p21 (CDKN1A). However, the precise mechanism is unknown.The mouse model of hypoxia-induced PH and in vitro experiments were used to assess the impact of microRNAs (miRNAs) on the expression of CDKN1A. In these experiments, the miRNA family miR-130 was identified to regulate the expression of CDKN1A. Transfection of HPASMC with miR-130 decreased the expression of CDKN1A and, in turn, significantly increased smooth muscle proliferation. Conversely, inhibition of miR-130 by anti-miRs and seed blockers increased the expression of CDKN1A. Reporter gene analysis proved a direct miR-130–CDKN1A target interaction. Exposure of HPASMC to hypoxia was found to induce the expression of miR-130 with concomitant decrease of CDKN1A. These findings were confirmed in the mouse model of hypoxia-induced pulmonary hypertension showing that the use of seed blockers against miR-130 restored the expression of CDKN1A.These data suggest that miRNA family miR-130 plays an important role in the repression of CDKN1A by hypoxia. miR-130 enhances hypoxia-induced smooth muscle proliferation and might be involved in the development of right ventricular hypertrophy and vascular remodeling in pulmonary hypertension.     

Altered pulmonary vasoreactivity in the chronically hypoxic lung

Prolonged exposure to alveolar hypoxia induces physiological changes in the pulmonary vasculature that result in the development of pulmonary hypertension. A hallmark of hypoxic pulmonary hypertension is an increase in vasomotor tone. In vivo, pulmonary arterial smooth muscle cell contraction is influenced by vasoconstrictor and vasodilator factors secreted from the endothelium, lung parenchyma and in the circulation. During chronic hypoxia, production of vasoconstrictors such as endothelin-1 and angiotensin II is enhanced locally in the lung, while synthesis of vasodilators may be reduced. Altered reactivity to these vasoactive agonists is another physiological consequence of chronic exposure to hypoxia. Enhanced contraction in response to endothelin-1 and angiotensin II, as well as depressed vasodilation in response to endothelium-derived vasodilators, has been documented in models of hypoxic pulmonary hypertension. Chronic hypoxia may also have direct effects on pulmonary vascular smooth muscle cells, modulating receptor population, ion channel activity or signal transduction pathways. Following prolonged hypoxic exposure, pulmonary vascular smooth muscle exhibits alterations in K+ current, membrane depolarization, elevation in resting cytosolic calcium and changes in signal transduction pathways. These changes in the electrophysiological parameters of pulmonary vascular smooth muscle cells are likely associated with an increase in basal tone. Thus, hypoxia-induced modifications in pulmonary arterial myocyte function, changes in synthesis of vasoactive factors and altered vasoresponsiveness to these agents may shift the environment in the lung to one of contraction instead of relaxation, resulting in increased pulmonary vascular resistance and elevated pulmonary arterial pressure.     

Regulatory effect of AMP-activated protein kinase on pulmonary hypertension induced by chronic hypoxia in rats: in vivo and in vitro studies

Activation of AMP-activated protein kinase (AMPK) plays an important role in cardiovascular protection. It can inhibit arterial smooth muscle cell proliferation and cardiac fibroblast collagen synthesis induced by anoxia. However, the role of AMPK-dependent signalling cascades in the pulmonary vascular system is currently unknown. This study aims to determine the effects of AMPK on pulmonary hypertension and pulmonary vessel remodelling induced by hypoxia in rats using in vivo and in vitro studies. In vivo study: pulmonary hypertension, right ventricular hypertrophy and pulmonary vascular remodelling were found in hypoxic rats. Meanwhile, AMPKα₁ and phosphorylated AMPKα₁ were increased markedly in pulmonary arterioles and lung tissues. Mean pulmonary arterial pressure, index of right ventricular hypertrophy and parameters of pulmonary vascular remodelling, including vessel wall area/total area, density of nuclei in medial smooth muscle cells, and thickness of the medial smooth muscle cell layer were markedly suppressed by AICAR, an AMPK agonist. In vitro study: the expression of AMPKα₁ and phosphorylated AMPKα₁ was increased in pulmonary artery smooth muscle cells (PASMCs) under hypoxic conditions. The effects of PASMC proliferation stimulated by hypoxia were reinforced by treatment with Compound C, an AMPK inhibitor. AICAR inhibited the proliferation of PASMCs stimulated by hypoxia. These findings suggest that AMPK is involved in the formation of hypoxia-induced pulmonary hypertension and pulmonary vessel remodelling. Up-regulating AMPK can contribute to decreasing pulmonary vessel remodelling and pulmonary hypertension induced by hypoxia.     

p53 Gene deficiency promotes hypoxia-induced pulmonary hypertension and vascular remodeling in mice

Chronic hypoxia induces pulmonary arterial remodeling, resulting in pulmonary hypertension and right ventricular hypertrophy. Hypoxia has been implicated as a physiological stimulus for p53 induction and hypoxia-inducible factor-1α (HIF-1α). However, the subcellular interactions between hypoxic exposure and expression of p53 and HIF-1α remain unclear. To examine the role of p53 and HIF-1α expression on hypoxia-induced pulmonary arterial remodeling, wild-type (WT) and p53 knockout (p53KO) mice were exposed to either normoxia or hypoxia for 8 wk. Following chronic hypoxia, both genotypes demonstrated elevated right ventricular pressures, right ventricular hypertrophy as measured by the ratio of the right ventricle to the left ventricle plus septum weights, and vascular remodeling. However, the right ventricular systolic pressures, the ratio of the right ventricle to the left ventricle plus septum weights, and the medial wall thickness of small vessels were significantly greater in the p53KO mice than in the WT mice. The p53KO mice had lower levels of p21 and miR34a expression, and higher levels of HIF-1α, VEGF, and PDGF expression than WT mice following chronic hypoxic exposure. This was associated with a higher proliferating cell nuclear antigen expression of pulmonary artery in p53KO mice. We conclude that p53 plays a critical role in the mitigation of hypoxia-induced small pulmonary arterial remodeling. By interacting with p21 and HIF-1α, p53 may suppress hypoxic pulmonary arterial remodeling and pulmonary arterial smooth muscle cell proliferation under hypoxia.