Silencer-regulated circLSM14A inhibits autophagy of pulmonary artery smooth muscle cells through parental protein LSM14A.

See related article, pages 1109–1114 Idiopathic pulmonary artery hypertension (IPH) is a rare illness with a poor prognosis. Whereas chronic intravenous prostacyclin relieves some of the symptoms of progressive dyspnea and prolongs survival, most patients ultimately require a lung transplant.1 Newer therapies such as nonintravenously administered prostacyclin derivatives,2,3,4 endothelin receptor blockers,5,6 and, to some extent, phosphodiesterase inhibitors,7 hold some promise as alternatives for intravenous prostacyclin, but current expectation is that, like prostacyclin, they will, at best, retard disease progression, serving as a bridge to transplant rather than as an alternative. The pathological features of IPH are loss of small distal precapillary pulmonary arteries, obliterative changes (plexogenic lesions) in more proximal pulmonary arteries associated with migration and proliferation of smooth muscle cells, and increased extracellular matrix deposition. There is also dysregulation of endothelial cells associated with increased proliferation.8 The mechanism underlying the evolution of these changes is unknown, so there was great interest when 2 groups independently identified a mutation in bone morphogenetic protein receptor 11 (BMP-RII) in 60% of families with IPH.9,10 A BMP-RII mutation also occurs in 20% of sporadic cases of IPH,11 but the biological connection between the mutation and the pathobiology of IPH has been relatively elusive. Recent studies using pulmonary artery smooth muscle cells from patients with IPH, including those with and without a BMP-RII mutation, showed similar abnormal proliferation in response to agents such as transforming growth factor-β (TGF-β) or BMP-2.12 In other studies, pulmonary artery smooth muscle cells were transfected with constructs encoding different mutant forms of BMP-RII expressing aberrant kinase or cytoplasmic domains, and impaired signaling was observed related to alterations in the induction of Smads and p38.13 Specifically, suppression of Smad1/5 and activation of p38 were related to smooth muscle cell proliferation. It …

Pulmonary hypertension is a pathophysiologic process characterized by progressive elevation of pulmonary vascular resistance and right heart failure, which is a common complication of many diseases. Pulmonary hypertension with no apparent causes (unknown etiology) is termed primary pulmonary hypertension or, more recently, idiopathic pulmonary arterial hypertension (IPAH). Before the availability of disease-specific (targeted) therapy (through the mid-1980s) the median life expectancy from the time of diagnosis in patients with this disease was 2.8 years.1-3 Modern treatment has markedly improved physical function and has extended survival, and the 5-year mortality is 50%.1 Although there is already more than 100 years of research history, the mechanisms of this disease are still not very clear.2 Recently, with the development of cell biology and molecular genetics, further research into the mechanisms responsible for pulmonary hypertension have been possible, which has helped in its diagnosis and treatment. It is believed that the mechanisms of pulmonary hypertension can not only be described by pathophysiology but involve multiple factors (pathways) like cellular, humoral and molecular genetics, etc. The increased contraction and remodeling of blood vessels and thrombosis are the major pathophysiological basis for the development of pulmonary hypertension.3 Endothelial cells, smooth muscle cells, fibroblasts and platelets all play important roles in the development of pulmonary hypertension. In addition, vasoconstrictive and vasodilating factors, proliferative stimulating and inhibiting factors, coagulating and anti-coagulating substances and different vasoactive substances are involved in its development. Currently, there is extensive work being done on the role of genetic factors in the development of pulmonary hypertension.4 CELLULAR MECHANISMS AND PULMONARY HYPERTENSION The pulmonary vascular remodeling is the major pathological basis of pulmonary hypertension. The structural changes in all three layers (inner, medial and external) of the pulmonary vascular wall have significant meaning in the incidence, development and recovery of the pulmonary hypertension. Endothelial cells An intact vascular endothelium, under normal physiological conditions, plays a very significant role in maintaining the phenotype of smooth muscle cells and the structure of blood vessels. Some abnormal conditions like hypoxia, mechanical injuries, inflammations, drugs and toxins, etc. affect the structure, function and metabolism of endothelial cells, which are the initial (primary) causes of pulmonary hypertension.5 The damage to the endothelium affects its barrier function and damages its link to smooth muscle cells. It also disturbs the equilibrium of the vasoactive substances produced by the endothelium and pulmonary circulation and their regulation of smooth muscle cells, which in turn promote the proliferation of smooth muscle cells and finally modulate the structure of pulmonary vessels. Endothelial damages not only causes disturbances in proliferation and apoptosis but also affect the coagulation of blood.6 In 90% of the lesions, endothelial cells do not express transforming growth factor (TGF)-β2 receptors, which suggests that tumor inhibiting genes are responsible in the development of IPAH.7 Smooth muscle cells During pulmonary artery hypertension, the static tunica media of smooth muscle cells are transformed to a proliferative synthetic state. The smooth muscle cells proliferate and enlarge resulting in hypertrophy of the tunica media. Moreover, the precursor cells of smooth muscle (like intermediate cells and pericytes), which do not differentiate under normal condition, differentiate into new smooth muscle cells. The partial muscular arteries and non muscular arteries get muscularisation which forms new muscular arteries.8 Our previous studies, as well as others, showed that the proliferative index (PI), the apoptotic indexes (AI) and the ratio of PI and AI of the pulmonary artery smooth muscle cells in rats with pulmonary hypertension were higher than those of normal rats. This indicates that hyperplasia and apoptosis of smooth muscle cells of the pulmonary artery occurs during the pulmonary vascular structural remodeling and the imbalance of hyperplasia and apoptosis also take part in the process. Moreover, caspases-3, bcl-2 and NF-κB genes take part in the regulatory mechanism for the smooth muscle cell apoptosis.9,10 Pulmonary smooth muscle cells also synthesize and secrete different vasoactive substances which regulate the pulmonary vascular structural remodeling and the pulmonary hypertension. Fibroblast The proliferation of the fibroblasts in the outer layer of blood vessels, and the abnormal deposition of connective tissues and the changes in extracellular matrix (ECM) are important components for pulmonary vascular structural remodeling. ECM includes collagen, elastin, etc. Our research team recently found that the expression of pulmonary artery collagen I, collagen III, procollagen I mRNA and procollagen III mRNA in rats with pulmonary hypertension were significantly elevated compared with the normal rats. The positive signals were mainly located in the media and adventitia of median and small pulmonary arteries. The expression of collagen degradation regulatory enzymes, tissue inhibitor of metalloproteinase-1 (TIMP-1) mRNA, metalloproteinase-1 (MMP-1) mRNA and the ratio of TIMP-1 and MMP-1 in the pulmonary artery were elevated in rats with pulmonary hypertension. These results suggested that with the development of pulmonary hypertension, collagen, as an important component of extracellular matrix, accumulated resulting from increased synthesis and decreased degradation of collagen.11 Our research also showed that elastin, another important ECM component, was also increased in hypoxic pulmonary hypertensive rats. Additionally, after 1 week of hypoxia the inner elastin lamina of the pulmonary artery of rats became thinned and the thickness of the inner elastin lamina was inversely proportional to the pulmonary artery pressure,12 which indicated that the changes in elastin also participated in the development of pulmonary hypertension. Platelets and thrombosis Functional disturbance of platelets and thrombosis play an important role in the development of IPAH. The damage to the endothelium of pulmonary blood vessels induces the activation and agglutination of platelets. The abnormality in the thrombomoduling system and fibinolysis system induce in situ thrombosis in the pulmonary artery.13 Platelets not only have anti-coagulant function but also produce constriction and remodulating active substances which cause remodulating of pulmonary blood vessels. Inflammatory cells The level of inflammatory cell factors like anti-nuclear antibody (ANA), interleukin (IL)-1 and IL-6 are found to be increased in some IPAH patients. Histology showed infiltration of macrophages and lymphocytes which indicated inflammatory cells might take part in the development of IPAH.14 Moreover, an inflammatory reaction plays a role, to some extent, in the development of pulmonary hypertension induced by connective tissue diseases and HIV infection. It is found that the immunosuppressive treatment is beneficial in some cases of lupus induced-pulmonary hypertension MOLECULAR MECHANISMS OF PULMONARY HYPERTENSION Endothelial cells, smooth muscle cells, fibroblasts, platelets and macrophages associated with blood vessels can produce different vasoactive substances. Under normal condition these substances exhibit a dynamic equilibrium; maintain the normal physiological structure and function of pulmonary vessels. The external stimuli (like high pulmonary blood flow, hypoxia and toxins) disturb the equilibrium and induce thrombosis, constriction and pulmonary vascular structural remodeling, which is an important mechanism in the development of pulmonary hypertension. Nitric oxide (NO) NO is closely related to the development of pulmonary hypertension. There exist different opinions regarding the role of NO in pulmonary hypertension. While not all conclusions are accordant. Most researchers believe that the expression of NO synthase (NOS) decreases during pulmonary hypertension resulting in a decrease in NO synthesis and, according to etiology and disease severity, the changes in NO levels also differ accordingly.15 Our research team, as well as others, found that long-term inhalation of NO or use of NO precursor L-arginine (L-Arg) or NO donor nitroglycerin alleviated pulmonary hypertension as well as pulmonary vascular structural remodeling in the hypoxic rats along with an increase in plasma NO. In contrast, NOS inhibitor significantly aggravated pulmonary vascular structural remodeling with downregulation of the endogenous NO/NOS pathway, which indicated NO played an important regulatory role in the development of pulmonary hypertension and pulmonary vascular structural remodeling.16,17 Recently, many hospitals reported the uses of NO inhalation, NO donors and precursor in the treatment of pulmonary hypertension and the effective treatment. Carbon monoxide (CO) Endogenous CO is mainly produced by the degradation of heme in the presence of heme oxygenase (HO). It relaxes the blood vessels and inhibits the proliferation of smooth muscle cells of blood vessels. Recent studies showed that there was the expression of HO in the smooth muscle cells and endothelial cells of pulmonary vessels, which indicated pulmonary circulation was one of the important places for production and release of endogenous CO. Our research team found that the CO/HO-1 system was increased in a time-dependent double-peak manner in hypoxic pulmonary hypertensive rats.18 For example, the content of CO in lung homogenates of rats from hypoxia day 1 and hypoxia day 3 groups was markedly increased compared with that of normal controls, while the content of CO in lung homogenates in the rats of hypoxia day 7 decreased to the baseline. Administration of zinc protoporphyrin (ZnPP), an inhibitor of HO-1, in the hypoxic rats decreased the content of CO in lung homogenate, decreased the apoptosis of the pulmonary artery smooth muscle cells and enhanced the proliferation of pulmonary artery smooth muscle cells and thus worsened hypoxic pulmonary hypertension and pulmonary vascular structural remodeling. Meanwhile, exogenous supply of CO had an adverse action.19 These results show that up-regulation of the CO/HO pathway plays an important role in the regulation of hypoxic pulmonary hypertension. Hydrogen sulfide (H2S) After the discovery of the role of NO and CO, H2S, known as toxic gas for a long time, is also described as a biologically active substance. It can be endogenously generated from cysteine in a reaction catalyzed by cystathionine β-synthesis (CBS) and cystathionine γ-lyase (CSE). The data from our research team, as well as others, demonstrated that H2S exhibited similar functions as NO and CO in the body, such as relaxation of blood vessels, inhibition of the proliferation of smooth muscle cells20 and promotion of the apoptosis of smooth muscle cells, etc. We also reported that H2S played a important role in pathophysiological processes of cardiovascular diseases, such as hypertension, pulmonary hypertension, shock and myocardial injury, etc. Our research team discovered that both the gene expression and activity of CSE were suppressed in lung tissues, and the plasma level of H2S was decreased under hypoxia. Furthermore, supplement of H2S donor molecules opposed the elevation of pulmonary arterial pressure and lessened the pulmonary vascular structure remodeling during hypoxic pulmonary hypertension, while DL-propargylglycine (PPG), a CSE inhibitor, aggravated hypoxic pulmonary hypertension. This indicates that the H2S/CSE pathway plays a significant role in the regulation of hypoxic pulmonary hypertension.21,22 NO, CO and H2S are three gaseous signaling molecules that have complicate interrelations in the regulation of pulmonary hypertension.23,24 Since 1987, NO, CO and more recently H2S, the endogenous gas molecules produced from metabolic pathway, have been recognized respectively as signal molecules involved in the regulation of homeostasis and to play important roles under physiological and pathophysiological conditions. The research into these endogenous gas signaling molecules opened a new avenue in life science. The biological regulatory effects of other endogenous gas molecules, which have previously been regarded as metabolic waste, are now a field of investigation in the life sciences and medicine. In recent years, we began to pay attention to the effects of endogenous sulfur dioxide (SO2) and its derivatives on mammalian and human physiology. SO2 can be produced endogenously from normal metabolism of sulfur containing amino acids. L-cysteine is oxidized via cysteine dioxygenase to L-cysteinesulfinate and the latter can proceed through transamination by glutamate oxaloacetate transaminase (GOT) to β-sulfinylpyruvate which decomposes spontaneously to pyruvate and SO2. During oxidative stress in mammals activated neutrophils can convert H2S to sulfite through a reduced form of a nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase-dependent process. Recently, our research team showed that SO2 could be endogenously generated in cardiovascular tissues and could exert important cardiovascular effects, such as vasorelaxant and negative inotropic effects. Moreover, SO2 might play a considerable role in the regulation of systemic circulatory pressure, pulmonary circulatory pressure and vascular structural remodeling in the pathogenesis of hypertension and hypoxic pulmonary hypertension. More studies of the significance of endogenous SO2 in the cardiovascular system under physiological and pathophysiological conditions need to be conducted.25-27 Vasoactive peptides and others vasoactive substances The arachidonic acid metabolic products include prostaglandin E1 (PGE1), PGE2, PGI2 and thromboxane. Among them, PGE2 and thromboxane constrict blood vessels whereas PGE1 and PGI2 dilate blood vessels. PGI2 has strong vasorelaxative effects. It inhibits proliferation of smooth muscle cells and agglutination of platelets. Disturbance in the metabolism of arachidonic acid and reduction in the expression of PGI2 synthesis enzymes occur in pulmonary hypertensive patients.28 Currently, PGI2 and similar functioning substances are successfully used in the treatment of pulmonary hypertension29 and in many countries it is recommended for treatment. In 1993, a new vasoactive polypeptide in chromaffin hemangioma called adrenomedullin (ADM) was found. It has vasorelaxative and hypotensive effects and also inhibits migration and proliferation of smooth muscles of blood vessels. There are different kinds of ADM receptors expressed in pulmonary tissues that combine with specific high affinity binding sites on ADM. The expression of ADM and its receptors increase in hypoxic hypertensive rat lung tissue. Serum ADM levels are also elevated. Our research team and others found that chronic infusion of ADM significantly decreased mean pulmonary artery pressure in hypoxic rats, lessened the muscularization of small pulmonary vessels, attenuated relative medial thickness and relative medial area of pulmonary arteries and alleviated the ultrastructural changes in pulmonary arteries of hypoxic rats. ADM inhibited the proliferation of pulmonary artery smooth muscle cells. Meanwhile, plasma proadrenomedullin N-terminal 20-peptide (PAMP) concentration and the expression of PAMP protein and mRNA in pulmonary arteries in rats with hypoxia treated with ADM were markedly decreased compared with the untreated hypoxic group. The results suggest that ADM ameliorates the development of hypoxic pulmonary vascular structural remodeling. Intramolecular regulation of ADM might play an important role in the regulation of hypoxic pulmonary hypertension by ADM.30,31 Recent studies showed that inhalation of ADM in IPAH patients reduced pulmonary artery pressure but did not change systemic arterial pressure and heart rate.32 This indicated that ADM could possibly become a new drug for the treatment of pulmonary hypertension. Endothelin-1 (ET-1) was discovered in 1988 and is a vasoconstrictive substance. ET stimulates and proliferation of cultured pulmonary artery smooth muscle cells in vitro. Its function is mediated by ETA and ETB receptors. Our research team and others found that ET precursors and ET-1, as well as the expressions of mRNA for ETA and ETB receptors, are significantly increased in the pulmonary arteries and lung tissues of pulmonary hypertensive rats.33 The ET receptor antagonist bosentan improves the hemodyanamics and functions in pulmonary hypertensive patients and is currently a recommended treatment for pulmonary hypertension in many countries.34 Angiotension (AT) is changed to AT II in the presence of angiotension trasferase, which is a strong vasoconstrictive substance. It also stimulates the proliferation of the smooth muscle cells of pulmonary vessels. It is already proven that angiotension trasferase antagonists, not only decrease pulmonary blood pressure, but also improve structural remodeling of pulmonary vessels.35 AT II mediates the high pulmonary blood flow-induced pulmonary vascular structural remodeling. 5-Hydroxytryptamine (serotonin, 5-HT) can constrict blood vessels and also stimulate the proliferation of smooth muscle cells. A significant raise of 5-HT is seen in pulmonary hypertensive patients. In 1999, Eddahibi and colleagues36 found that there was a significant raise in the expression of the 5-HT transporter in response to hypoxia in pulmonary smooth muscle cells. The deletion of the 5-HT transporter gene significantly protects from pulmonary hypertension in hypoxic rats. In IPAH patients, the 5-HT transporter shows polymorphism, which makes pulmonary artery smooth, cells sensitive to 5-HT's proliferation function.37 In addition, platelet-derived growth factor, vascular endothelial growth factor, epidermal growth factor, fibroblast growth factor, TGF, and the platelet-activating factor, urotensin, etc. may be involved in the development of pulmonary hypertension. Potassium channel The voltage dependent potassium channel (Kv, a subtype of potassium channel) is the major potassium channel which is responsible for the contraction of pulmonary artery smooth muscle cells. The inhibition of Kv results in a reduction in outflow of potassium from cells, which causes of depolarization of cell membrane and then the opening of calcium channels increase the calcium level in the cytoplasm, which resultes in vasoconstriction. The disturbance in the Kv channel plays an important role in the pathogenesis of IPAH. Selective Kv1.5 expression is reduced in IPAH patients, accompanied by Kv functional impairment, which leads to membrane depolarization and vasoconstriction.38 Dexfenfluraine plays a role in the treatment of pulmonary hypertension. This drug inhibits the activity of the Kv2.1 channel, which indicates potassium channel participates in the incidence of pulmonary hypertension.39 Potassium channels represent a new treatment target with potential therapeutic value for pulmonary hypertension patients. The regulation in the expression or activity of potassium channels can affect both pulmonary vascular tension and structure. GENETIC ISSUES IN PULMONARY HYPERTENSION Dresdale and colleagues40 discovered in 1954 that IPAH has genetic tendency. Later, a series of researches found that 6% of IPAH patients had a familial incidence and its clinical and pathological characteristics are similar to Sporadic IPAH patients. Based on the research of the families of IPAH patients, it was found that IPAH was an autosomal dominant inherited disease. But only 10%— 20% of the related mutation carriers show the features of pulmonary hypertension. The explication of features of pulmonary hypertension is more often seen in females than in males. Future generations of IPAH patients will gradually advance to a serious condition or there may be a skipped generation. In 1997, Morse et al41 found a susceptible gene located at 2q 31-33.41 On this basis, in 2000 Deng42 and Lane43 independently determined that the gene mutations in the bone morphogenetic protein receptor II (BMPR2) are one of the major causes in the development of IPAH. Currently, it is believed that a mutation in the BMPR2 gene causes a functional defect in BMPR2 receptors which is important in the development of IPAH. Germline mutations in the gene coding for the BMPR2 are present in more than 70% of familial pulmonary arterial hypentension (IPAH) and up to 26% of IPAH.44 Bone morphogenetic protein (BMP) belongs to the TGF-beta super family, and is synthesized and secreted by smooth muscle cells and endothelial cells. It mainly regulates those cells which exhibit major function in the embryonic development and stativity of tissues and inhibits proliferation of vascular smooth muscle cells and induces apoptosis. Similar to TGF-β, the regulatory signal transduction pathways of BMP involve a BMP type I receptor (BMPR1a and BMPR1b) and a BMP type II receptor (BMPR2). The type II receptor is the activator of the type I receptor, the two receptors form a complex unit. They activate Smad signaling protein and LIM-kinase to regulate gene transcription and maintain blood vessels.45 There are 46 types of BMPR2 gene mutations, 60% of which may lead to early termination of transcription. A point mutation and an anomaly in the structural domain of BMPR2 kinase may inhibit its receptor functions, rendering it unable to form a heterologous dimer complex or to lose kinase activity and block the signaling pathway. This can lead to excessive cell proliferation and apoptotic inhibition causing vascular structural remodeling and pulmonary hypertension. CONCLUSIONS Though the pathogenesis of most forms of pulmonary hypertension is unknown, there have been many recent developments, especially pertaining to the molecular genetics and cell biology of pulmonary hypertension. Since the range of medical conditions and environmental exposures associated with pulmonary hypertension is wide, it is difficult to envision a unifying pathogenic mechanism. Although there probably are genetic determinants, environmental exposures and acquired disorders that predispose patients to pulmonary hypertension, it is clear that none of the factors described in this review are sufficient alone to activate the pathways essential to the development of this vascular disease.

Platelet-derived growth factor is one of the major growth factors found in human and mammalian serum and tissues. Abnormal activation of platelet-derived growth factor signaling pathway through platelet-derived growth factor receptors may contribute to the development and progression of pulmonary vascular remodeling and obliterative vascular lesions in patients with pulmonary arterial hypertension. In this study, we examined the expression of platelet-derived growth factor receptor isoforms in pulmonary arterial smooth muscle and pulmonary arterial endothelial cells and investigated whether platelet-derived growth factor secreted from pulmonary arterial smooth muscle cell or pulmonary arterial endothelial cell promotes pulmonary arterial smooth muscle cell proliferation. Our results showed that the protein expression of platelet-derived growth factor receptor α and platelet-derived growth factor receptor β in pulmonary arterial smooth muscle cell was upregulated in patients with idiopathic pulmonary arterial hypertension compared to normal subjects. Platelet-derived growth factor activated platelet-derived growth factor receptor α and platelet-derived growth factor receptor β in pulmonary arterial smooth muscle cell, as determined by phosphorylation of platelet-derived growth factor receptor α and platelet-derived growth factor receptor β. The platelet-derived growth factor-mediated activation of platelet-derived growth factor receptor α/platelet-derived growth factor receptor β was enhanced in idiopathic pulmonary arterial hypertension-pulmonary arterial smooth muscle cell compared to normal cells. Expression level of platelet-derived growth factor-AA and platelet-derived growth factor-BB was greater in the conditioned media collected from idiopathic pulmonary arterial hypertension-pulmonary arterial endothelial cell than from normal pulmonary arterial endothelial cell. Furthermore, incubation of idiopathic pulmonary arterial hypertension-pulmonary arterial smooth muscle cell with conditioned culture media from normal pulmonary arterial endothelial cell induced more platelet-derived growth factor receptor α activation than in normal pulmonary arterial smooth muscle cell. Accordingly, the conditioned media from idiopathic pulmonary arterial hypertension-pulmonary arterial endothelial cell resulted in more pulmonary arterial smooth muscle cell proliferation than the media from normal pulmonary arterial endothelial cell. These data indicate that (a) the expression and activity of platelet-derived growth factor receptor are increased in idiopathic pulmonary arterial hypertension-pulmonary arterial smooth muscle cell compared to normal pulmonary arterial smooth muscle cell, and (b) pulmonary arterial endothelial cell from idiopathic pulmonary arterial hypertension patients secretes higher level of platelet-derived growth factor than pulmonary arterial endothelial cell from normal subjects. The enhanced secretion (and production) of platelet-derived growth factor from idiopathic pulmonary arterial hypertension-pulmonary arterial endothelial cell and upregulated platelet-derived growth factor receptor expression (and function) in idiopathic pulmonary arterial hypertension-pulmonary arterial smooth muscle cell may contribute to enhancing platelet-derived growth factor/platelet-derived growth factor receptor-associated pulmonary vascular remodeling in pulmonary arterial hypertension.

See related article, pages 1323–1330 Serotonin (5-HT, 5-hydroxytryptamine) has long been recognized as one of the most potent naturally occurring pulmonary vasoconstrictors.1 It was first implicated in the pathogenesis of pulmonary arterial hypertension (PAH) after an outbreak of the disease in Switzerland in the 1960’s among patients taking aminorex fumarate, an appetite suppressant that inhibits serotonin uptake by platelets.2 Since that time further outbreaks of PAH have been identified in Europe and the USA associated with the use of fenfluramine-derivate anorexigens,3–5 eventually leading to their withdrawal from the world market in 1997. Although this was, at least in retrospect, a predictable tragedy, it has ironically opened avenues of research into the biology of serotonin signaling in PAH. As fenfluramine-derivatives are substrates for the serotonin transporter (5-HTT, SERT) proteins,6 this suggests that abnormal SERT expression or functional activity could play a role in the pathogenesis of PAH. There is now a body of evidence supporting this hypothesis that provides hope for the development of effective therapeutic strategies targeting specific components of this signaling pathway in patients with these diseases. Most of the serotonin produced in the body is secreted by enterochromaffin cells of the intestine into the portal circulation where it is partially metabolized by the liver. However, levels of free circulating serotonin are maintained in the low nanomolar range through energy-dependent SERT-mediated transport into platelets. This led some researchers to hypothesize that fenfluramines might cause PAH by increasing free plasma levels of serotonin. However, this hypothesis is inconsistent with the observation that chronic treatment with fenfluramine-derivatives if anything reduces plasma levels of serotonin.6 This suggests that other SERT-related effects promote PAH in susceptible patients. This is supported by the observation that patients with idiopathic PAH have increased frequency of the so called L-type polymorphism …

PULMONARY ARTERIAL HYPERTENSION (PAH) is a progressive disease manifested by maladaptation of the pulmonary vasculature. The development of PAH can be influenced by genetic predisposition and/or by diverse endogenous or environmental stimuli. Regardless of the initial pathogenic factors, pulmonary vascular remodeling, sustained pulmonary vasoconstriction, in situ thrombosis, and increased pulmonary vascular wall stiffness are the major contributors to elevated pulmonary vascular resistance (PVR). The increase in PVR can lead to right ventricular failure and death in patients with PAH. While treatments for this disease are improving, it continues to be a life-threatening condition. MicroRNAs (miRNAs) have been implicated in the development and progression of PAH. MiRNAs are small, noncoding RNAs that regulate gene and protein expression by promoting degradation or suppressing translation of target mRNAs. Several studies have demonstrated aberrant expres

Excessive pulmonary artery (PA) smooth muscle cell (PASMC) proliferation and migration are implicated in the development of pathogenic pulmonary vascular remodeling characterized by concentric arterial wall thickening and arteriole muscularization in patients with pulmonary arterial hypertension (PAH). Pulmonary artery smooth muscle cell contractile-to-proliferative phenotypical transition is a process that promotes pulmonary vascular remodeling. A rise in cytosolic Ca2+ concentration [(Ca2+)cyt] in PASMCs is a trigger for pulmonary vasoconstriction and a stimulus for pulmonary vascular remodeling. Here, we report that the calcium homeostasis modulator (CALHM), a Ca2+ (and ATP) channel that is allosterically regulated by voltage and extracellular Ca2+, is upregulated during the PASMC contractile-to-proliferative phenotypical transition. Protein expression of CALHM1/2 in primary cultured PASMCs in media containing serum and growth factors (proliferative PASMC) was significantly greater than in freshly isolated PA (contractile PASMC) from the same rat. Upregulated CALHM1/2 in proliferative PASMCs were associated with an increased ratio of pAKT/AKT and pmTOR/mTOR and an increased expression of the cell proliferation marker PCNA, whereas serum starvation and rapamycin significantly downregulated CALHM1/2. Furthermore, CALHM1/2 were upregulated in freshly isolated PA from rats with monocrotaline (MCT)-induced PH and in primary cultured PASMC from patients with PAH in comparison to normal controls. Intraperitoneal injection of CGP 37157 (0.6 mg/kg, q8H), a non-selective blocker of CALHM channels, partially reversed established experimental PH. These data suggest that CALHM upregulation is involved in PASMC contractile-to-proliferative phenotypical transition. Ca2+ influx through upregulated CALHM1/2 may play an important role in the transition of sustained vasoconstriction to excessive vascular remodeling in PAH or precapillary PH. Calcium homeostasis modulator could potentially be a target to develop novel therapies for PAH.

Knowledge of molecular mechanisms underlying pulmonary arterial hypertension (PAH) continues to increase with the emerging theme that PAH is a heterogeneous disease involving multiple molecular abnormalities. Mutations in several genes have been identified in subsets of patients with PAH, and multiple signaling systems that influence vascular tone, function, and remodeling have been associated with PAH.1,2,3 In addition to mutations in BMPR2 , serotonin ( 5-HT ) and polymorphisms in its transporter ( SERT ) play a critical role in the pulmonary vascular smooth muscle hyperplasia and vascular remodeling found in PAH.4,5 Other genes and signals thought to contribute to the development of idiopathic pulmonary arterial hypertension (IPAH) include somatic mutations of BAX ,5 upregulation of Angiopoietin 1 ,6 transforming growth factor β1 polymorphisms,7 ALK1 mutation,8 SMAD8 mutation,9 and increased hyaluronic acid content associated with increased Hyaluronan Synthase 1 and decreased Hyaluronoglucosaminidase 1 gene expression.10 A recent observation also suggests that the noncanonical Wnt pathway is activated in IPAH.11 Article see p 2313 Several transient receptor potential canonical (TRPC) family …

Treatment of Pulmonary Arterial Hypertension: A Step Forward

miR-143 and miR-145 promote hypoxia-induced proliferation and migration of pulmonary arterial smooth muscle cells through regulating ABCA1 expression.

Pulmonary arterial hypertension (PAH) is a syndrome in which pulmonary arterial obstruction increases pulmonary vascular resistance, which leads to right ventricular (RV) failure and a 15% annual mortality rate. The present review highlights recent advances in the basic science of PAH. New concepts clarify the nature of PAH and provide molecular blueprints that explain how PAH is initiated and maintained. Five basic science concepts provide a framework to understand and treat PAH: (1) Endothelial dysfunction creates an imbalance that favors vasoconstriction, thrombosis, and mitogenesis. Restoration of this balance by inhibition of endothelin and thromboxane or augmentation of nitric oxide (NO) and prostacyclin is the paradigm on which most current therapy is based. (2) PAH has a genetic component. Mutations (bone morphogenetic protein receptor-2 [BMPR2]) and single-nucleotide polymorphisms (SNPs; ion channels and transporter genes) predispose to PAH. (3) Excess proliferation, impaired apoptosis, and glycolytic metabolism in pulmonary artery smooth muscle, fibroblasts, and endothelial cells suggest analogies to cancer. Many experimental therapies reduce PAH by decreasing the proliferation/apoptosis ratio; these include inhibitors of pyruvate dehydrogenase kinase (PDK), serotonin transporters (SERT), survivin, 3-hydroxy-3-methylglutaryl coenzyme A reductase, transcription factors (hypoxia-inducible factor [HIF]-1α and nuclear factor of activated T lymphocytes [NFAT]), and tyrosine kinases. Augmentation of voltage-gated K+ channels (Kv1.5) and BMPR2 signaling also addresses this imbalance. Tyrosine kinase inhibitors used to treat cancer are currently in phase 1 PAH trials. (4) Refractory vasoconstriction may occur due to rho kinase activation. Fewer than 20% of PAH patients respond to conventional vasodilators; however, refractory vasoconstriction may respond to rho kinase inhibitors. (5) The RV can be targeted therapeutically. Although increased afterload initiates RV failure, which is the major cause of death/dysfunction in PAH, the RV may be amenable to cardiac-targeted therapies. The RV in PAH has features of ischemic, hibernating myocardium. Guided by these new …

Pulmonary arterial hypertension (PAH) is a progressive disease characterized by elevated pulmonary vascular resistance (PVR) leading to right heart failure and premature death. The increased PVR results in part from pulmonary vascular remodeling and sustained pulmonary vasoconstriction. Excessive pulmonary vascular remodeling stems from increased pulmonary arterial smooth muscle cell (PASMC) proliferation and decreased PASMC apoptosis. A rise in cytosolic free Ca2+ concentration ([Ca2+]cyt) in PASMC is a major trigger for pulmonary vasoconstriction and a key stimulus for PASMC proliferation and migration, both contributing to the development of pulmonary vascular remodeling. PASMC from patients with idiopathic PAH (IPAH) have increased resting [Ca2+]cyt and enhanced Ca2+ influx. Enhanced Ca2+ entry into PASMC due to upregulation of membrane receptors and/or Ca2+ channels may contribute to PASMC contraction and proliferation and to pulmonary vasoconstriction and pulmonary vascular remodeling. We have shown that the extracellular Ca2+-sensing receptor (CaSR), which is a member of G protein-coupled receptor (GPCR) subfamily C, is upregulated, and the extracellular Ca2+-induced increase in [Ca2+]cyt is enhanced in PASMC from patients with IPAH in comparison to PASMC from normal subjects. Pharmacologically blockade of CaSR significantly attenuate the development and progression of experimental pulmonary hypertension in animals. Additionally, we have demonstrated that dihydropyridine Ca2+ channel blockers (e.g., nifedipine), which are used to treat PAH patients but are only effective in 15–20% of patients, activate CaSR resulting in an increase in [Ca2+]cyt in IPAH-PASMC, but not normal PASMC. Our data indicate that CaSR functionally couples with transient receptor potential canonical (TRPC) channels to mediate extracellular Ca2+-induced Ca2+ influx and increase in [Ca2+]cyt in IPAH-PASMC. Upregulated CaSR is necessary for the enhanced extracellular Ca2+-induced increase in [Ca2+]cyt and the augmented proliferation of PASMC in patients with IPAH. This review will highlight the pathogenic role of CaSR in the development and progression of PAH.

Pulmonary hypertension (PH) is a critical and dangerous disease in cardiovascular system. Pulmonary vascular remodeling is an important pathophysiological mechanism for the development of pulmonary arterial hypertension. Pulmonary artery smooth muscle cell (PASMC) proliferation, hypertrophy, and enhancing secretory activity are the main causes of pulmonary vascular remodeling. Previous studies have proven that various active substances and inflammatory factors, such as interleukin 6 (IL-6), IL-8, chemotactic factor for monocyte 1, etc., are involved in pulmonary vascular remodeling in PH. However, the underlying mechanisms of these active substances to promote the PASMC proliferation remain to be elucidated. In our study, we demonstrated that PASMC senescence, as a physiopathologic mechanism, played an essential role in hypoxia-induced PASMC proliferation. In the progression of PH, senescence PASMCs could contribute to PASMC proliferation via increasing the expression of paracrine IL-6 (senescence-associated secretory phenotype). In addition, we found that activated mTOR/S6K1 pathway can promote PASMC senescence and elevate hypoxia-induced PASMC proliferation. Further study revealed that the activation of mTOR/S6K1 pathway was responsible for senescence PASMCs inducing PASMC proliferation via paracrine IL-6. Targeted inhibition of PASMC senescence could effectively suppress PASMC proliferation and relieve pulmonary vascular remodeling in PH, indicating a potential for the exploration of novel anti-PH strategies.

Background and Objectives: Pulmonary Artery Hypertension (PAH) is considered as a malignant tumor in cardiovascular disease. Our previous study found that Calcium-Sensing Receptor (CaSR) is involved in pulmonary vascular remodeling in hypoxic pulmonary hypertension (HPH). However, the relationship of Pulmonary Artery Smooth Muscle Cell (PASMC) phenotypic switching, proliferation, and autophagy in CaSR-related HPH remain unclear. The purpose of this study was to detect the role of a CaSR antagonist, NPS2143, on the vascular remodeling by autophagy modulation under hypoxia. Methods: Hypoxic rat PAH model were simulated in vivo. Meanwhile, mean Pulmonary Artery Pressure (mPAP) was measured while RVI, WT%, and WA% indices were calculated. Immunohistochemistry and Western blot were used to detect phenotypic switching and cell proliferation in pulmonary arteriole. Cell viability was determined in vitro by CCK8 and cell cycle. Cell proliferation, phenotypic switching, autophagy level and PI3K/Akt/mTOR pathways were investigated in human PASMCs through mRNA or Western blot methods. Results: Rats with hypoxic-induced PAH had an increased mPAP, RVI, WT% and WA%. Moreover, expression of CaSR was significantly increased, followed by activation of autophagy (increased LC3b and decreased p62), phenotypic switching of PASMCs (reduced calponin, SMA-a and increased OPN) and pulmonary vascular remodeling. However, NPS2143 weakened these hypoxic effects. The results using hypoxic-induced human PASMCs confirmed that NPS2143 suppressed autophagy and reversed phenotypic switching in vitro by inhibiting PI3K/Akt/mTOR pathways. Conclusions: Our study demonstrates that NPS2143 was conducive to inhibit the proliferation and reverse phenotypic switching of PASMCs by regulating autophagy levels in HPH and vascular remodeling.

Activin-Like Kinase 5 (ALK5) Mediates Abnormal Proliferation of Vascular Smooth Muscle Cells from Patients with Familial Pulmonary Arterial Hypertension and Is Involved in the Progression of Experimental Pulmonary Arterial Hypertension Induced by Monocrotaline

Pulmonary hypertension is a devastating vascular disorder characterized by extensive pulmonary vascular remodeling, ultimately leading to right ventricular failure and death. Activation of PDGF (platelet-derived growth factor) signaling promotes the hyperproliferation of pulmonary arterial smooth muscle cells (PASMCs), thus contributing to the pulmonary vascular remodeling. However, the molecular mechanisms that govern hyperproliferation of PASMCs induced by PDGF remain largely unknown, including the contribution of long noncoding RNAs (lncRNAs). In this study, we aimed to identify a novel lncRNA regulated by PDGF implicated in PASMC proliferation in pulmonary vascular remodeling. RNA-sequencing analysis was conducted to identify a novel lncRNA named vessel-enriched lncRNA regulated by PDGF-BB (platelet-derived growth factor-BB; VELRP). Functional investigations of VELRP were performed using knockdown and overexpression strategies along with RNA sequencing. Validation of the function and potential mechanisms of VELRP was performed through Western blot, RNA immunoprecipitation, and chromatin immunoprecipitation assays. We identified a novel vessel-enriched lncRNA with an increased response to PDGF-BB stimulus. VELRP was identified as an evolutionarily conserved RNA molecule. Modulation of VELRP in PASMCs significantly altered cell proliferation. Mechanistically, VELRP enhances trimethylation of H3K4 (histone H3 lysine 4) by interacting with WDR5 (WD repeat-containing protein 5), leading to increased expression of CDK (cyclin-dependent kinase) 1, CDK2, and CDK4 and consequent hyperproliferation of PASMCs. The pathological relevance of VELRP upregulation in pulmonary artery was confirmed using rat pulmonary hypertension models in vivo, as well as in PASMCs from patients with idiopathic pulmonary arterial hypertension. Specific knockdown of VELRP in smooth muscle cells using adeno-associated virus type 9 SM22α (smooth muscle protein 22α) promoter-shRNA-mediated silencing of VELRP resulted in a significant decrease in right ventricular systolic pressure and vascular remodeling in rat pulmonary hypertension model. VELRP, as an lncRNA upregulated by PDGF-BB, mediates PASMC proliferation via WDR5/CDK signaling. In vivo studies demonstrate that targeted intervention of VELRP in smooth muscle cells can prevent the development of pulmonary hypertension.