TRPC6介导肺动脉高压大鼠肺动脉张力和肺动脉平滑肌细胞Ca~(2+)浓度的升高

经典瞬时感受器电位通道6(transient receptor potential channel6,TRPC6)蛋白是受体操纵性Ca2+通道(ROCC)的分子基础。本文旨在研究TRPC6/ROCC在野百合碱(monocrotaline,MCT)诱发的肺动脉高压大鼠模型中的作用。Sprague-Dawley大鼠随机分为正常对照组(CON组)和MCT组,CON组正常饲养三周,而MCT组按60mg/kg剂量一次性腹腔注射2%MCT,建立MCT诱导的慢性肺动脉高压大鼠模型。通过测定右心室收缩压(RVSP)和右心室重量指数(RVMI)、HE染色观察肺动脉血管形态,分析肺动脉结构重建。半定量RT-PCR和Western blot检测大鼠肺动脉TRPC6 mRNA和蛋白表达水平。血管张力实验中用可特异性激活ROCC、可透膜的DAG拟似物1-oleoyl-2-acetyl-sn-glycerol(OAG)检测大鼠离体肺动脉环的收缩效应。用荧光探针Fluo3-AM测定OAG诱导大鼠肺动脉平滑肌细胞(PASMCs)胞浆游离Ca2+浓度([Ca2+]i)。结果显示,与CON组相比,MCT组的RVSP、RVMI均明显增高(P0.01);形态学观察可见肺小动脉平滑肌层明显增厚,管腔减小;TRPC6的mRNA和蛋白质表达无明显变化。在CON组,OAG几乎不引起肺动脉环收缩,而在MCT组,肺动脉环的收缩反应显著增强,差别有显著性意义(P0.01)。相比较于CON组,MCT也可使OAG触发的PASMCs[Ca2+]i增量值显著升高(P0.05)。上述结果提示,MCT预处理对肺动脉TRPC6mRNA和蛋白质水平的表达无显著增强效应,但可促进TRPC6/ROCC介导的PASMCsCa2+内流和肺动脉张力升高,诱导大鼠产生肺动脉高压,并进一步诱发肺血管及右心室重构。     

肺纤维化初期肺动脉高压大鼠肺动脉反应性的变化

目的：观察肺纤维化初期肺动脉高压大鼠肺动脉血管反应性的变化。方法：66只雄性SD大鼠,随机分为博莱霉素（BLM）组和手术对照（Sham）组。BLM组为气管内一次性滴注BLM（5 mg/kg）;Sham组为气管内滴注等容量的生理盐水（NS）。应用离体血管张力检测技术测定大鼠肺动脉血管反应性变化;用HE显示肺动脉壁病理形态学变化;Masson染色检测肺纤维化程度;右心漂浮导管技术测定大鼠平均肺动脉压。结果：①BLM组大鼠的肺动脉血管（保留内皮和去内皮）对苯肾上腺素（PE）的收缩反应均弱于Sham组（P均〈0.05）。②BLM组大鼠肺动脉血管（保留内皮）对氯化乙酰胆碱（Ach）的舒张反应明显弱于Sham组（P〈0.01）。③Sham组有内皮的肺动脉血管对L-NAME和PE联合作用的收缩反应明显强于PE单独作用（P〈0.01）,而BLM组有内皮肺动脉血管对L-NAME和PE联合作用的收缩反应与对PE单独作用比,其差异无统计学意义（P〉0.05）。④BLM组肺动脉内皮细胞脱落。⑤BLM组大鼠肺组织呈现纤维增生初期的病理特征,且大鼠的平均肺动脉压明显高于Sham组（P〈0.05）。结论：肺纤维化形成初期肺动脉高压大鼠肺动脉血管反应性出现异常。     

NDUFA4L2 in smooth muscle promotes vascular remodeling in hypoxic pulmonary arterial hypertension

Pulmonary arterial hypertension (PAH) is characterized by a progressive increase in pulmonary vascular resistance and obliterative pulmonary vascular remodelling (PVR). The imbalance between the proliferation and apoptosis of pulmonary artery smooth muscle cells (PASMCs) is an important cause of PVR leading to PAH. Mitochondria play a key role in the production of hypoxia-induced pulmonary hypertension (HPH). However, there are still many issues worth studying in depth. In this study, we demonstrated that NADH dehydrogenase (ubiquinone) 1 alpha subcomplex 4 like 2 (NDUFA4L2) was a proliferation factor and increased in vivo and in vitro through various molecular biology experiments. HIF-1α was an upstream target of NDUFA4L2. The plasma levels of 4-hydroxynonene (4-HNE) were increased both in PAH patients and hypoxic PAH model rats. Knockdown of NDUFA4L2 decreased the levels of malondialdehyde (MDA) and 4-HNE in human PASMCs in hypoxia. Elevated MDA and 4-HNE levels might be associated with excessive ROS generation and increased expression of 5-lipoxygenase (5-LO) in hypoxia, but this effect was blocked by siNDUFA4L2. Further research found that p38-5-LO was a downstream signalling pathway of PASMCs proliferation induced by NDUFA4L2. Up-regulated NDUFA4L2 plays a critical role in the development of HPH, which mediates ROS production and proliferation of PASMCs, suggesting NDUFA4L2 as a potential new therapeutic target for PAH.     

姜黄素改善慢性肺动脉高压肺血管重塑作用途径的研究

目的：探究姜黄素改善慢性低O2高CO2大鼠肺动脉高压肺血管重塑作用途径的研究。方法：建立慢低O2高CO2肺血管重塑模型,以24只雄性大鼠为受试对象,随机分为四组（n=6）：I组（常氧空白对照组）,Ⅱ（低O2高CO2模型组）,Ⅲ组（色甘酸钠对照组）,Ⅳ组（姜黄素实验组）。将后3组动物放入常压低O2高CO2舱中,吸入O2浓度8％～11％,CO2浓度3％～5％,每天8h,每周6d,连续4周,Ⅲ组给予色甘酸钠以20mg／kg体重腹注射处理,Ⅳ组给予姜黄素混悬液按150mg/kg体重灌胃处理。光镜下、透射电镜下观察肺动脉血管壁及其周围大细胞超微结构形态学改变,甲苯胺蓝染色法和免疫组织化学法对肺动脉周围的肥大细胞及其脱颗粒状态进行性定量测定。结果：①电镜下,Ⅱ组肺细小动脉中膜平滑肌增生,外膜胶原纤维密集,内弹力板扭曲,内皮细胞起,血管外肥大细胞内颗粒减少,胞膜不完整;光镜下,Ⅱ组相比1组肺细小动脉管腔／管总面积（WA／TA）明显升（P〈0．05）、管腔／管总面积（EA／TA）明显降低（P〈0．05）,甲苯胺蓝染色肥大细胞细胞数（NMC）、肥大细胞脱颗率（DR）及免疫组化检测类胰蛋白酶阳性细胞数（TBS）高于I组（P〈0．05）;②干预后,电镜下,Ⅲ组、Ⅳ组血管结基本正常,平滑肌增生及胶原增生较Ⅱ组轻,肥大细胞膜完整;光镜下,两干预组相比Ⅱ组WA／TA明显降低（P〈0．05）、ET／TA明显升高（P〈0．05）,甲苯胺蓝染色NMC、DR、TBS阳性细胞数分别低于Ⅱ组（P〈0．05）。结论：姜素可通过Mc途径抑制慢性低O2高CO2导致的大鼠肺血管重塑改变。     

Bosentan in pulmonary arterial hypertension: a comparison between congenital heart disease and chronic pulmonary embolism

Background. In patients with pulmonary hypertension, it is unknown whether the treatment effect of bosentan is dependent on the duration of pulmonary vessel changes. Therefore, we studied the response to bosentan in patients with life-long pulmonary vessel changes (pulmonary arterial hypertension (PAH) due to congenital heart disease (CHD)) and in patients with subacutely induced pulmonary vessel changes (chronic thromboembolic pulmonary hypertension (CTEPH)). Methods. In this open-label study, 18 patients with PAH due to CHD and 16 patients with CTEPH were treated with bosentan for at least one year. All patients were evaluated at baseline and during follow-up by means of the six-minute walk distance (6-MWD) and laboratory tests. Results. Improvement of 6-MWD was comparable in patients with PAH due to CHD (444±112 m to 471±100 m, p=0.02), and in CTEPH (376±152 m to 423±141 m, p=0.03) after three months of treatment. After this improvement, 6-MWD stabilised in both groups. Conclusion. Although duration of pulmonary vessel changes is strikingly different in patients with PAH due to CHD and CTEPH, the effect of one year of bosentan treatment was comparable. The main treatment effect appears to be disease stabilisation and decreasing the rate of deterioration. (Neth Heart J 2009;17:334–8.)     

离子通道在肺动脉的分布及在低氧肺血管收缩反应中的作用

低氧性肺血管收缩反应（HPV）是指在急性低氧时,肺泡氧分压降到某一临界值,肺血管发生的快速、可逆的收缩反应,以纠正肺泡通气／灌流的不匹配。HPV的发生与肺动脉平滑肌细胞上K^＋、Ca^2＋、Cl^-通道的状态密切相关,而这些通道在不同部位的肺动脉上分布存在差异,因此不同部位的肺动脉在低氧中所表现的收缩反应程度也不同,本综述将对上述通道在肺动脉上的分布特点及其在HPV中的作用做一总结。     

Epistatic interactions in idiopathic pulmonary arterial hypertension

BACKGROUND:

Idiopathic pulmonary arterial hypertension (IPAH) is a poorly understood complex disorder, which results in progressive remodeling of the pulmonary artery that ultimately leads to right ventricular failure. A two-hit hypothesis has been implicated in pathogenesis of IPAH, according to which the vascular abnormalities characteristic of PAH are triggered by the accumulation of genetic and/or environmental insults in an already existing genetic background. The multifactor dimensionality reduction (MDR) analysis is a statistical method used to identify gene–gene interaction or epistasis and gene–environment interactions that are associated with a particular disease. The MDR method collapses high-dimensional genetic data into a single dimension, thus permitting interactions to be detected in relatively small sample sizes.

AIM:

To identify and characterize polymorphisms/genes that increases the susceptibility to IPAH using MDR analysis.

MATERIALS AND METHODS:

A total of 77 IPAH patients and 100 controls were genotyped for eight polymorphisms of five genes (5HTT, EDN1, NOS3, ALK-1, and PPAR-γ2). MDR method was adopted to determine gene–gene interactions that increase the risk of IPAH.

RESULTS:

With MDR method, the single-locus model of 5HTT (L/S) polymorphism and the combination of 5HTT(L/S), EDN1(K198N), and NOS3(G894T) polymorphisms in the three-locus model were attributed to be the best models for predicting susceptibility to IPAH, with a P value of 0.05.

CONCLUSION:

MDR method can be useful in understanding the role of epistatic and gene–environmental interactions in pathogenesis of IPAH.     

内毒诱导肺动脉内皮细胞生成过氧亚硝基阴离子的意义

目的：探讨牛肺动脉内皮细胞（BPAEC）产生过氧亚硝基阴离子（ONOO^-）的能力及其作用。方法：用流式细胞免疫荧光技术,定量检测内毒素主要成分脂多糖（LPS）诱导培养的BPAEC中ONOO^-生成标志物硝基酪氨酸（NT）的含量,观察ONOO^-对BPAEC形态变化的影响。结果：LPS可剂量依赖性诱导BPAEC产生ONOO^-明显增多,并为氮基胍部分翻转;ONOO^-可导致BPAEC明显回缩,胞体变小,细胞间隙增宽。结论：LPS诱导BPAEC产生增多的ONOO^-可能参与介导LPS对BPAEC本身的损伤效应。     

Chrysin ameliorates ANTU‐induced pulmonary edema and pulmonary arterial hypertension via modulation of VEGF and eNOs

Alpha‐naphthylthiourea (ANTU), a rodenticide induces lung toxicity. Chrysin a flavonoid possesses antioxidant, anti‐inflammatory, and antihypertensive potential. The aim of this study was to evaluate the efficacy of chrysin against ANTU‐induced pulmonary edema (PE) and pulmonary arterial hypertension (PAH) in laboratory rats. Sprague‐Dawley rats were used to induce PE (ANTU, 10 mg/kg, ip) and PAH (ANTU, 5 mg/kg, ip, 4 weeks). Animals were treated with chrysin (10, 20, and 40 mg/kg) and various biochemical, molecular, and histological parameters were evaluated. Acute administration of ANTU induces PE revealed by significant (P < 0.05) increase in relative lung weight, pleural effusion volume, lung edema, bronchoalveolar lavage fluid cell counts, total protein, 5‐hydroxytryptamine (5‐HT), lactate dehydrogenase (LDH), and γ‐glutamyl transferase (GGT), whereas pretreatment with chrysin (20 and 40 mg/kg, ip) significantly (P < 0.05) attenuated these ANTU‐induced biochemical and histological alterations. Repeated administration of ANTU caused induction of PAH evaluated by significant (P < 0.05) alterations in electrocardiographic, hemodynamic changes, and left ventricular function, whereas chrysin (20 and 40 mg/kg, p.o.) treatment significantly (P < 0.05) attenuated these alterations. ANTU‐induced hematological and serum biochemical (aspartate transaminase, alanine transaminase, LDH, and creatinine kinase MB) alterations were significantly (P < 0.05) inhibited by chrysin. It also significantly (P < 0.05) decreased elevated levels of oxido‐nitrosative stress in the right ventricle (RV) and lung. Chrysin significantly (P < 0.05) attenuated downregulated endothelial nitric oxide synthase and upregulated vascular endothelial growth factor messenger RNA and protein expressions both in the RV and pulmonary artery. Chrysin inhibited ANTU‐induced PE and PAH via modulation of inflammatory responses (5‐HT, LDH, and GGT), oxido‐nitrosative stress, and VEGF and eNOs levels.     

NAADP-induced intracellular calcium ion is mediated by the TPCs (two-pore channels) in hypoxia-induced pulmonary arterial hypertension

Pulmonary arterial hypertension (PAH) is a form of obstructive vascular disease. Chronic hypoxic exposure leads to excessive proliferation of pulmonary arterial smooth muscle cells and pulmonary arterial endothelial cells. This condition can potentially be aggravated by [Ca²⁺] _i mobilization. In the present study, hypoxia exposure of rat's model was established. Two-pore segment channels (TPCs) silencing was achieved in rats' models by injecting Lsh-TPC1 or Lsh-TPC2. The effects of TPC1/2 silencing on PAH were evaluated by H&E staining detecting pulmonary artery wall thickness and ELISA assay kit detecting NAADP concentrations in lung tissues. TPC1/2 silencing was achieved in PASMCs and PAECs, and cell proliferation was detected by MTT and BrdU incorporation assays. As the results shown, NAADP-activated [Ca²⁺]_i shows to be mediated via two-pore segment channels (TPCs) in PASMCs, with TPC1 being the dominant subtype. NAADP generation and TPC1/2 mRNA and protein levels were elevated in the hypoxia-induced rat PAH model; NAADP was positively correlated with TPC1 and TPC2 expression, respectively. In vivo, Lsh-TPC1 or Lsh-TPC2 infection significantly improved the mean pulmonary artery pressure and PAH morphology. In vitro, TPC1 silencing inhibited NAADP-AM-induced PASMC proliferation and [Ca²⁺]_i in PASMCs, whereas TPC2 silencing had minor effects during this process; TPC2 silencing attenuated NAADP-AM- induced [Ca²⁺]_i and ECM in endothelial cells, whereas TPC1 silencing barely ensued any physiological changes. In conclusion, TPC1/2 might provide a unifying mechanism within pulmonary arterial hypertension, which can potentially be regarded as a therapeutic target.     

脂肪干细胞移植对野百合碱诱发的肺动脉高压大鼠肺动脉压的量效和时效作用

目的探索脂肪干细胞（ADSC）移植治疗野百合碱（MCT）诱导的肺动脉高压（PAH）大鼠的适宜细胞数和干预时间。 方法（1）MCT的建模时效和量效：雄性SD大鼠48只分为正常对照组,20 mg/kg、30 mg/kg、40 mg/kg MCT组分别予腹腔注射生理盐水、MCT 20 mg/kg、30 mg/kg、40 mg/kg,4和8周后,右心室插管法检测平均肺动脉压（mPAP）,称重法计算右心室肥厚指数（RVHI）。（2）ADSC的治疗量效作用：雄性SD大鼠分别予腹腔注射MCT（30只）和生理盐水（30只）,1周后通过颈静脉注射分别移植0.5×10⁶、1.0×10⁶、3.0×10⁶、5.0×10⁶ADSC,其他组予等量生理盐水。移植3周后检测mPAP和RVHI。（3）ADSC的治疗时效作用：雄性SD大鼠30只,分别注射40 mg/kg MCT（24只）和生理盐水（6只）。MCT腹腔注射1 d,1、2周后分别移植1.0×10⁶个ADSC。MCT注射4周后检测mPAP和RVHI。多组间比较采用单因素或双因素方差分析,两两比较采用LSD检验。 结果（1）腹腔注射4周后,30 mg/ kg或40 mg/kg MCT组mPAP和RVHI均升高[mPAP值（24.89±3.31）mmHg,（27.19±2.11）mmHg比（15.80±0.42）mmHg,差异有统计学意义（P均< 0.05）;RVHI值0.42±0.06,0.47±0.04比0.25±0.02,差异有统计学意义（P均< 0.05）]。8周后,20 mg/kg或30 mg/ kg MCT组mPAP和RVHI均恢复正常,而40 mg/kg MCT组大鼠全部死亡。（2）40 mg/ kg MCT诱导的PAH大鼠mPAP和RVHI均升高。移植1.0×10⁶个ADSC可降低PAH大鼠的mPAP[（17.24±0.66）mmHg比（27.19±1.73）mmHg,P < 0.05]。移植0.5×10⁶、3.0×10⁶、5.0× 10⁶个ADSC不能降低PAH大鼠的mPAP和RVHI。（3）MCT腹腔注射1周和2周后,移植1.0×10⁶个ADSC可降低PAH大鼠的mPAP。 结论40 mg/kg MCT造模4周可建立稳定的PAH大鼠模型;造模1或2周后移植1.0×10⁶个ADSC能有效降低PAH大鼠的mPAP。     

低氧性肺动脉高压小鼠体内脂质水平的变化

目的：观察低氧性肺动脉高压小鼠体内脂质代谢的变化,探讨脂质代谢异常在低氧性肺动脉高压发生发展中的意义。方法：SPF级雄性C57BL/6小鼠20只,随机分为2组（n=10）：常氧组和低氧组。常压连续低氧3周（9%~11% O₂,23 h/d）复制慢性低氧性肺动脉高压模型,测定小鼠右心室压（RVSP）和右心室与左心室加室间隔重量比,Elisa法检测血浆中总胆固醇（TC）、低密度脂蛋白（LDL）、高密度脂蛋白（HDL）的含量;real-time PCR法检测肝组织中3-羟基-3-甲基戊二酸单酰辅酶A还原酶（HMGCR）、低密度脂蛋白受体（LDLR）、清道夫受体B1（SR-B1）、固醇调节元件结合因子2（SREBF2）等基因的表达。结果：低氧组小鼠RVSP、RV/（LV+S）显著高于常氧组（P<0.05）,血浆中HDL含量及HDL/LDL比值较常氧组显著降低（P<0.05）,肝组织中LDLR、SR-B1基因表达较常氧组显著下调（P<0.05）;RVSP与HDL/LDL比值及LDLR、SR-B1基因表达呈显著负相关（P<0.05）。结论：脂质代谢异常参与小鼠低氧性肺动脉高压的形成。     

Metabolomics reveals metabolite changes of patients with pulmonary arterial hypertension in China

The specific mechanism of pulmonary arterial hypertension (PAH) remains elusive. The present study aimed to explore the underlying mechanism of PAH through the identity of novel biomarkers for PAH using metabolomics approach. Serum samples from 40 patients with idiopathic PAH (IPAH), 20 patients with congenital heart disease‐associated PAH (CHD‐PAH) and 20 healthy controls were collected and analysed by ultra‐high‐performance liquid chromatography coupled with high‐resolution mass spectrometry (UPLC‐HRMS). Orthogonal partial least square‐discriminate analysis (OPLS‐DA) was applied to screen potential biomarkers. These results were validated in monocrotaline (MCT)‐induced PAH rat model. The OPLS‐DA model was successful in screening distinct metabolite signatures which distinguished IPAH and CHD‐PAH patients from healthy controls, respectively (26 and 15 metabolites). Unbiased analysis from OPLS‐DA identified 31 metabolites from PAH patients which were differentially regulated compared to the healthy controls. Our analysis showed dysregulation of the different metabolic pathways, including lipid metabolism, glucose metabolism, amino acid metabolism and phospholipid metabolism pathways in PAH patients compared to their healthy counterpart. Among these metabolites from dysregulated metabolic pathways, a panel of metabolites from lipid metabolism and fatty acid oxidation (lysophosphatidylcholine, phosphatidylcholine, perillic acid, palmitoleic acid, N‐acetylcholine‐d ‐sphingomyelin, oleic acid, palmitic acid and 2‐Octenoylcarnitine metabolites) were found to have a close association with PAH. The results from the analysis of both real‐time quantitative PCR and Western blot showed that expression of LDHA, CD36, FASN, PDK1 GLUT1 and CPT‐1 in right heart/lung were significantly up‐regulated in MCT group than the control group.     

Genetic counselling for pulmonary arterial hypertension: a matter of variable variability

We report three cases which highlight the complex considerations surrounding genetic counselling for pulmonary arterial hypertension (PAH). The first counselee developed PAH symptoms shortly after his daughter’s death from PAH and was diagnosed with a delay of 1 year. An early diagnosis of familial PAH was established in the second counselee. Oral therapy was initiated immediately, and her functional status has since remained stable. The third counselee was a healthy woman who struggled to cope with her risk for familial PAH, having lost two siblings from the disease. These cases show that incomplete penetrance and variable expression need particular attention during clinical assessment and genetic counselling of heritable PAH patients and family members.     

低氧对培养的不同内径的肺动脉平滑肌细胞增殖的影响

目的和方法：分离培养三种不同内径的肺动脉平滑肌细胞（PASMCs）,用^3H－TdR掺入速率和细胞计数作为细胞增殖的指标,观察低氧对其增殖作用的影响。结果：低氧对三种不同内径的PASMCs（内径分别为＞1000μm、500－800μm、300-400μm）增殖促进作用显著不同,其^3H－TdR掺入速率和细胞计数分别增加23.5％和11.1％、60.0％和33.8％、141.4％和52.0％,选择对低氧最敏感的PASMCs（内径为300－400μm）,进一步探讨低氧促PASMCs增殖作用的细胞机制：钙拮抗剂verapail、蛋白激酶C抑制剂staurosporine(Stau)和细胞Na-H交换抑制剂amiloride可显著降低低氧情况下PASMCs^3H－TdR掺入速率和细胞计数。结论：低氧对三种不同内径的PASMCs增殖促进作用显著不同; Ca^2 、蛋白激酶C和Na^2 －H^ 交换的激活,可能是低氧促PASMCs增殖的重要胞内信息转导机制。     

Identification of immune-associated signatures and potential therapeutic targets for pulmonary arterial hypertension

Pulmonary arterial hypertension (PAH) comprises a heterogeneous group of diseases with diverse aetiologies. It is characterized by increased pulmonary arterial pressure and right ventricular (RV) failure without specific drugs for treatment. Emerging evidence suggests that inflammation and autoimmune disorders are common features across all PAH phenotypes. This provides a novel idea to explore the characteristics of immunological disorders in PAH and identify immune-related genes or biomarkers for specific anti-remodelling regimens. In this study, we integrated three gene expression profiles and performed Gene Ontology (GO) and KEGG pathway analysis. CIBERSORT was utilized to estimate the abundance of tissue-infiltrating immune cells in PAH. The PPI network and machine learning were constructed to identify immune-related hub genes and then evaluate the relationship between hub genes and differential immune cells using ImmucellAI. Additionally, we implemented molecular docking to screen potential small-molecule compounds based on the obtained genes. Our findings demonstrated the density and distribution of infiltrating CD4 T cells in PAH and identified four immune-related genes (ROCK2, ATHL1, HSP90AA1 and ACTR2) as potential targets. We also listed 20 promising molecules, including TDI01953, pemetrexed acid and radotinib, for PAH treatment. These results provide a promising avenue for further research into immunological disorders in PAH and potential novel therapeutic targets.     

t-plasminogen activator inhibitor-1 polymorphism in idiopathic pulmonary arterial hypertension

AIM:

The aim of the present study was to identify the possible genotypic association of 3’UTR Hind III polymorphism of Plasminogen activator Inhibitor-1 (PAI-1) gene with idiopathic pulmonary arterial hypertension (IPAH).

BACKGROUND:

IPAH is a disorder with abnormally raised mean pulmonary arterial pressure and increase in the resistance to blood flow in pulmonary artery. One of the pathological features seen is development of intraluminal thrombin deposition leading to thrombosis. Plasminogen activator inhibitor-1 is an important inhibitor of the fibrinolytic system; its up-regulation may suppress fibrinolysis and result in an increased risk of thrombosis.

METHOD:

Blood samples from 54 IPAH patients and 100 healthy voluntary donors were analyzed by PCR-RFLP method for 3’UTR Hind III polymorphism.

RESULTS AND DISSCUSSION:

A significant association of Hd2 allele with the disease was observed. Raised mean level of right ventricular systolic pressure was observed in the Hd2/Hd2 genotypic patients, strengthening the role of Hd2 allele in the disease progression. Our data suggests an association of Hd2/Hd2 genotype, which may lead to the up-regulation of PAI-1 gene leading to increased levels of PAI-1, which is seen in IPAH. PAI-1 competes with plasminogen activators and hinders the normal mechanism of plasminogen activation system and leads to thrombosis and formation of plexiform lesions in the lung tissue, further strengthening its role in tissue remodeling and disease progression.     

Astragaloside IV improves pulmonary arterial hypertension by increasing the expression of CCN1 and activating the ERK1/2 pathway

The aim of the present study was to investigate the underlying mechanism of AS-IV and CCN1 in PAH and to evaluate whether the protective effect of AS-IV against PAH is associated with CCN1 and its related signalling pathway. In vivo, male SD rats were intraperitoneally injected with monocrotaline (MCT, 60 mg/kg) or exposed to hypoxia (10% oxygen) and gavaged with AS-IV (20, 40 and 80 mg/kg/day) to create a PAH model. In vitro, human pulmonary artery endothelial cells (hPAECs) were exposed to hypoxia (3% oxygen) or monocrotaline pyrrole (MCTP, 60 μg/mL) and treated with AS-IV (10, 20 and 40 μM), EGF (10 nM, ERK agonist), small interfering CCN1 (CCN1 siRNA) and recombinant CCN1 protein (rCCN1, 100 ng/mL). We identified the differences in the expression of genes in the lung tissues of PAH rats by proteomics. At the same time, we dynamically detected the expression of CCN1 by Western blot both in vivo and in vitro. The Western blot experimental results showed that the expression of CCN1 increased in the early stage of PAH and decreased in the advanced stage of PAH. The results showed that compared with the control group, MCT- and hypoxia-induced increased the hemodynamic parameters and apoptosis. AS-IV can improve PAH, as characterized by decreased hemodynamic parameters, vascular wall area ratio (WA%), vascular wall thickness ratio (WT%) and α-SMA expression and inhibition of cell apoptosis. Moreover, the improvement of PAH by AS-IV was accompanied by increased CCN1 expression, which activated the ERK1/2 signalling pathway. Meanwhile, CCN1 and p-ERK1/2 were inhibited by siCCN1 and promoted by rCCN1. EGF not only activated the ERK1/2 signalling pathway but also induced the expression of CCN1. In conclusion, AS-IV improves PAH by increasing the expression of CCN1 and activating the ERK1/2 signalling pathway. The results of our study provide a theoretical basis for additional study on the protective effect of AS-IV against PAH.     

Identification of plasma protein biomarkers associated with idiopathic pulmonary arterial hypertension

We have employed SELDI-TOF MS to screen for differentially expressed proteins in plasma samples from 27 patients with idiopathic pulmonary arterial hypertension (IPAH) and 26 healthy controls. One ion (m/z approximately 8600) that was found to be elevated in IPAH was validated by SELDI-TOF MS analysis of a second and separate set of plasma samples comprising 30 IPAH patients and 19 controls. The m/z 8600 was purified from plasma by sequential ion exchange and reverse-phase chromatographies and SDS-PAGE. It was identified, following trypsin digestion, by MS peptide analysis as the complement component, complement 4a (C4a) des Arg. Plasma levels of C4a des Arg measured by ELISA confirmed that the levels were significantly higher (p < 0.0001) in IPAH patients (2.12 +/- 0.27 microg/mL) compared with normal controls (0.53 +/- 0.05 microg/mL). A cut-off level of 0.6 microg/mL correctly classified 92% of IPAH patients and 80% of controls. Further studies will be needed to determine its performance as a diagnostic biomarker, whether used alone or in combination with other biomarkers. Nevertheless, this study demonstrates that putative biomarkers characteristic of IPAH can be identified using a conjoint SELDI-TOF MS - proteomics approach.