Effect of Panax notoginoside on pulmonary arterial pressure and expression of p38 MAPK in lung tissues of hypoxic rats

AIM: To observe the effect of Panax notoginoside (PNS) on the pulmonary artery pressure and the p38 mitogen-activated protein kinase(p38 MAPK) in lung tissues of rats treated with hypoxia. METHODS: Thirty adult male SD rats were randomly divided into 3 groups. The rats in normal control group were exposed to normal conditions, the rats in hypoxia group were exposed to isobaric hypoxia, and the rats in hypoxia+PNS group were treated with PNS under the condition of hypoxia. After 4 weeks of treatment, the mean pulmonary arterial pressure (mPAP) and the mean carotid arterial pressure (mCAP) were measured by cardiac catheterization. The heart was isolated, and the right ventricle (RV), left ventricle plus ventricular septum (LV+S) were weighed to calculate the ratio of RV/(LV+S).The quantity of phospho-p38 MAPK(p-p38 MAPK) in rat pulmonary arterioles was determined by the method of immunohistochemistry and the mRNA content of p38 MAPK was tested by RT-PCR. RESULTS: The mPAP and RV/(LV+S) in hypoxia group were higher than those in normal control group. The expression of p-p38 MAPK in rat pulmonary arterioles and p38 MAPK mRNA in the lung tissues were higher than those in normal control group (P<0.05). The mPAP, RV/(LV+S), the expression of p-p38 MAPK in rat pulmonary arterioles and p38 MAPK mRNA in the lung tissues in hypoxia+PNS group were significantly lower than those in hypoxia group (P<0.05).CONCLUSION: PNS possesses the preventive and therapeutic effect on hypoxic pulmonary hypertension by decreasing p-p38 MAPK and down-regulation of p38 MAPK mRNA in the lungs.     

Studies on the role of ET-1 and CGRP in the pathophysiology of ankylosing spondylitis and rheumatoid arthritis

AIM: To clarify the role of endothelin-1 (ET-1) and calcitonin gene related peptide(CGRP) in the pathophysiology of ankylosing spondylitis and rheumatoid arthritis. METHOD: Plasma/synovial fluid ET-1 and CGRP were measured by radioimmunoassay in patients with ankylosing spondylitis (AS) or rheumatoid arthritis (RA) and healthy control. RESULTS: ET-1 level in plasma of patients with AS and RA were significantly higher than that of healthy controls (P<0.01). No difference was found in plasma CGRP level between AS or RA and healthy control (P>0.05). CGRP level in synovial fluid was significantly higher than that in plasma (P<0.01), but ET-1 level was significantly lower than in plasma (P<0.01). CONCLUSION: These results suggest that ET-1 and CGRP play a pathogenic role in AS and RA.     

Chronic hypoxia altered the expression of ET-1 mRNA in response to acute hypoxia in pulmonary artery endothelial cells

AIM:To investigate the expression of ET-1 mRNA in porcine pulmonary artery endothelial cells cultured in normoxic and chronic hypoxic conditions, and their different responses to acute hypoxia were also evaluated.METHODS:Insituhybridization and image -analysis system were used. RESULTS:Acute hypoxia enhanced the expression of ET-1 mRNA in both normoxic and chronic hypoxic group. The increment was more significant in the latter group.CONCLUSION:Chronic hypoxia increased the expression of ET-1 mRNA in response to acute hypoxia in porcine pulmonary artery endothelial cells.     

Effect of CGRP on level of lung endogenous NO in rabbits with acute lung injury

AIM:To examine whether calcitonin gene-related peptide (CGRP) enhances nitric oxide (NO) level in pulmonary circulation blood and observe the influence of CGRP on mean pulmonary artery pressure (mPAP) in rabbits with acute lung injury (ALI) caused by oleic acid.METHODS: The level of NO was assessed by measuring the presence of nitrite in cervical artery blood by the Griess reaction, mPAP was measured with right ventricular catheter.RESULTS:The level of nitrite in cervical artery blood was significantly increased and the mPAP was markedly reduced after administration of CGRP intravenousely.CONCLUSION:CGRP enhanced the NO level of pulmonary circulation blood and reduces the mPAP significantly in rabbits with ALI.     

Changes of plasma and myocardial calcitonin gene-related peptide in myocardial stunning rats

AIM: To investigate changes of calcitonin gene-related peptide(CGRP) in myocardial stunning rats. METHODS: Rat in vivo myocardial stunning model was used. CGRP content in plasma and myocardium were determined by radioimmunoassay. RESULTS: Plasma level of CGRP increased significantly (P＜0.01), but in left ventricular myocardium CGRP decreased obviously (P＜0.05) in myocardial stunning group compared with the control group. CONCLUSION: CGRP content in the left ventricular myocardium was negatively correlated with plasma CGRP.     

Effects of A2a adenosine receptor on hypoxic pulmonary hypertension in rats treated with salidroside

AIM: To study the protective effect of A_2a adenosine receptor (A_2aAR) on hypoxic pulmonary hypertension in the rats treated with salidroside. METHODS: Sprague-Dawley rats were randomly divided into 6 groups: normal control group, hypoxia group, hypoxia+salidroside (low dose) group, hypoxia+salidroside (median dose) group, hypoxia+salidroside (high dose) group, and hypoxia+CGS-21680 (a selective agonist of A_2aAR) group. Pulmonary hypertension in the rats was produced for 4 weeks. Mean pulmonary artery pressure (mPAP), mean carotid arterial pressure (mCAP) and the weight ratio of right ventricle/(left ventricle+septum)［RV/(LV+S)］ were measured. The expression of A_2aAR in the pulmonary arterioles was determined by immunohistochemistry and in situ hybridization. The mRNA expression of A2aAR in the lung tissues was detected by real-time RT-PCR. The protein level of A_2aAR in the lung tissues was analyzed by Western blotting. RESULTS: The mPAP in hypoxia group was significantly higher than that in normal control group. The mPAP in hypoxia+salidroside (high dose) group and CGS-21680 group was significantly lower than that in hypoxia group. RV/(LV+S) in hypoxia group were significantly higher than that in normal control group. RV/(LV+S) in hypoxia+salidroside (median dose) group, hypoxia+salidroside (high dose) group and CGS-21680 group were lower than that in hypoxia group. The ratio of vessel wall area/vessel total area (WA/TA) in hypoxia group was significantly higher than that in normal control group. WA/TA in hypoxia+salidroside (low dose) group, hypoxia+salidroside (median dose) group, hypoxia+salidroside (high dose) group and CGS21680 group were obviously lower than that in hypoxia group. The expression of A_2aAR was significantly higher in hypoxia group than that in normal control group. The expression of A_2aAR in hypoxia+salidroside (high dose) group and CGS-21680 group was obviously higher than that in hypoxia group. CONCLUSION: The A_2aAR attenuates pulmonary vessel remodeling and pulmonary hypertension induced by hypoxia. Salidroside protects the pulmonary vessel from remodeling and inhibits the development of hypoxia-induced pulmonary hypertension by up-regulation of A_2aAR expression.     

Effect of hydroxylamine on pulmonary arterial hypertension induced by chronic hypoxic hypercapnia in rats

AIM: To explore the effects of hydroxylamine on the pulmonary arterial pressure in chronic hypoxic hypercapnic rats. METHODS: Twenty-four male Sprague-Dawley rats were randomly divided into 3 groups (8 rats in each group): the normal control group (NC), hypoxic hypercapnia+normal saline group (NS), hypoxic hypercapnia+hydroxylamine group (HA). The animals in NS and HA groups were kept in the O₂ (9%-11%) and CO₂ (5%-6%) cabin, 8 h a day and 6 days a week for 4 weeks. Before entering the cabin, the rats in HA group were administered with 1 mL hydroxylamine (12.5 mg/kg) by intraperitoneal injection, while the rats in NS group were given intraperitoneal injection of 1 mL saline solution. The mean pulmonary arterial pressure (mPAP) was measured by external jugular vein cannulation. The heart was removed, and the right ventricle (RV) and the left ventricle plus the septum (LV+S) were dissected. The ratio of the wet weight of the RV to that of the LV+S was calculated. The changes of the pulmonary vascular construction were observed under optical microscope. The concentration of H₂S in the plasma was measured with a spectrometer. The expression of cystathionine-γ-lyase (CSE) in the pulmonary arterioles and bronchi was measured by immunohistochemistry and RT-PCR. RESULTS: The values of mPAP, RV/(LV+S),vessel wall area/total area (WA/TA) and media thickness of pulmonary arterioles (PAMT) in NS group and HA group were significantly higher than those in NC group (P<0.05). The level of H₂S in the plasma, the content of CSE protein and the expression of CSE mRNA in NC group were significantly lower than those in NS group (P<0.05). The values of mPAP, RV/(LV+S), WA/TA and PAMT in HA group were significantly lower than those in NS group (P<0.05). The level of H₂S in the plasma, the content of CSE protein and the expression of CSE mRNA in HA group were significantly higher than those in NS group (P<0.05). CONCLUSION: Hydroxylamine may decrease the pulmonary arterial hypertension induced by chronic hypoxic hypercapnia in rats by increasing the level of H₂S in the plasma, the content of CSE protein and the mRNA expression of CSE, thus improving the pulmonary vascular structural remodeling.     

Roles of ERK pathway and Panax notoginoside treatment in hypoxic hypercapnia pulmonary hypertension in rats

AIM: To investigate the roles of Panax notoginoside (PNS) and ERK1/2 signaling pathway in the pathological process of chronic hypoxic hypercapnia pulmonary hypertension in rats.METHODS: The animal model of chronic hypoxic hypercapnia pulmonary hypertension was set up in 72 male Sprague-Dawley rats and the animals were randomly divided into 6 groups: normal (N) group, hypoxic hypercapnia for 3-day (H_3d) group, hypoxic hypercapnia for 1-week (H_1w) group, hypoxic hypercapnia for 2-week (H_2w) group, hypoxic hypercapnia for 4-week (H_4w) group and PNS treatment (H_p) group.The rats in H_p group were injected with PNS (50 mg·kg^-1·d^-1, ip) before placing the animals into the hypoxic hypercapnia chamber.The rats in other groups were injected with normal saline (2 mL/kg, ip).The morphological changes of the pulmonary artery were observed under microscope with HE staining.Western blotting was used to detect the protein expression of p-ERK.The protein levels of p-ERK in the lung tissues and pulmonary blood vessels were determined by immunohistochemistry.RESULTS: The ratios of WA/TA in H_1w, H_2w, H_4w and H_p groups were higher than that in N group (P<0.05).The ratio of WA/TA in H_p group was obviously lower than that in H_4w group (P<0.05).The protein expression of p-ERK was barely positive in N group, but was up-regulated in the pulmonary tissues in all hypoxic rats.Compared with N group, the protein level of p-ERK was markedly up-regulated in H_3d group, reached its peak in H_2w group, and tended to decline in H_4w group (P<0.05).In pulmonary arterial tunica intima and tunica media, p-ERK protein was dramatically expressed in all hypoxic rats compared with the control animals (P<0.05).In the lung tissues, the protein level of p-ERK in H_p group was lower than that in H_4w group (P<0.05).In pulmonary arterial tunica intima and tunica media, the protein level of p-ERK in H_p group was lower by 84.86% than that in H_4w group (P<0.05).CONCLUSION: ERK1/2 as a signal transducer may play an important role in the development of hypoxia and hypercapnia induced pulmonary hypertension.PNS inhibits the expression of ERK1/2, thus attenuating the development of pulmonary hypertension and improving pulmonary vascular remodeling.     

Role of α1 adrenoceptor in proliferation of hypoxic pulmonary artery smooth muscle cells

AIM: To investigate the role of α₁ and β₂ adrenoceptors(α₁AR and β₂AR) in the proliferation of hypoxic pulmonary artery smooth muscle cells (PASMCs).METHODS: PASMCs were isolated by an explant method from neonatal bovine pulmonary arteries. The cultured PASMCs were exposed to 6.6% O₂ for 6 h, 12 h and 24 h. The method of -TdR incorporation was used to measure the proliferation of PASMCs. _i was assayed with Fura-2/AM. The mRNA expression of α₁AR, β₂AR, c-fos and c-myc was determined by Northern blotting. The effects of activation of α₁AR and β₂AR, and inhibition of α₁AR on the above indexes were observed by treating PASMCs with different AR agonists and antagonists under hypoxic condition.RESULTS: Significant increase in TdR incorporation in hypoxic PASMCs with α₁AR activation was observed, and marked decrease in that was induced by α₁AR inhibition. However, no significant change was found after β₂AR activation. _i , the mRNA expression of c-fos, c-myc, α₁AR and β₂AR in PASMCs were increased after hypoxia.CONCLUSION: Hypoxia induces the increase in _i and mRNA expression of c-fos and c-myc, leading to the proliferation of PASMCs. The hypoxic proliferation of PASMCs is intervened by α₁AR, but not β₂AR. The remodeling of pulmonary arteriole and pulmonary hypertension may be involved in the processes of pulmonary arteriole constriction and proliferation induced by hypoxia through up-regulation of α₁AR.     

The preventive effect of Panax notoginseng saponins (PNS) on chronic hypoxic pulmonary hypertension in rats

AIM: To investigate the preventive effect of PNS on chronic hypoxic pulmonary hypertension in rats. METHODS: Pulmonary arterial pressure observation, hematocrit (Hct)measurement, biochemical analysis and transmission electron microscopy were conducted to investigate the role of PNS. RESULTS: (1)Mean pulmonary arterial pressure (mPAP), right ventricular mean pressure (RVMP) and Hct were significantly higher in hypoxia group (H group) than that of control group (C group) and were much lower in hypoxia with PNS group (HT group) than that in H group; (2) Nitric oxide (NO₂^-/NO₃^-) concentration and nitric oxide synthase (NOS) activity in the plasma and the lung tissue, total superoxide dismutase (T-SOD) and copper/zinc-containing enzyme (Cu/ZnSOD) activities in the plasma were all significantly lower in H group than that in C group and were much higher in HT group than that in H group, but NO₂^-/NO₃^- concentration and NOS activity were still markedly decreased in comparison with C group; (3) Injury of endothelial cells in pulmonary arteriole was improved obviously in HT group Compared with H group. CONCLUSIONS: These results suggest that PNSreduces the increase in mPAP, probably through adjusting NOlevel, anti-damaging effectof free radicals, inhibiting the injury of endothelial cells and decreasing Hct.     

Effect of ginsenoside Rb1 on proliferation and serotonin transporter and 5-HT1B R expression in hypoxia-induced rat pulmonary artery smooth muscle cells via Rho/Rho-kinase pathway

AIM: To observe the effect of ginsenoside Rb1 on the proliferation and the expression of serotonin transporter (SERT), 5-hydroxytryptamine 1B receptor (5HT_1BR) in rat pulmonary artery smooth muscle cells (PASMCs) under hypoxia condition and the relationship with Rho/Rho-kinase signal pathway.METHODS: PASMCs were isolated from the adult male SD rats and primarily cultured. The subcultured cells from the 4th generation to the 6th generation were harvested and divided into normal group, and hypoxia group, different concentrations of Rb1 incubation groups treated with 50, 100 and 200 mg/L ginsenoside Rb1 under hypoxia (HR50, HR100 and HR200 groups, respectively). The viability of the PASMCs was measured by CCK-8 assay. BrdU positive cells were determined using flow cytometry. The expression of serotonin transporter and 5HT_1BR at mRNA and protein levels was detected by RT-PCR and Western blot, respectively. The PASMCs were randomly divided into normal group, hypoxia group, HR200 group and hypoxia+Y-27632 incubation group (HY group). The mRNA expression of Rho-kinase and phosphorylated myosin phosphatase target subunit 1 (p-MYPT1) protein level were investigated by RT-PCR and Western blot, respectively.RESULTS: Compared with normal group, the proliferation of PASMCs in hypoxia group was significantly increased (P<0.01). The cell viability and the expression of SERT and 5HT_1BR at mRNA and protein levels in all different concentrations of Rb1 groups were obviously decreased compared with hypoxia group (P<0.05). The mRNA expression of Rho-kinase and protein level of p-MYPT1 were markedly decreased in HR200 group, and no significant difference compared with HY group was observed (P<0.01).CONCLUSION: Treatment with ginsenoside Rb1 might prevent hypoxia-induced proliferation of PASMCs and over-expression of SERT and 5HT_1BR through inhibiting the Rho/Rho-kinase pathway.     

Role of K+ channels in hypoxic pulmonary vasoconstriction in normal and chronic hypoxic rats

AIM:To investigate the role of K⁺ channels in the decreased hypoxic pulmonary vasoconstriction(HPV) in chronic hypoxic rats. METHODS:Blockers of three kinds of K⁺ channels, 4-AP(voltage dependent K⁺ channel blocker), TEA(Ca²⁺ activated K⁺ channel blocker), GLIB(ATP sensitive K⁺ channel blocker) were used in isolated perfused rat lungs to detect the role of K⁺ channels in HPV. RESULTS:In normal rats, 4-AP and TEA, but not GLIB, both elicited a significant increase in pulmonary artery baseline pressure, and also potentiated the acute hypoxic pulmonary vasoconstriction. In chronic hypoxic rats, the HPV is significantly decreased, while 4-AP, TEA, GLIB all elicited a significant but smaller increase in pulmonary artery baseline pressure. Additionally, all these three blockers potentiated the HPV stronger in chronic hypoxic rats than in control rats. CONCLUSION:The opening of Kv, K_Ca, K_ATP might modulate the hypoxic pulmonary vasoconstriction in isolated rat lungs, and the increase in this modulation by potassium channel in chronic hypoxic rats might play a role in its decrease in HPV.     

Dynamic changes of neurokinin A and calcitonin gene-related peptide levels in gastric mucosal and plasma in rats with celiac seawater-immersing trauma

AIM: To observe the dynamic changes of neurokinin A (NKA) and calcitonin gene-related peptide (CGRP) levels in gastric mucosal and plasma in rats after abdominal seawater-immersing trauma, and to investigate the influence of these two sensory neuropeptides on acute gastric mucosal lesion. METHODS: Thirty-two SD rats were randomly divided into four groups (normal group, celiac seawater-immersing trauma 1, 2 and 3 h groups). With emzyoimmunoassay and radioimmunoassay respectively, gastric mucosal and plasma NKA and CGRP levels in rats were measured.RESULTS: Compared with normal rats, with the seawater-immersing time prolonged, gastric mucosal NKA and CGRP levels in rats were progressively decreased (P<0.05), while plasma NKA and CGRP levels significantly elevated. CONCLUSION: Seawater-immersion is a harmful factor, it can lead to elevated plasma NKA and CGRP levels and decrease gastric mucosal NKA and CGRP levels.     

p38 MAPK specific inhibitor SB203580 down-regulates acute pulmonary thromboembolism-induced pulmonary artery hypertension and monocyte chemoattractant protein-1 in rats

AIM：To explore the hypothesis that initiation of pulmonary hypertension involves the up-regulation of monocyte chemoattractant protein-1 (MCP-1) in acute pulmonary thromboembolism (PTE), and to evaluate the role of p38 mitogen-activated protein kinase (MAPK) in this process. METHODS：One hundred and fifty male SD rats were randomly divided into five groups (n=30): normal control group, solvent control group, acute PTE group, acute PTE plus SB203580 (a p38 MAPK specific inhibitor) pretreatment group and acute PTE plus C1142 (a rodent chimeric monoclonal antibody neutralizing rat MCP-1) pretreatment group. Thirty rats in each group were further divided into 1, 4 and 8 h subgroups (n=10). A rat model of acute PTE was established by infusion of an autologous blood clot into the pulmonary artery through a polyethylene catheter. SB203580 or C1142, dissolved in 1% dimethyl sulfoxide (DMSO), was administered to the animals through caudal vein 1 h prior to the beginning of acute PTE modeling. Rats in normal control group and solvent control group were injected with normal saline and 1% DMSO, respectively. The mean pulmonary artery pressure (MPAP) and the mRNA and protein expression of MCP-1 were measured at each time point. RESULTS：Acute PTE elicited significant increase in MPAP, and up-regulated the expression of MCP-1. Pretreatment with SB203580 or C1142 significantly reduced MPAP, and down-regulated the expression of MCP-1. CONCLUSION：These findings suggest that MCP-1 is involved in the formation of acute PTE-induced pulmonary hypertension, and SB203580 down-regulates the expression of MCP-1 via p38 MAPK signaling pathway, thus attenuating pulmonary hypertension.     

Effects of rat mesenchymal stem cells modified by CGRP on proliferation and phenotype transformation of vascular smooth muscle cells in vitro

AIM：To explore the effects of rat mesenchymal stem cells (MSCs) modified by calcitonin gene-related peptide (CGRP) on the proliferation and phenotype transformation of rat vascular smooth muscle cells (VSMCs) in vitro. METHODS：Rat MSCs and VSMCs were isolated, cultured and identified. The MSCs were infected by lentivirus which carried genes encoding enhanced green fluorescent protein (EGFP) and CGRP. The expression levels of CGRP in CGRP-modified MSCs were detected using real-time PCR and enzyme-linked immunosorbent assay (ELISA). The prolife-ration and migratory abilities of VSMCs were evaluated by MTT assay, Trypan blue staining and scratch test. The expression levels of α-smooth muscle actin (α-SMA) and osteopontin (OPN) were assessed by Western blotting. RESULTS：Compared with MSCs and MSCs-EGFP groups, the expression levels of CGRP in MSCs-CGRP group were markedly increased (P<0.01). The results of MTT assay and scratch test demonstrated that the proliferation and migratory abilities of VSMCs in MSCs-CGRP group were significantly inhibited compared with MSCs and MSCs-EGFP groups (P<0.05). Furthermore, cell viability was >90% shown by Typan blue staining. Western blotting showed that the expression of α-SMA was increased and that of OPN was decreased in MSCs-CGRP group compared with MSCs and MSCs-EGFP groups (P<005). CONCLUSION：CGRP-modified MSCs could secrete CGRP protein and inhibit the proliferation and migration of VSMCs, which may be associated with deterring the phenotype transformation from contractile type to synthetic type. These results lay a foundation for gene therapy in vivo.     

Effects of hypoxia-inducible factor-1 signal pathway on proliferation in hypoxic pulmonary artery smooth muscle cells

AIM: To investigate the effect of hypoxia on the proliferation and apoptosis of pulmonary artery smooth muscle cells (PASMC); and to evaluate the role of hypoxia-inducible factor-1α(HIF-1α), iNOS, P-ERK1/2 protein expression in hypoxic pulmonary hypertension (HPH) pathogenesis.METHODS: Cultured rat PASMC were divided into normoxic group; hypoxic group; hypoxia+ADM(adrenomedulin) group, hypoxia+L-NAME(iNOS inhibitor) group; hypoxia+PD98059 group. Proliferation was investigated by MTT and PCNA. Apoptosis was examined by flow-cytometry. Westen blotting was used to measure protein expression of HIF-1α, P-ERK1/2 and iNOS. RESULTS: (1) A value of 24 h-hypoxia was significantly higher than that in the normoxic group (P<0.01). In the hypoxia+PD98059 group, ADM was significantly lower than that in hypoxia group, whereas A value of the hypoxia+L-NAME was significantly higher than that in hypoxic group and normoxic group (P<0.01). (2) PCNA was positive in PASMC after 24 h hypoxia (P<0.01). PD98059, ADM inhibited the expression of PCNA significantly (P<0.01), whereas L-NAME increased the expression of PCNA significantly (P<0.01). (3) Apoptosis index was not significantly difference among the different groups (P>0.05). (4) HIF-1α, iNOS and P-ERK1/2 expression was poorly positive in normoxic group, positive after hypoxia for 4h (P<0.01), reaching its peak at 8 h hypoxia (P<0.01), HIF-1α, P-ERK1/2 expression declined after 24 h hypoxia. L-NAME promoted the expression of HIF-1α, PD98059 inhibited the expression of HIF-1α, iNOS and P-ERK1/2 partly. ADM inhibited the expression of HIF-1α partly, promoted the expression of iNOS. CONCLUSION: (1) Hypoxia stimulates the proliferation of PASMC, and has no obvious effects on the apoptosis of PASMC. (2) HIF-1 plays an importent role in the proliferation of hypoxic PASMC.     

Effects of EGLN1 siRNA on growth of rat pulmonary artery smooth muscle cells under hypoxic condition

AIM: To observe whether EGLN1 gene is involved in the growth of pulmonary arterial smooth muscle cells (PASMCs) during hypoxia when EGLN1 gene expression was interference by siRNA. METHODS: The rat primary pulmonary arterial smooth muscle cells were cultured, and the specific lipidosome of EGLN1 siRNA was constructed and transfected into the PASMCs. The transfected PASMCs were cultured under hypoxia or normoxia conditions, respectively. The viability of the PASMCs was detected by CCK-8 assay. The protein expression of EGLN1 and vascular endothelial growth factor (VEGF) was determined by Western blot. RESULTS: The viability of the PASMCs was increased and the protein expression of VEGF was up-regulated in the PASMCs under hypoxic condition in a time-dependent manner. In hypoxia or normoxia condition, the viability and VEGF protein expression of the PASMCs were suppressed by EGLN1 siRNA. CONCLUSION: EGLN1 gene may involve in the growth of rat PASMCs by regulating VEGF protein level under hypoxic condition.     

Upregulative effect of CGRP on expression of CREB and phospho-CREB protein during focal cerebral ischemia/reperfusion in rat parietal cortex

AIM: To investigate the effect of calcitonin gene related peptide (CGRP) on the expression of cyclic AMP response element binding protein (CREB) and phosphorylated-CREB in rat parietal cortex during focal cerebral ischemia/reperfusion (I/R).METHODS: Focal cerebral ischemia/reperfusion model was induced by occlusion of the right middle cerebral artery using the intraluminal suture method. The expressions of CREB and phospho-CREB in the parietal cortex in different groups (sham group, focal cerebral ischemia/reperfusion group and CGRP group) were detected with immunohistochemistry and Western blotting, and the positive products were analyzed by image analysis system.RESULTS: There was definite expression of CREB in right parietal cortex in sham group, while it was lesser in I/R group than that in sham group, but it became more in CGRP group than that in I/R group (P<0.05). Phospho-CREB was barely detected in right parietal cortex in sham group and it became more in I/R group than that in sham group. The expression of phospho-CREB increased in CGRP group than that in I/R group of the right parietal cortex (P<0.05).CONCLUSION: CGRP upregulates the expression of CREB and phospho-CREB in the ischemic neurons of the parietal cortex during focal cerebral ischemia/reperfusion and CREB probably involves in the mechanism of protective role of CGRP to ischemic neurons.     

Effects of 15-HETE and ERK1/2 on pulmonary artery constriction in chronic hypoxic rats

AIM:The aim of the present study was to investigate whether the extracellular signal regulated kinase-1/2 (ERK1/2) pathway was involved in 15-hydroxyeicosatetraenoic acid (15-HETE)-induced chronic hypoxic pulmonary artery (PA) constriction and whether ERK1/2 activity was influenced by 15-HETE, for clarifying the mechanism of hypoxic pulmonary vasoconstriction (HPV). METHODS:Rats were placed in hypoxic box with fractional inspired oxygen (FiO2) 0.12 for 9 days to make hypoxic models, while those lived in FiO₂ 0.21 served as normal controls. Heart and lungs were taken out from chest and PA in diameter of 1-1.5 mm was isolated and cut into rings with 3 mm long for tension studies in organ baths. The ring tensions before and after adding 15-HETE were compared. Influences of ERK1/2 upstream kinase inhibitor U0126 as well as endothelium integrity on 15-HETE-induced HPV were observed. Expression and activity of ERK1/2 in cultured rat pulmonary artery smooth muscle cells (PASMCs) treated with 15-HETE for different times and concentrations were examined by Western blotting. RESULTS:15-HETE significantly constricted PA rings from hypoxic rats, and the response of the hypoxic rings were significantly greater than that of normoxic ones (P<0.05). U0126 significantly reduced vasoconstriction induced by 15-HETE both in endothelium-intact and -denuded rings (both were P<0.05). Western blotting results showed 15-HETE enhanced activity of ERK1/2 in PASMCs, increasing with concentration and decreasing with time. CONCLUSION:15-HETE upregulates activity of ERK1/2 in PASMCs of rats. The activation of ERK1/2 is an important step in 15-HETE- induced HPV in rats.