Baicalein inhibits monocrotaline-induced vascular wall thickening in rats with pulmonary hypertension

AIM:To investigate the effects of baicalein on pulmonary arterial hypertension (PAH) induced by monocrotaline (MCT) in rats, and its molecular mechanism was further explored. METHODS:Male SD rats (n=28) were randomly divided into 4 groups:control group, MCT group, MCT+baicalein 50 mg/kg group and MCT+baicalein 100 mg/kg group. The PAH model was established by subcutaneous injection of MCT. After 2 weeks of modeling, the rats in baicalein treatment groups were gavaged baicalein 50 and 100 mg·kg^-1·d^-1 for 14 d, the rats in control group were administered with saline. After 4 weeks of modeling, right ventricular systolic pressure (RVSP), right ventricular hypertrophy index (RVHI) and right ventricular mass index (RVMI) were detected. Masson staining was used to detect the degree of lung fibrosis. The pathomorphological changes of the pulmonary vessels were observed by HE staining. Western blot was used to detect the expression of α-smooth muscle actin (α-SMA) in the lung tissue and the phosphorylation p38, ERK and JNK in the artery. RESULTS:Compared with the control group, RVSP, RVHI and RVMI increased significantly in the MCT group (P<0.01). Pulmonary fibrosis and the thickening of pulmonary artery wall were observed. α-SMA was up-regulated and p38, ERK and JNK was activated significantly (P<0.01). Compared with the MCT group, baicalein (50 and 100 mg/kg) significantly decreased the RVSP, RVHI and RVMI (P<0.01). Lung fibrosis was reduced and the vascular wall thickening was decreased in baicalein-treated groups. Baicalein (50 and 100 mg/kg) inhibited the phosphorylation of p38, ERK and JNK compared with the MCT group (P<0.01). CONCLUSION:Baicalein ameliorates MCT-induced PAH by the inhibition of pulmonary artery wall thickening at least partially via MAPK signaling pathway.     

Effect of HGF gene transfection on human lymphoma xenografts in nude mice

AIM：To explore the promotion effect of hepatocyte growth factor (HGF) gene transfection on human lymphoma xenografts in nude mice. METHODS：The model of human lymphoma xenograft in nude mice was established by transplantation of Raji cells, which were transfected with recombinant plasmid pVITRO2-HGF harboring the HGF gene. The body weight of the nude mice and the tumor size were dynamically monitored and the tumor tissues were obtained after 8 weeks. Additionally, the methods of terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) and immunohistochemistry were used to detect the apoptotic index (AI) and microvessel density (MVD). RESULTS：The success rate of the human lymphoma xenografts in nude mice was 96.7%. The tumor volume in HGF transfection group was significantly greater than that in HGF transfection+VP-16 group and control groups (non-transfection group and empty vector group). The tumor volume in HGF transfection+VP-16 group was also bigger than that in control groups. No difference of the tumor volume between non-transfection group and empty vector group was observed. AI in HGF transfection group was substantially lower than that in control groups. AI in HGF transfection+VP-16 group showed a little higher than that in HGF transfection group, yet was still lower than that in control groups. MVD in HGF transfection group was extraordinary higher than that in control groups, but decreased after VP-16 induction (P<0.01), which was still higher than that in control groups. CONCLUSION：HGF gene transfection significantly promotes the growth of human lymphoma xenografts in nude mice and substantially inhibits the apoptosis presumably owing to promoting tumor angiogenesis and inhibiting tumor cell apoptosis.     

Alteration of pulmonary vascular structure and gaseous molecules in rats with pulmonary hypertension induced by high pulmonary blood flow

AIM: To examine the alteration of pathologic structure and gaseous molecules in rats with pulmonary hypertension induced by high pulmonary blood flow.METHODS: Aortocaval shunting was produced for 11 weeks in rats, and pulmonary hemodynamics was evaluated.Pulmonary vascular micro- and ultra- structure was also examined.Meanwhile,the concentration of plasma nitric oxide (NO) and carbon monoxide (CO) was measured by spectrophotometry.The expression of endothelial nitric oxide synthase (eNOS) and heme oxygenase-1 (HO-1) in pulmonary arteries was detected by immunohistochemistry.RESULTS: After 11- week aortocaval shunting,pulmonary artery mean pressure was significantly increased.Muscularization of small pulmonary vessels and relative medial thickness and area of pulmonary arteries were obviously increased in shunting rats compared with controls.Ultrastructure of intrapulmonary arteries changed obviously in shunting rats.Meanwhile,plasma NO concentration was increased and eNOS expression in pulmonary artery endothelial cells was significantly augmented in rats of shunting group.Plasma carbon monoxide level and HO-1 expression in puomonary artery smooth muscle cells,however,were not altered in shunting rats.CONCLUSIONS: Pulmonary vascular structural remodeling is the important pathologic basis of pulmonary hypertension induced by a left-to-right shunt,and NO other than CO might play an important regulating role in the development of high pulmonary blood flow-induced pulmonary hypertension.     

Effects of adipose tissue-derived mesenchymal stem cells on calcium channels of pulmonary artery in rats with pulmonary arterial hypertension induced by monocrotaline

AIM: To investigate the effects of adipose tissue-derived mesenchymal stem cells (ADMSCs) on calcium channels of pulmonary artery in monocrotaline (MCT)-induced pulmonary hypertensive rats.METHODS: ADMSCs were isolated from adipose tissue by collagenase digestion. Twenty-four Sprague-Dawley rats were randomly divided into 3 groups: normal control (Ctr) group, pulmonary arterial hypertension (PAH) group and ADMSCs transplantation group. Mean pulmonary arterial pressure (MPAP) was measured by catheterization, and right ventricular hypertrophy index (RVHI) was calculated. The expression of voltage-gated calcium channel α1c subunit (CaVα1c), sarcoplasmic/endoplasmic reticulum calcium ATPase 2a (SERCA-2a), inositol 1,4,5-triphosphate receptor 1(IP3R-1), transient receptor potential channel 1 (TRPC1) and TRPC6 at mRNA and protein levels in the pulmonary trunks was determined by RT-PCR and Western blotting, respectively.RESULTS: MPAP and RVHI were higher in PAH group than those in Ctr group, while those in ADMSCs group were significantly decreased as compared with PAH group. The expression of CaVα1c, TRPC1 and TRPC6 at mRNA and protein levels was obviously increased in PAH group as compared with Ctr group, while that in ADMSCs group was significantly decreased as compared with PAH group. Compared with Ctr group, the expression of SERCA-2a and IP3R-1 at mRNA and protein levels was obviously decreased in PAH group, while that in ADMSCs group was significantly increased as compared with PAH group.CONCLUSION: MPAP and RVHI are attenuated by ADMSCs in MCT-induced pulmonary hypertensive rats. The reduction of pulmonary arterial pressure by ADMSCs transplantation in MCT-induced pulmonary hypertensive rats may be related to the changes of calcium channels.     

Inhibitory effects of local transfection of vascular endothelial growth factor 165 gene on intimal hyperplasia of artery in rabbits after operation injury

AIM: To observe the effects of local transfection of vascular endothelial growth factor 165 (VEGF165) gene on inhibiting intimal hyperplasia and restenosis of artery in rabbits after operation injury, and its possible mechanisms. METHODS: Microsurgery injury was used to establish the intimal injury model of right external iliac artery in rabbits. 105 male New Zealand rabbits were randomly divided into 3 groups (35 rabbits in each group). Group A was physiological saline control group, group B was pBudCE4.1-transfected group, group C was pBudCE4.1/VEGF165-transfected group. The physiological saline, pBudCE4.1 and pBudCE4.1/VEGF165 transfection solutions were injected into injured vessel walls of above-mentioned groups. The injured vascular specimen was harvested for pathologic examination, electric microscope observation, RT-PCR examining and immunohistochemical staining. RESULTS: Rabbit intimal thickness and area of vessel walls in group C at every time point after operation were significantly less than those in group A and group B (P<0.01). The stenosis ratio of vessels in group C at 28 days after operation decreased by 51.6% and 49.8%, respectively, as compared with groups A and B. The expression of VEGF165 mRNA and VEGF165 positive cells in Group C were increased significantly than those in group A and B at every time point after operation (P<0.01). CONCLUSION: Local transfection of VEGF165 gene restrains intimal hyperplasia and restenosis of vessels, which lays a foundation for future gene therapy of vascular intimal hyperplasia.     

Changes of lung tissue and structure on bone marrow mesenchymal stem cells transplantation to treat the experimental pulmonary arterial hypertension in rats

AIM: To investigate the changes of lung tissue and structure on bone marrow mesenchymal stem cells (MMSCs) transplantation to treat the pulmonary arterial hypertension (PAH) in rats.METHODS: MMSCs cells were collected from bone marrow of SD rat’s femoral’s tibial bones, mix cultured with Hoechst 33342 fluorescence dye in vitro. Sixty healthy male Sprague-Dawley rats (weighing 180 g±5 g) were randomly divided into 3 groups: normal control group (group C), MMSCs transplanted group (group M), pulmonary model group (group H). The rats of the two latter groups were given a single subcutaneous monocrotaline (50 mg/kg) to induce the model of PAH. The rats of normal control group were injected respectively a single subcutaneous isotonic Na chloride (6 mL/kg). The three groups were fed in the same condition. After 3 weeks, 5×109 cells/L MMSCs with l mL phosphate-buffered saline were infused into the rats respectively in group M by sublingual vein and 1 mL L-DMEM was administered in group H. Nothing was given to group C. The viability, right ventricle systolic pressure (RVSP), right ventricle hypertrophy index, the microstructure and ultrastructural changes of small pulmonary vascular were observed after 28 d.RESULTS: The viability of administrated MMSCs 28 d after PAH was 100% in transplanted group, and it nearly completely prevented the increase in right ventricular systolic pressure (RVSP) compared to PAH alone ［(32.20±2.32)mmHg vs (48.30±1.56)mmHg, respectively, P<0.05］, right ventricle hypertrophy index also degraded (38.80%±3.24% vs 45.10%±3.43%, respectively, P<0.05). The pulmonary arteries remodeling and ultrastructural changes of lung, such as blood gas barrier, chondriosome, osmiophilic lamellar body and so on, were improved significantly in MMSCs transplanted group. The changes were similar to the results of normal control group, but they were better than that in pulmonary arterial hypertension model group, which had conspicuous significance of statistics. MMSCs labeled with Hoechst 33342 were observed that they permanent planted and differentiated into plentiful collateral circulations in lung observed by fluorescence microscope. The pulmonary arteries remodeling were attenuated effectively in MMSCs transplanted group. Pulmonary fibrosis and tumorous were not observed in rats of MMSCs transplanted group.CONCLUSION: These results indicate that MMSCs reduce and even reverse the progression of pulmonary arterial hypertension (PAH) induced by crotaline. It restores the structure and function of microvasculature with marked improvement of tissue and structure changes of the lung.     

Haemodynamic changes in high altitude pulmonary edema and effects of oxygen breathing

AIM:To explore the pathogenic mechanism of high altitude pulmonary edema(HAPE).METHODS:Haemodynamic changes and effects of 100 percent oxygen breathing were measured by Swan-Ganz thermistor catheters, high altitude healthy volunteers were served as controls.RESULTS:The important features of haemodynamic changes in HAPE: (1)Pulmonary arterial pressure was raised; (2)Pulmonary arterial resistance and cardiac output were raised; (3)Pulmonary artery wedge pressures and right atrial pressure were normal; (4)Pulmonary arterial pressure and resistance were induced by oxygen breathing.CONCLUSIONS:The normal pulmonary artery wedge pressures with a high cardiac output indicated that HAPE was recognized as a form of noncardiogenic pulmonary edema. The pulmonary hypertension may play an important role in the development of HAPE.     

Effects of early intervention of adipose-derived mesenchymal stem cells on function of pulmonary arterioles in rats with pulmonary arterial hypertension induced by monocrotaline

AIM：To investigate the effects of early intervention of adipose-derived mesenchymal stem cells (ADMSCs) on the function of pulmonary arterioles in monocrotaline (MCT)-induced pulmonary arterial hypertensive (PAH) rats. METHODS：ADMSCs were obtained from subcutaneous adipose tissue in the rat groin region and isolated by collagenase digestion. Ninety male SD rats were randomly divided into normal control group, PAH group and early intervention of ADMSCs (ADMSCs-EI) group. PAH was induced by MCT at dose of 40 mg/kg intraperitoneally. ADMSCs were delivered via left jugular vein 1 weeks after MCT administration. At the 1st, 2nd and 3rd weeks after transplantation of ADMSCs, mean pulmonary arterial pressure (MPAP) was measured by catheterization. Pulmonary arteriolar endothelium-dependent relaxation (EDdR), endothelium-independent relaxation (EDiR) and vasoconstrictive function (VCF) were evaluated by isolated vascular ring tension. The potency of vascular contraction and relaxation was expressed as the pD2 value. RESULTS：Compared with control group, pulmonary arteriolar EDdR in PAH group was obviously decreased 2 weeks after MCT administration. Meanwhile, compared with PAH group, pulmonary arteriolar EDdR in ADMSCs-EI group was obviously increased 1 week after early transplantation of ADMSCs. Compared with control group, MPAP was significantly increased and pulmonary arteriolar EDdR and EDiR were significantly decreased in PAH group 3 and 4 weeks after MCT administation. Meanwhile, compared with PAH group, MPAP was markedly decreased and pulmonary arteriolar EDdR and EDiR were markedly increased in ADMSCs-EI group 2 and 3 weeks after early transplantation of ADMSCs. However, pulmonary arteriolar VCF was not different among all groups. CONCLUSION：Early intervention of ADMSCs significantly ameliorates the impaired pulmonary arteriolar EDdR and EDiR, and greatly decreases MPAP in MCT-induced PAH rats.     

Effects of Danshensu on TGF-β1/Smads signaling pathways in rats with pulmonary fibrosis

AIM: To study the preventive and curative roles of Danshensu (DA) in bleomycin (BLM)-induced pulmonary fibrosis in rats. METHODS: Pulmonary fibrosis was induced in SD rats by intratracheal instillation of BLM. The rats were intraperitoneally injected with dexamethasone (1 mg·kg-1·d-1, DXM group), DA (15 mg·kg-1·d-1, DA group), or physiological saline (2 mL·d-1, BLM group). Normal controls (NC group) received physiological saline both intratracheally and intraperitoneally. At the 28th day after modeling, the histological changes of the lungs were evaluated by hematoxylin-eosin (HE) and Masson’s trichrome staining. The protein levels of α-smooth muscle actin (α-SMA) in the lung tissues were detected by the method of immunohistochemistry. The mRNA expression of transforming growth factor beta 1 (TGF-β1), Smad3 and Smad7 was assessed by real-time fluorescence quantitative PCR. RESULTS: Compared with BLM group, the degree of inflammation and fibrosis of the lung in DA group was obviously reduced, and so was the expression of α-SMA in the lung tissues. The mRNA expression of TGF-β1 and Smad3 in the lung tissues of the rats decreased and the mRNA expression of Smad7 increased. CONCLUSION: DA alleviates BLM-induced pulmonary fibrosis in rats in the early stage by inhibiting the expression of TGF-β1/Smad3 and stimulating the expression of Smad7 in the lung tissues.     

Inhibition of plasminogen activator inhibitor-1 by small interfering RNA attenuates pulmonary fibrosis in rats

AIM: To examine the effects of silencing of plasminogen activator inhibitor-1 (PAI-1) expression by small interfering RNA (siRNA) on bleomycin (BLM)-induced rat pulmonary fibrosis. METHODS: Total 72 Wistar rats were divided into 4 groups: control, BLM, BLM+non-specific siRNA (BLM+N), and BLM+ PAI-1 siRNA (BLM+P). Pulmonary fibrosis was induced by intratracheal injection of BLM (5 mg/kg), whereas equal volume of normal saline was used in control group. After the administration of BLM or normal saline, the rats were treated with tracheal injection of PAI-1-siRNA (7.5 nmol/0.2 mL per rat) in BLM+P group, non-specific siRNA (7.5 nmol/0.2 mL per rat) in BLM+N group, and 0.2 mL normal saline in BLM group and control group, twice a week, 8 times in 28 d. On day 7, 14, and 28, the rats (n=6 at each time point) were sacrificed. The bronchoalveolar lavage fluid (BALF) from the left lung was harvested to examine the activity of PAI-1. The mRNA expression of collagen type Ⅲ, α-smooth muscle actin (α-SMA) and tissue inhibitor of metalloproteinase-1 (TIMP-1) in the middle lobe of the right lung was detected by RT-PCR. RESULTS: PAI-1 activity and the expression of collagen type Ⅲ, α-SMA and TIMP-1 were increased in BLM group on day 7, 14 and 28. Intratracheal injection of PAI-1 siRNA twice a week continuously reduced PAI-1 activity in the BALF (P<0.05),and decreased the expression of collagen type Ⅲ, α-SMA and TIMP-1 in the fibrotic lung tissues on day 7, 14 and 28. Statistical differences in the expression of collagen type Ⅲ, α-SMA and TIMP-1 between BLM+P group and BLM group at the same time point were observed. CONCLUSION: Intratracheal injection of PAI-1 siRNA twice a week continuously inhibits the expression of PAI-1. PAI-1 siRNA ameliorates BLM-induced pulmonary fibrosis by down-regulation of TIMP-1 expression.     

Pathogenesis of endothelial-mesenchymal transition in pulmonary arterial hypertension

Endothelial-mesenchymal transition (EndMT) is a biological process through which endothelial cells change their endothelial phenotype into a mesenchymal or myofibroblastic phenotype with the expression of mesenchymal markers such as α-smooth muscle actin (α-SMA) and vimentin. When the pulmonary vascular endothelial cells are exposed to hypoxia and inflammatory stimulation, vascular smooth muscle cells in the outer and middle membrane accumulate through EndMT process, leading to pulmonary vascular remodeling and pulmonary arterial hypertension (PAH). Transforming growth factor β (TGF-β)/bone morphogentic protein (BMP) signaling pathways promote EndMT process by bone morphogentic protein receptor 2 (BMPR2) gene mutation which up-regulates high mobility group protein A1 (HMGA1) gene and contributes to protein expression such as Slug and Snail, thus resulting in PAH. In brief, TGF-β/BMP signaling pathways and related regulators play an important role in pulmonary vascular reconstruction and the formation of PAH.     

Effect of human tissue factor pathway inhibitor gene transfection in vein grafts on thrombus formation and early patency rate

AIM: To reduce thrombus formation after coronary artery bypass graft,we investigated the antithrombotic effect of human tissue factor pathway inhibitor (TFPI) gene delivery on vein grafts. METHODS: The eukaryotic expressed plasmid vector pCMV-(Kozak)TFPI was constructed. Through pressurizing infusion,vein endotheliocytes were transfected with cationic liposome containing the plasmid vector pCMV-(Kozak)TFPI. After operation, vein grafts were harvested at third day for immunohistochemical, RT-PCR and Western blotting analysis of exogenous gene expression and for observation of thrombus formation by pathological method and scanning electron microscopy. At 30th days, the patency rate was recorded by vessel Doppler ultrasonography. RESULTS: Human TFPI mRNA and protein were detected in TFPI gene transferred vein grafts. Thrombosis was found in 8 animals of empty plasmid control group and in 7 animals of empty control group, but in only 1 of the TFPI group (P<0.05). Thirty days after operation, 5 vein grafts were occluded in both empty plasmid control group and empty control group, but none of vein grafts were occluded in TFPI group (P<0.05). The endothelial surfaces of the vein grafts in both control groups were covered with aggregated erythrocytes and platelets, but not in TFPI group. CONCLUSION: Human tissue factor pathway inhibitor gene transfection reduces thrombus formation and improves early patency rate in vein grafts.     

Effect of angiontensin-(1-7) on development of monocrotaline induced pulmonary arterial hypertension and vascular remodeling

AIM: To investigate the effect of angiotensin-(1-7) ［Ang-(1-7)］ on the development of monocrotaline (MCT) induced pulmonary arterial hypertension and vascular remodeling.METHODS: 60 Sprgue-Dawely rats were randomly assigned into three groups: control group, PAH group and PAH+Ang-(1-7) group. Rats in PAH group and PAH +Ang-(1-7) group received 60 mg/kg MCT injection subcutaneously and after 24 h received either saline or 24 μg·kg-1·h-1 of Ang-(1-7) injection via osmotic minipumps for 4 weeks. These rats in control group were firstly injected saline subcutaneously and then received saline injection via osmotic minipumps.RESULTS: After 4 weeks, in PAH group, right ventricular systolic pressure (RVSP), right ventricular hypertrophy index (RVHI), percentage of wall thickness (WT%) and percentage of wall area (WA%) of pulmonary artery were significantly increased and NO concentration, the level of endothelial nitric oxide synthase (eNOS), eNOS Ser1177-phosphorylation were significantly decreased compared with control group. However, RVSP, RVHI, WT %, WA % were dramatically decreased in PAH +Ang-(1-7) group and NO concentration, the level of eNOS protein, eNOS Ser1177-phosphorylation were significantly increased compared with PAH group.CONCLUSION: These results suggest that Ang-(1-7) could prevent the development of monocrotaline induced pulmonary arterial hypertension and vascular remodeling, which appears to be associated with up-regulation of NO concentration and the level of eNOS protein, eNOS Ser1177-phosphorylation.     

Effects of Foxp3 transfection on splenocyte functions in asthma mice

AIM: To study the expression and the effects of Foxp3 on the immunologic functions by transfecting the Foxp3 eukaryotic expression plasmid into the splenocytes of the asthma mice. METHODS: The mice were sensitized and challenged by ovalbumin to make asthma model. The splenocytes were harvested and cultured. The Foxp3 expression vector pcDNA3.1(-)-Foxp3 was transfected into the splenocytes with electroporation. The splenocytes transfected with empty vector and control splenocytes (non-transfected) were also set up. The expression of Foxp3 at mRNA and protein levels was detected by RT-PCR and Western blotting, respectively. The proportion of CD4⁺CD25⁺ Treg cells/CD4⁺ cells was measured by flow cytometry. Proliferation of the splenocytes was analyzed with MTT assay. ELISA was used to determine the levels of interleukin 4 (IL-4) and interferon γ (IFN-γ) in the supernatant of the splenocytes. RESULTS: The expression of Foxp3 at mRNA and protein levels in transfection group was significantly higher than that in empty vector group and control group. The proportion of CD4⁺CD25⁺Treg cells/CD4⁺ cells in transfection group was higher than that in empty vector group and control group. The proliferation of transfected cells was markedly inhibited compared with empty vector group and control group. The levels of IL-4 and IFN-γ were significantly lower in transfection group than those in empty vector group and control group. CONCLUSION: The transfected Foxp3 gene overexpresses in the splenocytes of asthma mice. Foxp3 increases the number of CD4⁺CD25⁺ T cells and inhibits the proliferation and production of Th1/Th2 cytokines in splenocytes.     

Effect of propofol on expression of PKC mRNA in pulmonary injury induced by ischemia-reperfusion in rabbits

AIM: To investigate the effect of propofol on expression of protein kinase C (PKC) mRNA during pulmonary ischemia and reperfusion injury (PIRI) in rabbits. METHODS: Single lung ischemia and reperfusion animal model was used in vivo. The rabbits were randomly divided into three groups (n=9 in each): sham operated group (sham), PIR group (I-R) and PIR+ propofol group (PPF). Changes of several parameters including malondialdehyde (MDA), superoxide dismutase (SOD), wet to dry ratio of lung tissue weight (W/D) and index of quantitative assessment of histologic lung injury (IQA) were measured at 60 minutes after reperfusion in lung tissue. Meanwhile the location and expression of PKC mRNA were observed. Lung tissue was also prepared for light microscopic and electron microscopic observation at 60 minutes after reperfusion. RESULTS: As compared with group I-R, PKC mRNA strongly expressed in intima and extima of small pulmonary artery as well as thin-wall vessels (mostly small pulmonary veins) in PPF group. The average optical density values of PKC-α、δ and θ mRNA in small pulmonary veins PPF in group showed significantly higher than that in I-R group (all P<0.01). SOD increased and MDA, W/D and IQA decreased at 60 minutes after reperfusion in lung tissue (P<0.01 and P<0.05). Abnormal changes of the lung tissue in morphology were lessen markedly in PPF group. CONCLUSIONS: Propofol produces notable protective effects on PIRI in rabbits by activating PKC-α， δ and θ mRNA expression in lung tissue, raising NO level, dropping OFR level and decreasing lipid peroxidation.     

Up-dating research of the factors in pulmonary artery hypertension

Pulmonary artery hypertension is a serious disease in respiratory system and a key tache in the mechanism of pulmonary-heart disease. The pathological changes include the contraction and remodeling of the pulmonary vessels. There are more and more studies on the pulmonary artery hypertension because of its refractory character and the following increasing mortality. This article summarizes the updating research of the factors and mechanism studied on pulmonary artery hypertension recently, to provide a new view for the clinical and basic medical investigation.     

Relationship between airway inflammation and pulmonary arterial remodeling in rats with chronic bronchitis and emphysema

AIM:To study the effects of airway and pulmonary inflammation on pulmonary arterial remodeling in rats with chronic bronchitis (CB) and emphysema.METHODS:Twenty-four male Wistar rats were divided into three groups (n=8): Group A: four-weeks CB and emphysema;Group B: six-weeks CB and emphysema group;Group C: normal control.The rat model of CB and emphysema was established by intratracheal instillation of lipopolysaccharide (LPS) and daily exposure to cigarette smog.The arterial blood gas analysis,pulmonary hemodynamics changes and cell counts in bronchoalveolar lavage fluid (BALF) were measured.The pathomorphological changes of airway inflammation,alveoli destruction and pulmonary arterial remodeling were observed by HE straining and triple straining.RESULTS:(1) The characteristic pathological changes of CB and emphysema were observed in group A and B.Neutrophils were the main cells infiltrated into the walls of airway in group A.Lymphocytes and macrophages were the main cells in group B.(2) Right ventricular systolic pressure (RVSP),mean pulmonary arterial pressure (mPAP),the ratio of the weight of right ventricle/left ventricle and septum (RV/LV+S) in group A and B were significantly higher than those in group C (P<0.05).The amount of muscular artery (MA) in group A and B were significantly higher than that in group C (P<0.05).(3) In group A and B,the levels of MA,RVSP,mPAP and RV/LV+S was correlated positively with the average alveolar area,the total cell counts and differential cell counts of neutrophils,lymphocytes and macrophages in BALF,and the level of infiltration into the walls of airway,respectively (P<0.05).The positive correlation was observed with the percentage of neutrophils,lymphocytes and macrophages between group A and B (P<0.05).The amounts of MA were also correlated positively with RVSP,mPAP and RV/LV+S (P<0.05).CONCLUSIONS:(1) The pulmonary artery hypertension,the right ventricular hypertrophy and the pulmonary arterial remodeling appeared before hypoxia.These may be related with the degree of the pulmonary inflammation.(2) The characteristic of pulmonary arterial remodeling was small artery organization,and correlated positively with the changes of hemodynamics.     

Expression of programmed cell death factor 4 in pulmonary fibrosis model

AIM:To detect the expression of programmed cell death protein 4 (PDCD4) in pulmonary fibrosis model and its effect on cell viability and pulmonary fibrosis indicators, and to explore its mechanism. METHODS:The expression level of PDCD4, α-smooth muscle actin (α-SMA) and collagen type I (COL-I) in human embryonic lung fibroblasts cell line HFL-1 group and HFL-1+TGF-β1 group were detected by Western blot and RT-qPCR. The plasmid pEZ-M03-PDCD4 and empty vector pEZ-M03 were transfected into myofibroblasts (MB), and the protein expression level of PDCD4 was detected by Western blot. The protein levels of p-AKT and AKT in blank control group, pEZ-M03-PDCD4 group, pEZ-M03 group and LY294002 group and the expression of cell cycle-related proteins c-Myc and cyclin D1 were determined by Western blot. The effect of PDCD4 on the viability of MB was measured by CCK-8 assay. The hydroxyproline content in the culture supernatant of HFL-1 cells and transfected MB was detected by hydroxyproline digestion method. The expression of PDCD4 in the lung tissues of the mice in model group and control group was detected by Western blot. RESULTS:Compared with HFL-1 group, the expression of α-SMA and COL-I at mRNA and protein levels in HFL-1+TGF-β1 group was significantly increased (P<0.01), the mRNA expression of PDCD4 was not significantly changed, while PDCD4 protein was significantly down-regulated (P<0.01). Compared with blank control group and pEZ-M03 group, the protein expression of PDCD4 in pEZ-M03-PDCD4 group was significantly increased (P<0.05), the protein expression of c-Myc and cyclin D1 was significantly decreased (P<0.05), the cell viability was also significantly inhibited (P<0.01), and the content of hydroxyproline in the culture supernatant was significantly reduced (P<0.05). Compared with blank control group, the protein levels of p-AKT were significantly decreased in pEZ-M03-PDCD4 group and LY294002 group, and no significant difference between blank control group and pEZ-M03 control group was observed. Compared with control group, PDCD4 expression was decreased in model group (P<0.01).CONCLUSION:PDCD4 is low expressed in the process of pulmonary fibrosis. Over-expression of PDCD4 inhibits the viability of MB, decreases the content of hydroxyproline, and inhibits the PI3K/AKT signaling pathway.     

Role of EndoMT in HHPH based on TGF-β/Smads signaling pathway and regulated by Yiqi-Wenyang-Huoxue-Huatan formula

AIM: To investigate the relationship between transforming growth factor-β (TGF-β)/Smads signaling pathway and pulmonary arterial endothelial-mesenchymal transition (EndoMT) in hypoxia-hypercapnia pulmonary hypertension (HHPH) process and the regulatory effect of Yiqi-Wenyang-Huoxue-Huatan formula (YWHHF). METHODS: Healthy male SD rats were randomly divided into 5 groups:normal control (N) group, hypoxia-hypercapnia (HH) group, high-dose YWHHF (YH) group, middle-dose YWHHF (YM) group and low-dose YWHHF (YL) group. The rats in N group was housed in normoxic environment, and the rats in the other 4 groups were housed in hypoxia-hypercapnia environment (9%~11% O₂ and 5%~6% CO₂) for 4 weeks, 8 h/d, 6 d/week. The excess water vapor was absorbed by anhydrous CaCl₂, and CO₂ was absorbed by sodium hydroxide. The rats in YWHHF groups were put into the oxygen chamber before the same volume of YWHHF at different concentrations were given (200 g/L for YH group, 100 g/L for YM group and 50 g/L for YL group). The average pulmonary artery pressure and the average carotid artery pressure were measured during the operation. After operation, the right ventricular free wall and left ventricle plus interventricular septum were collected for determining the right ventricular hypertrophy index. Moreover, the morphological changes of the lung tissues were observed under light microscope. The mRNA and protein levels of α-smooth muscle actin (α-SMA), CD31, TGF-β1 and Smad2/3, and the protein level of p-Smad2/3 were detected by RT-PCR and Western blot. RESULTS: Compared with N group, the pulmonary artery mean pressure, the mRNA and protein expression of α-SMA, TGF-β1 and Smad2/3, and the protein level of p-Smad2/3 were increased, the levels of CD31 were decreased (P<0.05), and the lung tissue damage was observed in the other 4 groups. Compared with HH group, the pulmonary artery mean pressure, the mRNA and protein expression of α-SMA, TGF-β1 and Smad2/3, and the protein level of p-Smad2/3 were decreased, while the mRNA and protein levels of CD31 were increased. Moreover, the lung tissue damage was reduced in YH, YM and YL groups. CONCLUSION: TGF-β/Smads pathway may be involved in the process of EndoMT under hypoxia and hypercapnia condition, and YWHHF may reduce EndoMT by inhibiting the expression of TGF-β/Smads pathway-related molecules.     

Effects of c-fos antisense oligoneuleotide and p21 genetic transfection on the intimal proliferation of venous autografts in rabbits

AIM: To investigate the effects of c-fos antisense oligoneuleotide and p21 genetic transfection on the intimal proliferation of venous autografts. METHODS: The external jugule veins were autografted into common carotid arteries in the same side in 20 New Zealand rabbits, which were divided evenly into experimental and control group randomly. The transplanted veins of experimental group were immersed in the adenovirus-mediated p21 gene solution for 15 minutes just before anastomosis and coated with c-fos antisense oligoneucleotide glue gel just after anastomosis, while the control was only treated with empty vector. The transplanted vascular sample were taken at 2 weeks after operation. The intimal thickness (IT), degree of restenosis (DR), expression of proliferating cell nuclear antigen (PCNA), quantity of VSMC were determined by immunohischemistry. RESULTS: The IT, DR and expression of PCNA, VSMC were decreased, compared to control group. CONCLUSION: Transfection of c-fos antisense oligoneuleotide and p21 gene inhibits the intimal proliferation of venous antografs.