Rat lung microvascular endothelial cells (LMVEC) were cocultured with rat pulmonary arterial smooth muscle cells (PASMC). The cultures were divided into 5 groups: normal group (N), lipopolysaccharide (LPS) 2h (L 2), LPS 8h (L 8), LPS 16h (L 16 ), and LPS 24h (L 24 ). The activity of IL 6 in LMVEC and PASMC supernatants was determined by radio immunity method, and the expression level of IL 6 mRNA in LMVEC was detected with RT PCR. The results showed that the activity of IL 6 in LMVEC and PASMC was enhanced by LPS at L 2, reaching the higher level after 8 hour stimulation. Furthermore, the activity of IL 6 was stronger in homotypic groups than that of coculture groups. The expression of IL 6 mRNA in LMVEC was increased in L2 group, and highest in L6 group, but lowered in L12 group, though it was still stronger than that of N. It suggest that IL 6 mRNA and the activity of IL 6 in LMVEC and PASMC were enhanced in both homotypic and coculture groups by LPS. And the mRNA level and activity of IL 6 were higher in homotypic groups than that of coculture groups.

Objective To explore the effect of hypoxia on the expression of Janus kinase (JAK) in cocultured pulmonary arterial smooth muscle cells (PASMC) Methods Rat PASMC were cocultured with lung microvascular endothelial cells (LMVEC), the expression of JAK protein in PASMC was detected with Immunocytochemical method Results Expressions of JAK1 and JAK3 Protein were seen excessive positive (++++) in the cytoplasm of simple PASMC in hypoxia, and better positive (+++) in cocultured PASMC in hypoxia Compared with those of the control group, the expressions of JAK1 and JAK3 protein obviously increased in PASMC in hypoxia for 2 h, reached highest in those with hypoxia for 6 h, and decreased in those with hypoxia for 12 hours hypoxia, but still higher than those in hypoxia for 2 h Conclusion Hypoxia modulates the signal pathways and improves cell proliferation by the enhancement of the expressions of JAK1 and JAK2 proteins in co cultured PASMC

Objective To observe the proliferation and phenotype-switching of pulmonary arterial smooth muscle cell (PASMC) induced by hypoxia and interfered by Ad-PKGIα. And to investigate the potential regulative role of PKGIα gene in the molecule mechanism of hypoxia pulmonary vessel remodeling (HPVR). Methods To establish the pure PASMC cultured by tissue-sticking methods. Semi-quantitative reverse transcription and polymerase chain reaction (RT-PCR) and Western blot were used to examine the PKGIα mRNA and protein expression after PASMC were transfected by Ad-PKG. The mRNA and protein expressive change of smooth muscle α actin(SM-α-actin) determined the degree of cell phenotype-switching. The changes of PASMC proliferation were determined by flow cytometry and ~3H-TdR incorporated way. Results Ad-PKGIα could transfect into PASMC and highly express. Hypoxia down-regulated the expression of SM-α-actin protein (44. 25±5.34 in normoxia, 32. 18±4. 19 in 12 h hypoxia condition, 21.90 ±2. 44 in 24 h hypoxia condition, P < 0. 05), that could be blocked by the transfeetion of Ad-PKGIα. Hypoxia could push PASMC mitosis and proliferating(~3H-TdR incorporated way: 7570 ± 371 in normoxia,12 020± 831 in 12 h hypoxia condition,14 924 ± 1491 in 24 h hypoxia condition, P <0. 05), that could be blocked by the transfection of Ad-PKGIα, too. Conclusions The results suggested that PKGIα signaling pathway might play an important role in the molecule mechanism of HPVR. And PKGIα gene might be a target point of gene therapy.

Objective　To study the expressions of some apoptosis related to genes in lung tissues and their relationship to the pathogenesis in rats to hypoxic pulmonary hypertension (HPH). Methods　A total of 60 rats were employed and equally divided into 5 groups, i.e. control, HPH, hemin, Tin Protoporphyrin (Snpp) and low concentration CO groups. Of them, the rats from middle 3 groups were treated with hypoxia under normal pressure for 7 h in every day, 3 weeks except Sunday. Hemin and Tin Protoporphyrin (Snpp) were given half an hour before hypoxia while low concentration CO after 2 h of hypoxia. The expression of bcl-2, bax, Fas and Fas ligand (FasL) were detected with immunohistochemical staining, in situ cell death detection, DNA fragment detection, and in situ hybridization. Results　In low concentration CO and hemin group, typical DNA ladder bands and apoptotic cells were found in lung tissues, and the expressions of Fas, FasL, bax, and bcl-2 mRNA were increased in the epithelial cells of alveoli, vascular and bronchial walls. And the expression of bcl-2 mRNA was decreased than in hypoxic group. Conclusion　CO can regulate the expression of bcl-2, bax, Fas and FasL, these apoptotic genes may involve in the regulation of cell apoptosis of lung tissues under HPH and play important roles in the genesis and development of chronic obstructive lung disorders.

AIM:To explore the effect of carbon monoxide (CO) on hypoxic proliferation of rat pulmonary arterial smooth muscle cell(PASMC). METHODS:Rats were randomly divided into four groups, including: (1)N:Normoxia group (21% O 2); (2)H:Hypoxia group (1% O 2); (3)CO+H:CO+hypoxia group (3% CO+1% O 2),tested by immunocytochemical analysis,[3H]-TdR incorporation and flow cytometry (FCM). RESULTS: (1)Flow cytometric DNA analysis revealed that hypoxia induced significantly enhancement of G 2/M phase and decreased G 0/G 1 phase of PASMC ( P

AIM: To study the expression level of heme oxygenase -1 and -2 in pulmonary arterial smooth muscle cell(PASMC) treated with carbon monoxide(CO). METHODS: (1) The expression of HO in cultured rat PASMC was determined by immunohistochemistry stain. (2) The levels of HO mRNA and protein were evaluated by reverse transcription- polymerase chain reaction(RT-PCR) and Western blot. RESULTS: (1) PASMC cytoplasm showed yellow (++-+++) in hypoxia group, the buffy color(++++) in CO group as cells were stained by HO-1 antibody. (2) The expression level of HO-1 mRNA (1.25?0.37) in hypoxia group was higher than that of normal group (0.12?0.04). It was much higher (3.52?0.68) in CO group, than that of hypoxia group. (3)The content of HO-1 protein (1.07?0.15) in hypoxia 48 h group was higher than that of hypoxia 24 h group(0.52?0.04). It was higher(3.65?0.43) in CO group than that of hypoxia group.It was highest in CO 48 h group. CONCLUSION: Low density of CO and hypoxia enhanced the expression of HO-1 mRNA and protein in PASMC when cell were treated in hypoxia or CO,indicating that HO-1 has regulatory effect on PASMC during hypoxia.

Objective To establish a cell line of rat lung microvascular endothelial cells (LMVEC) and investigate the activity of tyrosine kinase receptor (TKR) in them after lipopolysaccharide (LPS) treatment Methods Rat LMVEC were cultured with lung tissue block pasted methods, and were identified by using immuocytochemical methods with anti human CD31 and factor Ⅷ antibody, by testing their binding with the Lectin Bandeiraea simplicifoliaⅠwith immunofluoresence staining, and by measuring the activity of angiotensin converting enzyme (ACE) with ACE kit The cells were treated with LPS at 1?g/ml for 2, 8, 16, 24 h respectively The activity of TKR in the cells was determined with radioimmunoassay Results The self cultured LMVEC in flask showed regular cobblestone morphology and positive for binding of the Lectin Bandeiraea simplicifoliaⅠand anti human CD31 antibody The TKR activity in LMVEC increased in 2 h group, reached the highest in 8 h group, and fell in 16 h group, but still significantly higher than that in normal group Conclusion TKR activity increases in the cytoplasm of LMVEC, but decreases in the cell membrane, suggesting that TKR and its correlated signal pathways may exert important modulation in the mechanism of acute lung injury and systemic inflammatory response syndrome

Objective To compare the growth curve, protein content, calcium ion content of pulmonary arterial smooth muscle cells (PASMCs) cultured by enzyme digestion and tissue-sticking methods. To investigate whether PASMCs cultured by two ways could be offered to one series of signaling pathway studies. Methods The protein content was determined by upgraded-lowry, the calcium ion content by fluorescence indicator. The growth of PASMCs was observed by light microscope. Results There existed certain difference in protein content, growth curve between PASMCs cultured by the two methods, especially in calcium ion content (P

AIM: To explore the effect of exogenous carbon monoxide (CO) on the hypoxic pulmonary arterial hypertension in the rat. METHODS: 60 rats were randomly divided into 5 groups: Normal group\, hypoxic group\, Hermin group\, Znpp group and CO group. HO-1 activity was examined by spectrophotometer and lung tissues was stained by immunohistochemical method. HO-1 mRNA level was tested by reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: (1) The pulmonary arterial pressure (18.52?3.24 mmHg) in CO group was higher than anoxic group and lower than hypoxic group (P

Objective To analyze the change in the concentration of IL-17 and IL-10 inflammatory factors among the deadaptation personnel who returned from the plateau.Methods A total 21 healthy males were investigated who averaged 25 years in age, lived permanently in the plains (200 m), and once stayed to the plateau (Lasha) for 6 months.Their venous blood was collected at three time points:the day before ascending to the plateau(control), the second day after return to the plains(d2) and the 30th day(d30), respectively.Their serum was seperated from the whole blood and the level of IL-17A and IL-10 was detected by ELISA method.Results The concentration of IL-17A and the IL-17A/IL-10 ratio were significant increased at d2 and d30, respectively, compared with control (P<0.05).Compared with d2, IL-17A and the IL-17A/IL-10 ratio were decreased obviously at d30(P <0.05).The level of IL-10 at d2 and d30 was significantly reduced compared with control ( P <0.05), but increased at d30 compared with d2.Correlative analysis showed that there was a negative correlation in the levels of IL-10 and IL-17A between control, d2 and d30, respectively (r1=0.948, P<0.05;r2=0.969, P<0.05;r3=0.972, P<0.05).A significant negative correlation was observed in the alteration levels of IL-10 and IL-17A between the three groups(r4=-0.793, P<0.05; r5=-0.756, P<0.05). Conclusion The concentration of inflammatory factors among the plateau deadaptation patients is imbalanced, but it is gradually reduced with time.The mechanism is still not clear.