Rat Copd Airway Fibroblasts Transfection Rhsg Gene After Observation Of Cell Proliferation, Apoptosis And Tgf - Beta 1, Pdgf Expression Level

Objective:This study was constructed to carry proliferation inhibition gene (rHSG) and eukaryotic expression vector, transfected into chronic obstructive pulmonary disease (COPD) in rat airway fibroblasts, observe the expression level relationship between COPD rat airway fibroblast proliferation, apoptosis and transforming growth factor (TGF-β1), the platelet-derived growth factor (PDGF), To lay the foundation for lung diseases such as chronic obstructive pulmonary disease cause by intervention in airway fibroblast proliferation and provide theory basis for find out the progress to delay the COPD disease.Methods:Using the SD male rats construct COPD rat model, applying tissue explants wall method, in vitro COPD rat airway primary fibroblast cells, and cells were identified with vimentin, take three generations of cells for subsequent experiments. Through molecular cloning, build carry rHSG gene eukaryotic expression vector, and the experimental groups:control group (COPD model untransfected), empty vector pEGFP-nl (and) transfection pEGFP-rHSG group, experimental cationic liposomes as transfection medium, the recombinant plasmid was transfected into a model of COPD rat airway fibroblasts, were infected with24hours,48hours,3days,5days,7days, using the MTT assay observed the capacity of transfection fibroblasts proliferative; by western blot was detected the protein content after transfection of rHSG, TGF-β1and PDGF; Total RNA was extracted, reverse transcription polymerase chain reaction (RT-PCR), detection the expression levels of TGF-β1, PDGF and rHSGmRNA. The data were statistically analyzed using the application SPSS18.O software, get by the experimental data express by x±s, using one-way ANOVA analysis of the mean, standard deviation, P<0.05was considered significant statistically significance.Results:COPD rat airway fibroblasts in primary culture success vimentin identification positive after immunohistochemistry. As the rHSG gene transfection in rat airway fibroblasts prolonged the inhibition of fibroblast proliferation in rHSG gene transfection group were significantly higher than the model control group; mRNA expression of TGF-β1and PDGF two genes reduction appear in the transfection in24hours, reduce the value of mRNA expression reached a maximum in7days, the amount of mRNA expression also reduced in5days to7days, but the magnitude was reduced smaller than before, each transfection group, TGF-(31and_PDGFmRNA express the amount of difference to meet the P<0.05.when pEGFP-rHSG transfected into cells to24hours, PDGFmRNA expression begins to increase quantity, added value reached a maximum at7days, the transfected with the5d-7d expression still continue to increase, but the increase reduced than the previously, rHSG/Mfn2mRNA expression differences were to meet (P<0.05) in each transfection group.Conclusion:The balance relationship between rHSG gene and TGF-β1, PDGF gene is broken, is the vital factors in COPD airway remodeling. rHSG gene transfection group with longer transfected time enough to inhibit obviously infection COPD airway into fiber cell proliferation, instead COPD airway fibroblasts, TGF-β1and PDGF With the rHSG gene infection time to extend the low expression, confirmed that when the exogenous rHSG Gene transfected to the COPD rat airway fibroblasts can regulate the over expression of TGF-β1and PDGF genes, thereby enabling the airway fibroblast proliferation was inhibited. Therefore the efficient rHSG gene transfected with the COPD airway remodeling in progress may slow down to a certain extent.