Effects Of Mechanical Pre - Stretch On The Expression Of HMGB 1 In Alveolar Type Ⅱ Epithelial Cells Induced By Excessive Mechanical Distraction

Part I Preconditioning of cyclic stretch inhibits the HMGB 1 expression induced by excessive cyclic stretch in type II alveolar epithelial cellObjectives:To investigate the effects of preconditioning of cyclic stretch on excessive cyclic stretch (CS) induced expression of high mobility group box 1 (HMGB 1) in type II alveolar epithelial cell.Methods:We culture A549 cell in vitro. A549 cells were randomly divided into six groups; I group (without medication), II group (20% CS for 6 hours); III (preconditioned by 5% CS for 15 min, then stimulated by 20% CS for 6 hours); IV (preconditioned by 5% CS for 30 min, then stimulated by 20% CS for 6 hours); V group (preconditioned by 5% CS for 60 min, then stimulated by 20% CS for 6 hours); VI group (preconditioned by 5% CS for 120 min, then stimulated by 20% CS for 6 hours). The expression levels of HMGB 1 was measured using Enzyme-Linked Immunosorbnent Assay (ELISA); the expression levels of HMGB 1 mRNA and HMGB 1 reportor mRNA were measured using Real-time polymerase chain reaction (RT-PCR).The quantification of Polymerase signal transducer and activator of transcription 3 (p-STAT3) was measured using Western blot analysis.Results:Compared with I group, the expression levels of HMGB1 and p-STAT3 significantly increased in II group CP<0.05). Compared with III group, at 30 min、60 min and 120 min of precondition of CS, the expression levels of HMGB1 and p-STAT3 decreased (P<0.05), which reached the peak at 60 min of precondition (P<0.05).Conclusions:Preconditioning of cyclic stretch inhibits the HMGB 1 expression induced by pathologically mechanical stretch in type II alveolar epithelial cell.Part II Preconditioning of cyclic stretch attenuated the HMGB 1 expression by through SOCS3/STAT3 pathwayObjectives:To investigate the effects of SOCS3/STAT3 pathway in preconditioning of cyclic stretch attenuated the HMGB 1 expression in type II alveolar epithelial cell.Methods:On the one hand, A549 cells were randomly divided into five groups:Control group, negative group, siRNA-SOCS3-lgroup, siRNA-SOCS3-2 group, siRNA-SOCS3-3 group. The expression levels of SOCS 3 mRNA was measured using RT-PCR. On the other hand, A549 cells were randomly divided into four groups; 20% CS group (20% CS for 6 hours),20% CS+5% CS+SOCS3 siRNA group (transfection of SOCS3 siRNA,5% CS for 60 min, then stimulated by 20% CS for 6 hours); 20% CS+5% CS group (5% CS for 60 min, then stimulated by 20% CS for 6 hours); 20%CS+5% CS+negative-siRNA group (transfection of negative-siRNA,5% CS for 60 min, then stimulated by 20% CS for 6 hours). The expression levels of HMGB 1 was measured using ELIS A; the expression levels of HMGB 1 mRNA was measured using RT-PCR. The quantification of p-STAT3 was measured using Western blot analysis.Results:The first experiment shown that the expression levels of HMGB 1 mRNA were significantly decreased in siRNA-SOCS3-1 group, siRNA-SOCS3-2 group and siRNA-SOCS3-3 group (P<0.05). The most obvious suppression of HMGB 1 mRNA was obtained in siRNA-SOCS3-2 group (P<0.05). The following experiments choice of siRNA were siRNA-SOCS3-2. In the second experiment, the expression levels of HMGB 1, HMGB 1m RNA and p-STAT3 significantly decreased in 5% CS+20% group compared to the 20% CS group (P <0.05). Preconditioning of cyclic stretch failed to suppress expression of HMGB 1, HMGB 1 mRNA and p-STAT3 in cells transfected with SOCS3 siRNA.Conclusions:Preconditioning of cyclic stretch attenuated the HMGB 1 expression by through SOCS3/STAT3 pathway...