| Acute and chronic inflammatory lung diseases such as ARDS/IPF are often associated with epithelial cell injury/loss.Resolvin D1(RvD1)is a lipid mediator derived from polyunsaturated fatty acids and is biosynthesized during the resolution phase of inflammatory response,which exerts potent anti-inflammatory and promotes resolution of inflammatory lung diseases.In addition,RvD1 can play a protective role on LPS-induced ALI.But the underlying mechanisms are not fully understood.In this project,we focused on effect and mechanism of RvD1 on Bronchial Epithelial cells and type Ⅱ alveolar epithelial cells,and tried to explore the mechanism and effect of RvD1 on lung inflammation resolution.Three parts of experiment were involved:Part ⅠEffects and Mechanisms of RvD1 on Alveolar Epithelial Wound Repair and Cell SurvivalRvD1 is biosynthesized during the resolution phase of inflammatory response,which exerts potent anti-inflammatory and promotes resolution of inflammatory lung diseases.We hypothesised that RvD1 has a trophic role to protect against alveolar epithelial damage and promote repair.Methods: Human primary alveolar type II cells(ATII cells)were isolated and cultured.ATII cells were mechanically wounded using a scratch wound assay.Wound repair with or without RvD1 was assessed by video microscopy.Cell proliferation was measured by BRDU incorporation.Cell viability was measured using cell titer.m RNA expression was measured by realtime-PCR.Protein expression was measured by Western-blot.Results: RvD1 promoted wound repair and proliferation in ATII cells through activation of ALX receptor and the PI3K/AKT signaling pathway.s Fas L induced cell death was reduced by addition of RvD1 suggesting that RvD1 reduced apoptotic cell death.Conclusion: Our data suggests that RvD1 may have therapeutic potential in lung inflammtion to promote alveolar epithelial repair and prevent Fas L induced apoptotic cell death in addition to its known anti-inflammatory effects.Part Ⅱ Resolvin D1 inhibit TGF-? induced EMT in Primary human alveolar type II cellsEpithelial–mesenchymal transition(EMT)is a process in which epithelial cells gradually acquire amesenchymal(fibroblast-like)cell phenotype.To explore effects and mechanisms of RvD1 on TGF-? induced EMT in Primary human alveolar type II cells.Methods: Human primary alveolar type II cells(ATII cells)were isolated and cultured.EMT was induced in ATII cells with TGF-? treatment.m RNA expression was measured by realtime-PCR.Protein expression was measured by Western-blot.Results: RvD1 inhibit TGF-? induced EMT in Primary human alveolar type II cells via activation of ALX receptor.Conclusion: Our data suggests that Inhibit TGF-? induced EMT is potentially vitally important for RvD1 if used as a novel therapy in both acute and chronic inflammatory lung diseases such as ARDS and IPF.Part Ⅲ RvD1 reduces lipopolysaccharide-induced expression of cyclooxygenase-2 in human bronchial epithelial cellsTo explore the effects of RvD1 on expression of cyclooxygenase-2(COX-2)in human bronchial epithelial cells(HBECs).Methods: HBECs were incubated with various concentrations(0.1、1、10ug/ml)of LPS for 9 h,or 1 μg/ml LPS for different times(3h、6h、9h).The COX-2 m RNA and prostaglandin E2(PGE2)of HBECs was measured.Then HBECs incubated in DMEM medium containing LPS(1ug/ml)for 9h were deal with the RvD1(0、100、400nmol/m L).After incubation,the supernatant was collected and the amount of PGE2 was measured by ELISA.The cells were harvested,and COX-2 m RNA and protein were analyzed by RT-PCR and Western-blot respectively.Results: LPS increased the expression of COX-2 m RNA and production of PGE2 in a dose and time dependent manner in HBECs.Induction of COX-2 m RNA and protein by LPS were inhibited by RvD1 in a dose dependent manner.RvD1 also significantly decreased LPS induced production of PGE2.Conclusion: RvD1 down regulates LPS induced expression of the COX-2 and consequently inhibits production of PGE2 in cultured HBECs. |
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