The Possible Mechanism Of Wenshen Yiqi Granules On The Treatment Of COPD:Activation Of SIRT1/PPARγ Pathway And Inhibition Of NLRP3 Inflammasome

[Objective]Study on the effects of Wenshen Yiqi Granules on inflammation,apoptosis,NLRP3 inflammasome and SIRT1/PPARγexpression in COPD rats.And investigate the mechanism of Wenshen Yiqi Granules activation of SIRT1/PPARγ to inhibit NLRP3 inflammation,reducing the inflammation and apoptosis in AECII cells.[Method]1.Animal experiments:Study the effects of Wenshen Yiqi Granules on inflammation,,apoptosis,NLRP3 inflammasome and expression of SIRT1/PPARγ in COPD ratsCOPD rat model established by smoking and airway injection of LPS.Different doses of Wenshen Yiqi granules and dexamethasone(DEX)were used to intervene COPD rats.Magogee staining was used to detect the leukocyte classification and number in rat bronchoalveolar lavage fluid;HE staining was used to observe the infiltration of inflammatory cells in rat lung tissue;TUNEL staining was used to detect the apoptosis of rat lung tissue;MPO kit was used to detect the MPO activity of lung tissue;ELISA was used to detect the contents of INF-γ,TNF-α,IL-18 and IL-1β in rat serum,bronchoalveolar lavage fluid,and lung tissue;Western blot and qRT-PCR methods were used to detect the SIRT1,PPARγ,NLRP3 inflammasome and apoptosis-related proteins expression and mRNA levels in rat lung tissue;The expression and distribution of PPARy and SIRT1 in lung tissue were detected by histochemistry.2.Cell experiment:Explore the mechanism of Wenshen Yiqi Granules activation of SIRT1/PPARγ pathway and inhibition of the activation of NLRP3 inflammasome and reducing the inflammation and apoptosis of type Ⅱ alveolar epithelial cells1.The effects of Wenshen Yiqi medicated serum on SIRTl/PPARypathway,NLRP3 inflammasome,apoptosis and inflammatory factor secretion in CSE-treated type Ⅱ alveolar epithelial cells:Wenshen Yiqi medicated serum treated 10%CSE-treated type Ⅱ alveolar epithelial cells(AECII)cells.Cell viability was detected by CCK-8;cell proliferation was detected by EdU staining;cell apoptosis was detected by flow cytometry;INF-γ,TNF-α,IL-18 and IL-1β in cell supernatant were detected by ELISA.Western blot and RT-PCR were used to detect the expression of SIRT1,PPARy,NLRP3 inflammasome and apoptosis-related proteins in cells.2.Explore the mechanism of Wenshen Yiqi Granules activation of SIRT1/PPARγpathway,inhibition of the activation of NLRP3 inflammasome and reducing cell inflammation and apoptosis:The adenovirus vector was constructed to verify its gene silencing and overexpression effects on PPARy,SIRT1 and NLRP3.The combination of Wenshen Yiqi medicated serum with SIRT1/PPARy/NLRP3 pathway interfered treated COPD cell model.Cell viability was detected by CCK-8;cell proliferation was detected by EdU staining;cell apoptosis was detected by flow cytometry;INF-γ,TNF-α,IL-18 and IL-1β contents in cell supernatant were detected by ELISA.Western blot and RT-PCR were used to detect the expression of NLRP3 inflammasome and apoptosis-related proteins in cells.[Results]Wenshen Yiqi Granules may significantly improve the pulmonary inflammatory injury in COPD rats,inhibit the level of MPO,apoptosis in lung tissue,INF-γ,TNF-α,IL-18 and IL-1β in serum,bronchoalveolar lavage fluid and lung tissue homogenate,inhibit the protein expression and mRNA level of NLRP3,ACS,Caspase-1,IL-18,IL-1β and BAX in lung tissue,promote the expression of BCL2 and the distribution of PPARγ and SIRT1 in lung tissue.Treating CSE-induced COPD cell model with Wenshen Yiqi containing serum,the cell viability increased,the apoptosis decreased,the contents of INF-γ,TNF-α,IL-18 and IL-1β decreased,and the protein expressions and mRNAs of BCL2,PPARy and SIRT1 increased,the level of NLRP3,ACS,Caspase-1,IL-18,IL-1β,BAX protein expression and mRNA decreased.Knockdown of PPARy or overexpression of NLRP3 reversed the amelioration effect of Wenshen Yiqi-containing serum on CSE-induced COPD cell injury,and overexpression of SIRT1 enhanced the amelioration effect of Wenshen Yiqi-containing serum on CSE-induced COPD cell damage.[Conclusion]Wenshen Yiqi Granules activate SIRT1/PPARγ pathway to inhibit the activation of NLRP3 inflammasome,reducing COPD injury and inflammatory response.