Molecular Mechanism Of Qingfei Yucuo Decoction Improving Acne Inflammation By Classical Pyrolysis Pathway

BackgroundAcne is a kind of chronic inflammatory skin disease of the sebaceous glands and the hair follicles.Inflammation is accompanied by the occurrence and end of acne.The Qingfei Yucuo Decoction,which has a good clinical effect in treating acne,is based on the Pipa Qingfei Yin of《Yi Zong Jin Jian》.ObjectiveProviding a scientific basis for its clinical treatment of acne through regulating TLR2/NF-κB pathway,impacting NLRP3 inflammasome and P.acnes-induced Caspase-1-dependent cell pyrolysis in vivo and in vitro experiments,and clarifying the molecular mechanism of acne treatment.Methods1、Studying the effect of Qingfei Yucuo Decoction on TLR2/NF-κB pathway,NLRP3 inflammasome and cell pyrolysis in vivo:Acne model was established by subcutaneous injection of Propionibacterium acnes in the external auricle of rats.Signs and HE slices were used to evaluate the effect of Qingfei Yucuo Decoction on auricle skin of rats;The tissue inflammatory factors IL-8,TNF-α,IL-1β and IL-18 were detected by enzyme-linked immunosorbent assay;TLR2 and P-IκBα and P-p65 protein expression were detced by Western blot;NLRP3,Caspase-1,IL-1β and IL-18 mRNA levels were determined by Real-time quantitative PCR;NLRP3 and IL-1βprotein expression were evaluated by immunohistochemistry.2、Exploring the effect of Qingfei Yucuo Decoction on P.acnes-induced pyrolysis of THP-1 cells:Co-culture of P.acnes and THP-1 cells was used to establish acne model in vitro;Cell Infection experiment was used to study the effect of Qingfei Yucuo Decoction on the infection of THP-1 Cells by P.acnes;Flow cytometry was used to investigate THP-1 pyrolysis rate;Spectrophotometry was used to detect its effect on the activity of Caspase-1;Fluorescent probe method was used to determine its effect on mtROS;Real-time quantitative PCR and Western blotting were used to detect the effect of Qingfei Yucuo Decoction on the expression of NLRP3,Caspase-1,IL-1β mRNA and protein.3、LC-MS/MS method was used to identify the active ingredients of Qingfei Yucuo Decoction.The effects of baicalin on NF-κB/MAPK pathway and NLRP3 inflammasome were studied in vivo and in vitro.Results1、Qingfei Yucuo Decoction could improve erythema,swelling and inflammatory cell infiltration of rat ears,reduce the levels of inflammatory factors IL-8,TNF-α,IL-1β and IL-18,inhibit the expression of TLR2,P-IκBα and P-p65 protein;Qingfei Yucuo Decoction aslo could down-regulate the mRNA levels of NLRP3,IL-1β and Caspase-1,inhibit the expression of NLRP3 and IL-1β protein and reduce the pyrolysis.2、Qingfei Yucuo Decoction could prevent P.acnes from infecting THP-1 cells,inhibit mtROS production,reduce NLRP3,Caspase-1 and IL-1β mRNA levels and protein expression,weaken caspase-1 activation and decrease Caspase-1 enzyme3、LC-MS/MS identification analysis showed that Qingfei Yucuo Decoction contained baicalin.Baicalin inhibited the production of IL-1β and IL-18,downregulated the phosphorylation of IκBα,JNK and ERK1/2,inhibited p65 entry into the nucleus,and down-regulated the expression of NLRP3,pro-Caspase-1,IL-1β mRNA and protein in vitro and in vivo.ConclusionsQingfei Yucuo Decoction can reduce Caspase-1 dependent pyrolysis caused by P.acnes,improve skin inflammation by regulating TLR2/NF-κB pathway and inhibit the activation of NLRP3 inflammasome.Baicalin can improve P.acnes-induced skin inflammation by down-regulate the NF-κB/MAPK pathway and inhibit activation of NLRP3 inflammasome,which has a good correlation with the mechanism of Qingfei Yucuo Decoction exerting anti-acne inflammation.In this study,the mechanism of Qingfei Yucuo Decoction in treating acne was revealed from the perspective of pharmacological effects and material basis.