Qianliexiaotang Intervenes TGF-?1/Smads Signaling Pathway To Regulate The Differentiation Of PrF Cells In EAP Mice

Objective : During the development of chronic prostatitis(CP),Prostate Fibroblast(PFR)is proliferated by inflammation,which in turn forms fibrosis of the prostate body,making it difficult for drugs to penetrate into the gland.It is an important factor leading to the characteristics of high recurrence and low cure rate of CP.Transforming growth factor-beta1(TGF-?1)is more complex,but it mainly acts through membrane receptors and Smads signaling pathways.The TGF-1/Smads-related signaling pathway has a strong regulatory effect on the differentiation of Th17 and fibroblasts,and has been extensively studied in recent years.TGF-1 plays a more complex role in the inflammatory process with fibrosis,and has a strong correlation with the overexpression and sustained high concentration of TGF-?1 in tissues.It can promote the differentiation of Th17 cells and inhibit the proliferation of PrF at lower concentrations.However,at higher concentrations,it can inhibit the differentiation of Th17 cells and promote the proliferation of PrF.There is evidence that this fibrosis is reversible through treatment.This study was to investigate the relationship between the expression of TGF-?1 and Smad4 and the phosphorylation of Smad7 in the TGF-?1/Smads signal transduction pathway,and to clarify the regulation of Qianliexiao Decoction on the differentiation of PrF cells in EAP mice.,provideexperimental basis for follow-up research.Methods: Animal Modeling: A mouse model of autoimmune prostatitis(EAP)with damp-heat syndrome was established using "immunoassay +TCM method".Serum preparation after intervention: Male C57BL/6 mice were intragastrically administered with Qianliexiao Decoction for 7consecutive days.After 1 hour of the last intervention,blood was taken from the eyeballs,collected in non-anticoagulant tubes,centrifuged at3500 r for 10 min,and serum was separated and extracted in a water bath at 56 °C.Inactivated for 30 min,filtered through 0.22 ?m microporous membrane filtration,labeled separately,and stored in a refrigerator at-20 °C for use.Cell experiment: primary cultured EAP mouse prostate fibroblasts were stimulated with purified TGF-?1 concentration of 5 ng/ml medium.After modeling,the PrF in logarithmic growth phase was randomly divided into blank groups(replace fresh).Serum-free medium),normal serogroup,positive control group(medium containing TGF-?1 concentration 5ng/ml),treatment group(high,medium and low dose groups,serum preparation after intervention,serum addition after intervention in each group)Medium)6 replicate wells per group,tested at 37 ° C,5% CO2 incubator for 24 h.The expression levels of TGF-?1 and Smad4 and the phosphorylation level of Smad7 were detected by Western Blot.The proliferation of PrF in each group was detected by CCK8 method,and the apoptosis was detected by flow cytometry.Each group was tested three times in parallel.Results:1.Effect of Qianliexiao Decoction on proliferation of PrF cells stimulated by TGF-?1Compared with the normal cell control group,the proliferation of PrF cells in the TGF-?1 stimulation group was significant.Compared with the normal cell control group,Qianliexiao Decoction had a significant difference between the low dose and the middle dose(P<0.05).There was no significant difference in the high dose group(p>0.05).Compared with the TGF-?1 stimulation group,the proliferation of PrF cells in the serum of Qianliexiao Decoction was significantly inhibited(P<0.05),and the higher the concentration of drug-containing serum,the greater the inhibition.2.Effect of Qianliexiao Decoction on apoptosis of PrF cells stimulated by TGF-?1Compared with the normal cell control group,the apoptotic rate of Qianliexiaotang drug-containing serum was significantly different(P<0.01),and the higher the serum concentration,the higher the apoptotic rate.Effects of Qianliexiao Decoction on TGF-?1/Smads Signal Transduction Pathway in TGF-?1 Stimulated PrF Cells Compared with normal control cells,the expression of Smad4,Samd7 and TGF-?1 protein in TGF-?1-stimulated group was significantly increased(P<0.05).Compared with TGF-?1-stimulated group,it was compared with TGF-?1-stimulated group.The serum containing Tang could significantly down-regulate the expression of Smad4 and TGF-?1protein in PrF cells(P<0.01),and increase the expression level of Samd7(P<0.01).Conclusion: 1.Qianliexiaotang drug-containing serum does not affect the normal growth of PrF.2.TGF-?1 can stimulate the proliferation of PrF,up-regulate theexpression of TGF-?1,Smad4,down-regulate the expression of Smad7,and activate the TGF-?1/Smads pathway.3.Qianliexiaotang can inhibit the proliferation of PrF,which may be related to the decrease of TGF-?1,Smad4 expression,and the increase of Smad7 expression,which blocks the TGF-?1/Smads signaling pathway.