健脾养正消癥方调控USP51抑制低黏附性胃癌进展的机制OA

Objective:To investigate the mechanisms by which Jianpi Yangzheng Xiaozheng prescription(JPYZXZ)treats poorly cohesive gastric carcinoma(PC-GC)through regulation of ubiquitin-specific peptidase 51(USP51).Methods:In vitro experiments:Cell viability and proliferation of PC-GC cell lines(MKN-45 and HGC-27)treated with different concentrations of JPYZXZ(2,4,6 g·L-1)were assessed using Cell Counting Kit-8(CCK-8)and colony formation assays.Cell migration was evaluated by wound healing(scratch)and Transwell assays.The mRNA and protein expression levels of USP51,zinc finger E-box-binding homeobox 1(ZEB1),and epithelial-mesenchymal transition(EMT)-related markers(e.g.,E-cadherin)were detected by quantitative real-time PCR(Real-time PCR)and Western blot,respectively.Subsequently,stable MKN-45 and HGC-27 cell lines with USP51 knockdown(sh-USP51)and overexpression(oe-USP51)were constructed.Their migration ability and EMT-related protein expression were further evaluated by scratch assay,Transwell assay,and Western blot.Invivo experiments:A subcutaneous xenograft model of MKN-45 human gastric cancer was established in BALB/c nude mice.Thirty mice were randomly divided into six groups(NC,NC+JPYZXZ,sh-USP51,sh-USP51+JPYZXZ,oe-USP51,and oe-USP51+JPYZXZ),with five mice in each group.After successful modeling,mice in the treatment groups were administered JPYZXZ(30 g·kg-1)by gavage for 28 days.Body weight and tumor volume were monitored during the experiment.The expression levels of USP51 and EMT-related proteins in tumor tissues were detected by Western blot and immunohistochemistry(IHC).Results:Compared with the blank group,the colony formation rate,wound healing rate,and number of migrated cells in MKN-45 and HGC-27 cells were significantly reduced in all JPYZXZ groups and the 5-fluorouracil(5-FU)group(P＜0.05).The mRNA and protein expression levels of USP51 were decreased(P＜0.05).The expression of ZEB1 and mesenchymal phenotype proteins(e.g.,N-cadherin and vimentin)was reduced(P＜0.05),whereas the expression of the epithelial marker E-cadherin was increased(P＜0.05).Compared with the control group,USP51 expression was decreased in the sh-USP51 group and increased in the oe-USP51 group(P＜0.05).Compared with the NC group,USP51 knockdown significantly reduced the migration and proliferation of gastric cancer cells(P＜0.01),decreased the expression of ZEB1 and EMT-related proteins,and increased E-cadherin expression(P＜0.05).In vivo results showed that JPYZXZ significantly inhibited the growth of xenograft tumors in nude mice(P＜0.05)and markedly reversed the abnormal expression of EMT-related proteins in tumor tissues(P＜0.05).Conclusion:The therapeutic mechanisms of JPYZXZ in PC-GC may be associated with inhibition of the EMT process via regulation of the USP51-ZEB1 signaling pathway.