Objective:To investigate the effects and possible mechanisms of action of Curcuma wenyujin Y.H.Chen et C.Ling n-Butyl alcohol extract(CWNAE)on repression of human gastric cancer(GC)AGS cell invasion induced by co-cult...Objective:To investigate the effects and possible mechanisms of action of Curcuma wenyujin Y.H.Chen et C.Ling n-Butyl alcohol extract(CWNAE)on repression of human gastric cancer(GC)AGS cell invasion induced by co-culturing with Helicobacter pylori(HP).Methods:AGS cells were cultured with HP of positive or negative cytotoxin-associated gene A(Cag A)and vacuolating cytotoxin gene A(Vac A)expression(Cag A+/-or Vac A+/-)and divided into 5 group.Group A was cultured without HP as a control,Group B with HPCagA+VacA+,Group C with HPCagA-VacA-,Group D with HPCagA+VacA+and CWNAE,and Group E with HPCag A-Vac Aand CWNAE.Methylthiazolyldiphenyl-tetrazolium bromide(MTT)and tumor invasion assays,examinations of morphology and ultramicroscopic structures,quantitative real-time polymerase chain reaction and Western blots were performed to measure the effects and uncover the mechanisms behind these effects of HPCagA+VacA+and CWNAE on the epithelial-mesenchymal transition(EMT)of AGS cells.Results:The 10%inhibitory concentration of CWNAE against AGS cells after a 48 h incubation was 19.73±1.30μg/m L.More AGS cells were elongated after co-culturing with HPCagA+VacA+than after culturing with HPCagA-VacA-.In tumor invasion assays,HPCagA+VacA+significantly enhanced the invasiveness of AGS cells compared to the other experimental groups(all P value<0.05),and this effect was inhibited by CWNAE.Treatment with CWNAE normalized tight junctions and reduced the number of pseudopodia of AGS cells co-cultured with HPCagA+VacA+.HPCagA+VacA+up-regulated zincfinger ebox binding homeobox 1(ZEB1)in AGS cells after co-culturing for 24 h.Expression of caudal type homeobox transcription factor(CDX-2)and claudin-2 was significantly increased by HPCagA+VacA+(P<0.05),but not by HPCagA-VacA-.Conclusions:HPCagA+VacA+promoted the invasiveness of AGS cells through up-regulation of ZEB1 transcription and claudin-2 and CDX-2 expression.CWNAE inhibited these effects of HPCagA+VacA+on AGS cells by down-regulating ZEB1 transcription,and CDX-2 and claudin-2 expression.展开更多
基金Supported by the National Natural Science Foundation of China(No.81202822)Fund of Zhejiang Province Administration of Traditional Chinese Medicine,China(No.2016ZA092/2017ZKL008)Key Laboratory of Pathology and Physiology of Digestive Tract Diseases in Zhejiang Province,China
文摘Objective:To investigate the effects and possible mechanisms of action of Curcuma wenyujin Y.H.Chen et C.Ling n-Butyl alcohol extract(CWNAE)on repression of human gastric cancer(GC)AGS cell invasion induced by co-culturing with Helicobacter pylori(HP).Methods:AGS cells were cultured with HP of positive or negative cytotoxin-associated gene A(Cag A)and vacuolating cytotoxin gene A(Vac A)expression(Cag A+/-or Vac A+/-)and divided into 5 group.Group A was cultured without HP as a control,Group B with HPCagA+VacA+,Group C with HPCagA-VacA-,Group D with HPCagA+VacA+and CWNAE,and Group E with HPCag A-Vac Aand CWNAE.Methylthiazolyldiphenyl-tetrazolium bromide(MTT)and tumor invasion assays,examinations of morphology and ultramicroscopic structures,quantitative real-time polymerase chain reaction and Western blots were performed to measure the effects and uncover the mechanisms behind these effects of HPCagA+VacA+and CWNAE on the epithelial-mesenchymal transition(EMT)of AGS cells.Results:The 10%inhibitory concentration of CWNAE against AGS cells after a 48 h incubation was 19.73±1.30μg/m L.More AGS cells were elongated after co-culturing with HPCagA+VacA+than after culturing with HPCagA-VacA-.In tumor invasion assays,HPCagA+VacA+significantly enhanced the invasiveness of AGS cells compared to the other experimental groups(all P value<0.05),and this effect was inhibited by CWNAE.Treatment with CWNAE normalized tight junctions and reduced the number of pseudopodia of AGS cells co-cultured with HPCagA+VacA+.HPCagA+VacA+up-regulated zincfinger ebox binding homeobox 1(ZEB1)in AGS cells after co-culturing for 24 h.Expression of caudal type homeobox transcription factor(CDX-2)and claudin-2 was significantly increased by HPCagA+VacA+(P<0.05),but not by HPCagA-VacA-.Conclusions:HPCagA+VacA+promoted the invasiveness of AGS cells through up-regulation of ZEB1 transcription and claudin-2 and CDX-2 expression.CWNAE inhibited these effects of HPCagA+VacA+on AGS cells by down-regulating ZEB1 transcription,and CDX-2 and claudin-2 expression.
