| ?Background and aims? Chemotherapy is one of the main treatments for advanced gastric cancer,but the general effect is not ideal because of the prevalence of chemotherapy drug resistance.Cancer stem cell(CSC)is a major source of chemotherapy resistance.Hedgehog(Hh)signaling pathway is linked with cancer stem cells maintain and chemoresistance in various kinds of cancers.ABCG2 is also very important in cancer chemoresistance.The purpose of this study is to research the possible mechanism of chemotherapy resistance and propose the new Gli1-ABCG2 signaling axis.It may help to explain the mechanism of chemoresistance from a new point and provide new ideas and new targets for effective prevention and treatment of gastric cancer.?Methods? Firstly,the IC50 values of CDDP and 5Fu in gastric cancer cells N87 and AGS were determine by AlamarBlue,and the cell induced model was established according to the IC50.Then,the expression levels of Hedgehog signaling pathway target genes Gli1,Gli2 and PTCH1 were detected by QRT-PCR.The IC50 and cancer stem cell properties were determined followed by Gli1 knockdown through lentivirus or Gli1 overexpression through retrovirus.Cancer stem cell properties included sphere formation by serum free suspension culture of gastric cancer cells,the percentage of side populatin(SP)cells and CD90 positive cells by flow cytometry.ABCG2,the downstream target gene of Gli1,was predicted by bioinformatics and verified by ChIP assay.The effects of ABCG2 on the IC50 values and cancer stem cell properties were investigated by inhibiting ABCG2 function or overexpressing ABCG2.The role of Gli1-ABCG2 signaling axis in chemoresistance of gastric cancer was validated in NSG mice.Finally,the expression of ABCG2 in 180 cases of gastric cancer was explored by IHC,and the relationship between the expression level of ABCG2 and clinicopathological parameters or survival was analyzed.?Results? In this study,we found that the Hedgehog signaling pathway target genes Gli1,Gli2 and PTCH1 were activated in gastric cancer cells N87 and AGS induced by CDDP or 5Fu for 48 h.Further,IC50 values of chemotherapeutic drugs and cancer stem cell properties were decreased after Gli1 knockdown,including the proportion of SP cells was reduced,the spheres became small and formation efficiency was decreased,stem cell surface molecule CD90 expression level was also dropped.The IC50 and cancer stem cell properties were increased after Gli1 overexpression.After screening,it was found that the expression level of ABCG2 was consistent with that of Gli1,and the direct regulation of transcription factor Gli1 on ABCG2 was confirmed by ChIP assay.ABCG2 also can affect the drug sensitivity of gastric cancer cells and cancer stem cell properties through inhibition of ABCG2 function or overexpression of ABCG2.At the same time,the role of Gli1-ABCG2 signaling axis in chemotherapy resistance of gastric cancer was also verified in NSG mice.The expression level of ABCG2 was related to the differentiation and survival of patients with gastric cancer.?Conclusions? Hedgehog signaling pathway was activated in gastric cancer cells induced by chemotherapy drugs in vitro.The target gene of Hh,Gli1 can promote the drug resistance in gastric cancer by regulating cancer stem cell properties.The expression level of ABCG2 was consistent with that of Gli1,and the direct regulation of transcription factor Gli1 on ABCG2 was confirmed by ChIP assay.ABCG2,as a function execution gene of Gli1,mediated the cancer stem cell properties and lead to chemotherapy resistance in gastric cancer.The role of Gli1-ABCG2 signaling axis in chemotherapy resistance was also identified in NSG mice.The expression level of ABCG2 in gastric cancer was related to the differentiation and prognosis of gastric cancer. |
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