[Fusobacterium nucleatum upregulates ABCG2 by activating the E-cadherin/β-catenin signaling pathway to promote oxaliplatin resistance in colorectal cancer].

Abstract

Objective: To investigate whether F.nucleatum affects the chemotherapy resistance of colorectal cancer by regulating ABC subfamily G subtype 2 (ABCG2), in view of the fact that the correlation between the two has not been reported. Methods: Oxaliplatin was used to interfere with colorectal cancer cells and co-cultured with F.nucleatum to establish a chemotherapy-induced model of microbial infection. Calcein AM/PI cell staining, trypan blue staining, and cell counting kit 8 (CCK-8) method were used to detect cell activity. Real-time fluorescence quantitative polymerase chain reaction and western blot were used to detect ABCG2 mRNA and protein expression levels in colorectal cancer cells. The target gene was knocked down by constructing shRNA plasmids. The HCT-116 cell F.nucleatum infection model was constructed and transcriptome sequencing and Kyoto Encyclopedia of Genes and Genomes (KEGG) and gene ontology (GO) analyses were performed to determine the differential gene expression enrichment pathways. Genistein (GST) was used as an E-cadherin blocker, and triclabendazole sulfoxide (TCBZSO) was used as an ABCG2 blocker. Immunofluorescence was used to detect E-cadherin and β-catenin protein expressions and intracellular localization levels. The subcutaneous xenograft model of nude mice was constructed in vivo, and the expression level of ABCG2 protein in tissues was detected by immunohistochemical staining. Results: CCK-8 results showed that F. nucleatum+oxaliplatin group [HT-29, (92.26±1.66)%; HCT-116, (82.13±1.84)%] cell relative survival rate was higher than that of oxaliplatin group [HT-29, (79.64±3.72)%; HCT-116, (67.56±2.96)%; P＜0.001]. The relative survival rate of oxaliplatin and F. nucleatum co-culture group with ABCG2-knockdown HCT-116 cells [(61.44±1.48)%] was lower than that of F. nucleatum and oxaliplatin co-cultured with HCT-116 cells [(69.29±4.45)%, P=0.015]. GO enrichment analysis showed that HCT-116 cells co-cultured with F.nucleatum were significantly enriched in "β-catenin binding", "cadherin binding" and "regulation of Wnt signaling pathway". RT-qPCR results showed that the relative expression of ABCG2 mRNA in F.nucleatum + genistein group was significantly lower than that in F.nucleatum group (P＜0.001). The results in vivo showed that the tumor weight in the F.n+L-OHP+TCBZSO group [(0.12±0.02)g] was lower than that in the F.n+L-OHP group [(0.33±0.05)g, P＜0.001]. Immunohistochemistry suggested that the promotion of ABCG2 protein expression by F. nucleatum was blocked after TCBZSO intervention. Conclusion: F. nucleatum up-regulates ABCG2 expression through E-cadherin/β-catenin signaling pathway to promote colorectal cancer resistance to oxaliplatin.