| Colorectal cancer(CRC)is one of the malignant tumors with high incidence rate and mortality in the world.In recent years,the incidence rate of colorectal cancer has increased year by year and the proportion of early-onset cases has increased.Research on colorectal cancer related to pathogens has become a hot direction in the current medical field.Safe and effective biological regulation methods have also become an important means of intervention and prevention.More and more studies have confirmed that Fusobacterium nucleatum(Fn)live bacteria and their metabolites are the main triggers for inducing and promoting CRC proliferation and metastasis.However,the mechanism of action between Fusobacterium nucleatum extracellular vesicles(Fnevs)and CRC cells has not been reported yet.This study focuses on CRC cell lines(HT-29,Caco-2)and Fusobacterium nucleatum ATCC25586.By combining electron microscopy,it was determined that Fn secretes extracellular vesicles in the form of "outer membrane vesicles".The extracted and purified Fnevs were characterized,and their active protein components carrying 579 parental bacteria were determined through mass spectrometry analysis;Fluorescence staining and in,co culture techniques confirmed that Fnevs can be recognized and internalized by CRC cells;The results of cell proliferation,migration and invasion,flow cytometryand cell scratch experiments showed that Fnevs promoted the proliferation,migration,and invasion of CRC cells,and inhibited tumor cell apoptosis;And it was found that it can enhance the antioxidant stress capacity and SOD enzyme activity level of CRC cells.The results of transcriptome and metabolomics showed that the differentially expressed genes and metabolites of CRC cells after Fnevs treatment were mostly involved in tumor cell proliferation;Joint analysis found that Fnevs regulates the cancer center carbon metabolism pathway of CRC cells,activates the RAS/ERK signaling pathway,increases the phosphorylation level of C-Myc and the expression of the SLC1A5 gene encoding glutamine transporters on the cell membrane surface,promotes the utilization of glutamine by CRC cells,enhances the TCA cycle and proline biosynthesis.In the exploration of CRC biological prevention and control,the best strain to intervene in Fn growth activity and biofilm formation was screened for six types of Bifidobacterium(Bifidobacterium longum,Bifidobacterium brevis,Bifidobacterium adolescens,Bifidobacterium animalis,Bifidobacterium bifidum bifidum,and Bifidobacterium infantis)during their stable developmental stage metabolites(Cell free fermentation supernatant(CFS)),And the minimum concentration for inhibiting biofilm synthesis was determined to be 12.5%.Through non target metabolomics analysis,it was found that CFS of Bifidobacterium longum contains abundant carboxylic acids,derivatives,and organic heterocyclic compounds.The research results indicate that a 25% concentration of CFS from Bifidobacterium longum can effectively inhibit the proliferation,migration,and invasion of CRC cells induced by Fnevs,and this concentration does not damage normal colon epithelial cells NCM460.Based on the joint analysis of differentially expressed genes and differentially expressed metabolites in CRC cells after intervention,it was found that Bifidobacterium longum CFS effectively controlled the promoting effect of Fnevs on CRC cell transcription and metabolism.This study preliminarily explored the mechanism of the effect of extracellular vesicles of Fusobacterium nucleatum on the proliferation of colorectal cancer cells.It was found that the postbiotic element of Bifidobacterium longum can serve as an effective biological regulatory means to inhibit CRC induced by Fusobacterium nucleatum and extracellular vesicles,laying a theoretical foundation for the feasibility and effectiveness of biological prevention and control of CRC in the future. |
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