Trimethylamine N-Oxide dysregulates the expression of tight junctions through Highly Upregulated Liver Carcinoma (HULC) in cellular model of colorectal cancer.

Backgrounds and Aim: Colorectal cancer (CRC) pathogenesis is correlated with dysregulation of tight junction. This study aimed to investigate the molecular mechanism by which trimethylamine N-oxide (TMAO) alters the expression of tight junction proteins in a colorectal cancer (CRC) cell line. Material and Method: The study utilized the CRISPR/Cas13 system for targeted knock down of HULC in Caco-2 cells, followed by treatment with trimethylamine N-Oxide (TMAO). Tight junction components, including ZO-1, Claudin-1, and Occludin, were analyzed using real-time quantitative polymerase chain reaction (RT-qPCR). To investigate the role of the P38MAPK pathway, the specific inhibitor SB203580 was used in cells treated with TMAO to comprehensively assess tight junction regulation. Statistical analysis was performed using one-way ANOVA to compare the mean ± SD between different groups, followed by paired comparisons using the t-test. Results: Cells treated with TMAO showed a significant upregulation of the oncogenic long non-coding RNA (lncRNA) HULC (Highly Upregulated in Liver Cancer), , accompanied by increased expression of p38 MAPK. Interestingly, a significant downregulation of ZO-1 and Claudin-1 was observed as a result of TMAO treatment, which was modulated by the HULC/p38 MAPK axis. However, Occludin expression was also reduced by TMAO, but it remained unaffected by the HULC/p38 MAPK pathway. Conclusion: This study revealed a novel TMAO/HULC/p38 MAPK axis involved in the regulation of tight junctions in a colorectal cancer cell line model. TMAO treatment significantly reduced the expression of ZO-1, Claudin-1, and Occludin. Further in vivo research is strongly recommended to clarify the impact of TMAO on the integrity of colorectal cancer cells.