MTX-induced gastrointestinal reactions in RA: Prevotella enrichment, gut dysbiosis, and PI3K/Akt/Ras/AMPK pathways.

OBJECTIVES

To investigate the role of gut microbiota in methotrexate (MTX)-induced gastrointestinal reactions (MRGR) in patients with rheumatoid arthritis (RA).

METHODS

As a prospective, single-center, convenience sampling study, stool samples were obtained from 28 RA patients (male: female = 10:18) at Lanzhou University Second Hospital who were undergoing MTX treatment for analysis of their gut microbiota using 16S rRNA gene sequencing. Clinical disease activity (CDAI) and MRGR were assessed after two months of MTX therapy. All data collection periods exceeded one year. Intestinal germ-free mice, generated through antibiotic treatment, received fecal microbiota transplantation (FMT) from the patients, followed by varying doses of MTX to observe MRGR. Intestinal transcriptomics and markers related to intestinal barrier function were subsequently examined.

RESULTS

Females (84.6%) and high disease activity (CDAI scores, 39.6 ± 11.2 vs 26.3 ± 9.2) were prone to have MRGR in RA patients. Patients with MRGR (PT-GR) showed lower gut microbial diversity versus non-MRGR (PT-noGR). Prevotella abundance, positively correlated with CDAI and MRGR (p < 0.05), was elevated in PT-GR. Administering 10 mg/kg MTX to mice caused intestinal damage. FMT-GR-MTX mice exhibited weight loss (95.2%), morphological deterioration (86.4%), and reduced tight junction proteins (Claudin-1:72.4%; ZO-1:81.2%). Transcriptomics linked upregulated Gβγ/CREB/Atp4b to PI3K/Akt/Ras pathways and downregulated PFK2/PP2 to AMPK signaling in MRGR.

CONCLUSION

Our study identified notable gut microbiota alterations in RA patients prone to MRGR, with changes in intestinal gene expression and reduced intestinal barrier function potentially contributing to MRGR. These findings suggest potential strategies to mitigate MRGR in RA patients undergoing MTX treatment. Key Points • The RA-related MRGR is correlated with the intestinal microbiota. • Females, low gut diversity, and Prevotella enrichment are MRGR risks in RA. • Upregulated DEGs in MRGR linked to PI3K/Akt, Ras pathways. • Downregulated DEGs in MRGR focus on the AMPK pathway.