A high-fiber diet synergizes with Prevotella copri and exacerbates rheumatoid arthritis

Abstract

Both preclinical and established rheumatoid arthritis (RA) patients display alterations in the gut microbiome. Prevotella spp. are preferentially enriched in a subset of RA patients. Here, we isolated a Prevotella strain, P. copri RA, from the feces of RA patients and showed that colonization of P. copri RA exacerbated arthritis in a collagen-induced arthritis (CIA) model. With the presence of P. copri RA colonization, a high-fiber diet exacerbated arthritis via microbial alterations and intestinal inflammation. Colonization of P. copri together with a high-fiber diet enabled the digestion of complex fiber, which led to the overproduction of organic acids, including fumarate, succinate and short-chain fatty acids. Succinate promoted proinflammatory responses in macrophages, and supplementation with succinate exacerbated arthritis in the CIA model. Our findings highlight the importance of dysbiosis when evaluating the effects of dietary interventions on RA pathogenesis and provide new insight into dietary interventions or microbiome modifications to improve RA management.

Keywords: Dysbiosis; High-fiber diet; Microbiota-derived metabolites; Prevotella copri; Rheumatoid arthritis.

Fig. 1

P. copri RA colonization exacerbated arthritis. A Illustration of the colonization experiments with P. copri RA (PC) and B. thetaiotamicron (BT). B Confirmation of P. copri RA and B. thetaiotamicron colonization by agarose gel electrophoresis. gPC indicates P. copri genomic DNA (gPC), and B. thetaiotaomicron genomic DNA (gBT) was used as a positive control; PC, BT, and CON represent total bacterial DNA exacted from the feces of the corresponding group; M, DNA ladder. C Clinical scores of collagen-induced arthritis in the PC (n = 4), BT (n = 4) and CON (n = 3) groups. D Serum anti-bovine type II collagen IgG level. E Representative histology of joints. The black arrow indicates synovial lining-cell hyperplasia, and the blue box indicates stromal cells. F Histological scores of joints. G Levels of fecal IgA. H Serum LPS levels. Each symbol in (D, F, G, and H) represents an animal. Error bars indicate the SEM (C, D, F–H). P values were determined by two-way ANOVA (C), one-way ANOVA (D, F, G, H). *p < 0.05, **p < 0.01, ***p < 0.001, ^#p < 0.05, ^##p < 0.01, ^###p < 0.001

Fig. 2

P. copri RA-induced disease severity was promoted by a high-fiber diet. A Schematic of CIA mice with dietary intervention. B Confirmation of P. copri RA colonization by quantitative PCR. Control means P. copri genomic DNA; PC + FCD means DNA exacted from FCD group feces; PC + NCD means DNA exacted from NCD group feces. C Anti-bovine type II collagen IgG levels in the serum of PC + FCD (n = 4) or PC + NCD (n = 4) mice. D Clinical scores of collagen-induced arthritis. E Body weight of mice in the two groups. F Flow cytometric analysis of CD4⁺Th17⁺ cells in mesenteric lymph nodes. G Histological evaluation of the CIA colon and joint tissues. The scale bar is 100 μm. H The histology scores of the colon tissues. I The histology scores of the synovial tissues. Each symbol in (C, F, H, and I) represents an animal. P values were determined by two-way ANOVA (D, E), the Mann‒Whitney test (F, H, I), and Student’s t test (C). *p < 0.05, **p < 0.01, ****p < 0.0001. Abbreviations: PC P. copri RA, NCD non-fiber-containing diet, FCD fiber-containing diet

Fig. 3

The impact of dietary intervention on the colon transcriptome. A Volcano plots showing the expression of significantly downregulated (blue) and upregulated (red) genes in the FCD and NCD groups. Blue and red dots represent significant DEGs, with the lines denoting a cutoff P value of <0.05. B Heatmap showing the expression of genes involved in the colons of mice subjected to dietary intervention. The red square indicates upregulated genes, and the blue square indicates downregulated genes. For RNA-sequencing data, n = 4 per group. Key genes are highlighted in red. C Gene Ontology (GO) enrichment analysis among the significantly regulated transcripts from the colons of the FCD and NCD groups

Fig. 4

Dietary intervention affected the gut microbiota colonized with P. copri RA. A Shannon diversity index between the FCD and NCD groups. (p value = 0.0198). B Principal coordinate analysis (PCoA) of the microbial composition based on Bray Curtis. (p value = 0.031, permutational multivariate analysis of variance (PERMANOVA) by Adonis). C Taxonomy stack distribution of phyla. D Relative abundance of Akkermansiaceae. E Relative abundance of Bacteroidaceae. F Mucus layer thickness of the colon. Each symbol in (F) represents an animal

Fig. 5

Unique carbohydrate-active enzyme (CAZyme) classes in the P. copri RA strain compared with P. copri 18205. A The distribution of CAZyme classes in P. copri 18205 and P. copri RA. B Presence or absence of glycoside hydrolases (GHs) in P. copri 18205 vs. P. copri RA. Differentially encoded GHs are listed. Abbreviations: CBM carbohydrate binding module, CE carbohydrate esterase, GH glycoside hydrolase, GT glycosyltransferase, PL polysaccharide lyase

Fig. 6

High-fiber diet with P. copri RA colonization induced overproduction of inflammatory microbial metabolites and contributed to RA development. A, B The levels of fumarate and succinate in cecal samples. C–H Cecum SCFA levels. Error bars indicate the SEM (A–H). I The level of IL-1β in LPS-stimulated BMDMs pretreated with the designated acids. J Schematic of the sodium succinate treatment. K Clinical scores of collagen-induced arthritis in the succinate group (n = 4) and control group (n = 4). L Anti-bovine type II collagen IgG levels in serum. M Serum IL-6 levels. N Heatmap showing the expression of genes involved in the colon of mice subjected to sodium succinate treatment and control treatment. Each symbol in (A–H, L, and M) represents each animal. Error bars indicate the SEM. P values were determined by two-way ANOVA (K), and t test (A–I, L, M). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001