Host-specific effects of Eubacterium species on Rg3-mediated modulation of osteosarcopenia in a genetically diverse mouse population

Abstract

Background: Osteosarcopenia, characterized by the simultaneous loss of bone and muscle mass, is a serious health problem in the aging population. This study investigated the interplay between host genetics, gut microbiota, and musculoskeletal health in a mouse model of osteosarcopenia, exploring the therapeutic potential of gut microbiota modulation.

Methods: We examined the effects of Rg3, a phytochemical, on osteosarcopenia and its interactions with host genetics and gut microbiota in six founder strains of the Collaborative Cross (CC) population. Subsequently, we evaluated the therapeutic potential of Eubacterium nodatum (EN) and Eubacterium ventriosum (EV), two gut microbes identified as significant correlates of Rg3-mediated osteosarcopenia improvement, in selected C57BL/6 J (B6) and 129S1/SvImJ (129S1) mouse strains.

Results: Rg3 treatment altered gut microbiota composition aligned with osteosarcopenia phenotypes, which response varied depending on host genetics. This finding enabled the identification of two microbes in the Eubacterium genus, potential mediator of Rg3 effect on osteosarcopenia. Oral administration of EN and EV differentially impacted bone density, muscle mass, exercise performance, and related gene expression in a mouse strain-specific manner. In 129S1 mice, EN and EV significantly improved these parameters, effectively reversing osteosarcopenic phenotypes. Mechanistic investigations revealed that these effects were mediated through the modulation of osteoblast differentiation and protein degradation pathways. In contrast, EN and EV did not significantly improve osteosarcopenic phenotypes in B6 mice, although they did modulate mitochondrial biogenesis and microbial diversity.

Conclusions: Our findings underscore the complex interplay between host genetics and the gut microbiota in osteosarcopenia and emphasize the need for personalized treatment strategies. EN and EV exhibit strain-specific therapeutic effects, suggesting that tailoring microbial interventions to individual genetic backgrounds may be crucial for optimizing treatment outcomes. Video Abstract.

Fig. 1

Experimental design for two independent studies. A Experimental design for the collaborative founder strain study. B Experimental design for the validation study using C57BL/6 J and 129S1/SvImJ strains

Fig. 2

Rg3 treatment alters fecal microbiota composition in six Collaborative Cross (CC) founder strains. A Relative abundance of gut microbiota at the phylum level following Rg3 treatment in the six CC founder strains. B Strain-by-treatment interaction boxplots for three alpha diversity metrics (Shannon diversity, Faith’s PD, and observed ASV) following Rg3 treatment. C Comparison of alpha diversity metrics across the six founder strains with and without Rg3 treatment. Asterisks indicate statistically significant differences (*p < 0.05; **p < 0.01) as determined using Wilcoxon test. D Principal coordinate analysis (PCoA) plots depicting beta diversity of the gut microbiota in the six founder strains with and without Rg3 treatment. Symbols represent individual samples, and colors denote strain identity. E Proportion of differentially abundant taxa identified by Rg3 treatment within each strain. A/J (AJ); C57BL/6 J (B6); 129S1/SvImJ (129); NOD/ShiLtJ (NOD); NZO/HILtJ (NZO); and PWK/PhJ (PWK)

Fig. 3

Abundance of EN and EV and their association with osteosarcopenic phenotypes in CC founder strains. A, B Association between osteosarcopenic traits and variance in Bray–Curtis distance of fecal (A) and cecal (B) microbiota composition in six CC founder strains. Significant associations (p-values) are included in the bar graph. C, D Differentially abundant genera between control and Rg3 treatment for fecal (C) and cecal (D) microbiota in six CC founder strains. Colors represent the strain and sample information. Asterisks indicate statistically significant differences (*p < 0.05; **p < 0.01) determined using ANCOM2.1 and Wilcoxon test. E, F Effect of Rg3 on the relative abundance of EN and EV from all mice (E) and individual strains (F). G, H Spearman correlation plots between grip strength and bacterial abundance of (G) EN in the feces and (H) EV in the cecum by sample treatment. A/J (AJ); C57BL/6 J (B6); 129S1/SvImJ (129); NOD/ShiLtJ (NOD); NZO/HILtJ (NZO); and PWK/PhJ (PWK)

Fig. 4

Effect of EN and EV on muscle atrophy markers in DEX-treated B6 and 129S1 mice. A Representative hematoxylin and eosin (H&E) staining of the gastrocnemius muscle and (B) quantification of the average muscle fiber cross-sectional area. C Grip strength normalized to body weight (D–F) and muscle weight relative to body weight for gastrocnemius (D), tibialis (E), and soleus (F) muscles. G, H Effect of EN and EV on mRNA expression of protein degradation markers (Fbxo32 and Trim63) in DEX-treated B6 and 129S1 mice. Data are represented as mean ± SEM. Statistical significance was determined using one-way ANOVA followed by Tukey’s post hoc test. *p < 0.05, **p < 0.01, ns (not significant). C57BL/6 J (B6); 129S1/SvImJ (129S1)

Fig. 5

Effect of EN and EV on bone microarchitecture markers in DEX-treated B6 and 129S1 mice. Microarchitecture was assessed using (A) micro-computed tomography in femur, B bone mineral density (BMD), C bone volume to total volume (BV/TV), D serum osteocalcin, E trabecular thickness (Tb.Th), F trabecular number (Tb.N), G and H mRNA expression of genes associated with osteoblast differentiation. Data are represented as the mean ± standard error of the mean (SEM). Statistical significance was determined using one-way ANOVA followed by Tukey’s post hoc test. Asterisks indicate significance levels: *p < 0.05, **p < 0.01, ***p < 0.001, ns (not significant). C57BL/6 J (B6); 129S1/SvImJ (129S1)

Fig. 6

Effect of EN and EV on endurance-related markers and gut microbial diversity in DEX-treated mice. A Distance to exhaustion, B, C mRNA expression of (B) Ppargc1a, and (C) Tfam. D, E Relative abundance of bacterial taxa at the (D) phylum and (E) class levels is displayed in stack bar plot. F–H Alpha and beta diversity metrics are presented. F Observed generate amplicon sequence variants (ASVs), G Faith’s PD, and (H) PCoA plot depicting beta diversity (Bray–Curtis) are shown. I–K Abundance of specific bacterial genera is presented for (I) Romboutsia, J Rikenella, and (K) Lachnoclostridium. L–N Spearman correlation coefficients between the abundance of each bacteria genus and time to exhaustion are presented. Data are represented as the mean ± standard error of the mean (SEM). Statistical significance was determined by one-way ANOVA followed by Tukey’s post hoc test. Asterisks indicate significance levels: *p < 0.05, **p < 0.01, ***p < 0.001, ns (not significant). C57BL/6 J (B6); 129S1/SvImJ (129S1)