Fig. 2

Microbiota governs BMMSC lineage commitment. a Oil Red O staining. Left panel: BMMSCs from GF mice show less Oil Red O-positive adipocytes after adipogenic induction in vitro compared to BMMSCs derived from SPF (middle panel) and ConvD (right panel) mice (bar = 100 μm). b Western blot analysis shows that SPF-derived and ConvD-derived BMMSCs express higher adipogenic key transcription factor PPAR-γ and adipogenic marker LPL compared to germ-free BMMSCs (left and middle). Real-time PCR shows PPAR-γ and LPL mRNA highly expressed in germ-free derived BMMSCs (right). c Alizarin Red staining shows GF-derived BMMSCs formed more mineralized nodule in vitro compared to SPF and ConvD groups. d Western blot analysis shows that GF BMMSCs expressed higher OCN and Runx2 after osteogenic induction in vitro (left). Quantification of Runx2 western blot intensity (right). e In-vivo bone regeneration capacity of BMMSCs. Left panels of micro-CT and H&E staining show the initial bone defect size in the rat mandibles. Second panels of micro-CT and H&E staining show the bone defect size after GF BMMSC treatment. Third panels of micro-CT and H&E staining show bone defect size after SPF BMMSC treatment. Right panels show the bone defect size after ConvD BMMSC treatment (bar = 200 μm). All experimental data verified in at least three independent experiments. Error bars represent the SEM from the mean values. **P < 0.001; *P < 0.05. ConvD conventionalized, GF germ free, SPF specific pathogen free, N.S. not significant, PPAR peroxisome proliferator-activated receptor, LPL lipoprotein lipase, Runx2 runt-related transcription factor 2, OCN osteocalcin