Post-Weaning Microbiota Colonization in Germ-Free Mice Compensates for Impaired Skeletal Muscle Maturation but Not for Peripheral Nerve Fiber Abnormalities

Summary: Previous work demonstrated that the absence of a complex gut microbiota (GM) impairs postnatal development of peripheral nerves and motor targets in germ-free (GF) mice. In this study, germ free (GF) mice were colonized with a diverse gut microbiota (GM) at weaning (EX-GF) to evaluate whether this intervention could reverse the observed phenotypic alterations. Although colonization partially restored microbial diversity, it failed to rescue molecular and structural deviations in peripheral nerves. Axon hypermyelination was exacerbated, and alterations in gene expression in both nerves and dorsal root ganglia (DRG), along with elongated nodes of Ranvier, persisted. In contrast, muscle defects were fully reversed: skeletal muscle atrophy was alleviated, muscle mass regulatory pathways were reactivated, proteomic profiles normalized, and postsynaptic neuromuscular junction (NMJ) fragmentation was reduced. However, presynaptic NMJ deviations remained. Targeted metabolomic analysis revealed normalization of serum indole-3-propionate levels following GM colonization, suggesting a role in muscle recovery. These findings underscore the differential sensitivity of neuromuscular system components to the gut microbiota and point to a critical developmental window during which microbiota-derived signals are essential for proper neuromuscular maturation and long-term maintenance.Overall design: Fecal pellets from 63-76-day-old adult C57BL6 JZtm CGM mice (Complex Gut Microbiota; n = 4) and EX-GF mice (generated by co-housing GF with CGM from weaning onward; n = 5) were collected, stored at -80 C and sent to Novogene for DNA isolation and shotgun metagenomics for taxonomical profiling of the gut microbiota.Contributors: Sonia Calabro, Davide Pellegrino, Chiara Cicconetti, Svenja Kankowski, Sajjad Farzin, Francesca Bertone, Mira Fruhwein, Francesca Anselmi, Dario Bizzotto, Marijana Basic, Silvia Bolsega, Matthias Steglich, Lutz Wiehlmann, Cecilia Gelfi, Daniele Capitanio, Karin Kleigrewe, Salvatore Oliviero, Giovanna Gambarotta, Kirsten Haastert-Talini, Matilde Cescon, Giulia Ronchi