Extended Data Fig. 5: Indole-3-proprionate-dependent axonal regeneration.
From: The gut metabolite indole-3 propionate promotes nerve regeneration and repair
a. Schematic of the experiment presented in b and c. Mice were gavaged with 10 mg/kg IPA daily for 10 days followed by 3 days of regeneration time post sciatic nerve crush. b. Micrographs of representative longitudinal sections of sciatic nerves immunostained with SCG-10. Scale bar: 1000 µm. The dashed line indicates the crush site. c. Quantification of the percentage of fibers past the crush site at the indicated distances and normalised to the number of fibers at the crush site. revealed no difference in regeneration potential when treated with 10 mg/kg/day. (N = 4 biologically independent animals per group with a bilateral sciatic nerve crush, two-way-ANOVA with Sidak multiple comparisons test, data are presented as mean values +/ − SEM). d. Quantification of IPA concentration in serum at 20 min, 1 h, 6 h and 12 h post gavage of 20 mg/kg IPA. e. Quantification of IPA concentration in naïve DRG tissue at 20 min, 1 h, 6 h and 12 h post gavage of 20 mg/kg IPA. f. Schematic of the experiment presented in g and h. Mice received daily intraperitoneal injections of 20 mg/kg IPA for 10 days, followed by 3 days of regeneration time post sciatic nerve crush. g. Micrographs of representative longitudinal sections of sciatic nerves immunostained with SCG-10. Scale bar: 1000 µm. The dashed line indicates the crush site. h. Quantification of the percentage of fibers past the crush site at the indicated distances and normalised to the number of fibers at the crush site in nerves of mice injected intraperitoneally with 20 mg/kg/day IPA (N = 4 biologically independent animals per group, ****P < 0.0001, two-way-ANOVA with Sidak test, data are presented as mean values +/ − SEM). i. DRG neurons were treated with 10 μM, 100 μM and 1000 μM IPA. Representative images of DRG neurons immunostained with ßIII-Tub. Scale bar: 50 µm. j. Quantification of neurite outgrowth by measuring the length of neurites (N = 4 independent experiments per group; one-way ANOVA, Dunnett’s multiple comparisons test, examined over two experiments, data are presented as mean values +/ − SEM). k+m. Micrographs of representative sections of DRG slices 72h post dextran injection to the nerve co-immunostained for neuronal markers NF200 and IB4. Scale bar: 200 µm. l+n. Quantification of the percentage of double positive NF200+ and IB4+ neurons in IPA and PBS post-injury treated sciatic DRG (N = 5 biologically independent animals per group, Student’s unpaired t-test, data are presented as mean values +/ − SEM).
