Activation of the BMP4/Smad1 Pathway Promotes Retinal Ganglion Cell Survival and Axon Regeneration

Purpose: We investigate if the BMP4/Smad1 intracellular signaling pathway is neuroprotective and axogenic in adult rodent retinal ganglion cells (RGC) in vivo and in vitro.

Methods: Adult retinal cultures were prepared from intact and after optic nerve crush (ONC) injured rats that have been stimulated to survive and regenerate using an intravitreal peripheral nerve (PN) graft. Laser capture microdissection (LCM) then was used to isolate RGC with and without neurites. Quantitative RT-PCR determined changes in BMP4/Smad1 signaling pathway mRNA. Immunohistochemistry confirmed localization of BMP4 and activation of Smad1 in ONC+PN-stimulated RGC in vivo. BMP4 peptide was used to stimulate RGC survival and neurite/axon regeneration in vitro and in vivo. Finally, the rapamycin sensitivity of the effects was determined in BMP4-stimulated RGC in vitro and in vivo.

Results: In retinal cultures prepared from intact and ONC+PN-stimulated rats, RGC with neurites had upregulated regeneration-related and BMP4/Smad1 signaling pathway mRNA levels, while low levels of these mRNAs were present in RGC isolated without neurites. An optimal dose of 200 ng/mL BMP4 peptide in vitro promoted approximately 30% RGC survival and disinhibited RGC neurite outgrowth, despite the presence of inhibitory CNS myelin extracts. BMP4 also promoted approximately 30% RGC survival in vivo and stimulated significant RGC axon regeneration at 100, 200, and 400 μm beyond the lesion site. Finally, the response of RGC to BMP4 treatment in vitro and in vivo was rapamycin-insensitive.

Conclusions: Activation of the BMP4/Smad1 pathway promotes survival and axon regeneration independent of mTOR and, therefore, may be of therapeutic interest.