Abstract
Purpose :
During postnatal development, mammalian neurons of the central nervous system (CNS) lose their ability to elongate projections. Extrinsic and intrinsic factors, both stimulatory and inhibitory, regulate this process. Krüppel-like factors have emerged as important intrinsic factors involved in the control of axonal growth. Our previous work showed that KLF13 is an intrinsic inhibitor of axonal regeneration. The action mechanisms underlaying this effect include KLF13-dependent inhibition of signaling pathways involved in axonal growth and regeneration, including the JAK/STAT pathway, which is the canonical mediator of growth hormone (GH) signaling. In addition to its primary role as regulator of growth, GH is known for its neurotrophic actions in the CNS.
Methods :
To test the hypothesis that depletion of KLF13 would potentiate the regenerative effects of GH, we established two experimental paradigms in retinal ganglion cells (RGC) of mice: 1) AAV-mediated ectopic expression in RGCs, and 2) peripheral GH injection. Initially, we packaged the mouse Gh coding sequence into AAV viral particles of the PHP.eB seroptype. The viral vectors were then injected intravitreally into the eyes of adult WT mice and, twenty-one days later, we confirmed by Western Blot that AAV-mGH causes ectopic expression of GH in mouse retinas.
Results :
Retinal flat mount immunohistochemistry (IHC) for GH showed that that the viral vector transduced RGCs and other retinal cells with a 35% of efficency. On the other hand, we performed subcutaneous injections of bovine GH (2 mg/kg) in both WT and Klf13-/- mice and analyzed GH-mediated signaling pathway activation in the retina, this lead to a 35% increase on RGC survival. Our results showed that JAK/STAT and PI3/AKT signaling pathways were activated by GH treatment, as evidenced by STAT3/STAT5 and AKT phosphorylation, respectively. We also found that GH-mediated phosphorylation of STAT3 in retinas of Klf13-/- mice was higher compared to WT.
Conclusions :
Our results suggest that, by targeting the KLF13, the GH-dependent axon regeneration may be potentiated.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.