PURPOSE: To determine the in vivo expression of neurotrophins (NTs) and nerve regeneration-associated genes (RAGs) after surgically creating a hinged lamellar corneal flap in thy1-YFP mice. METHODS: Lamellar corneal flaps with multiple hinges were created in thy1-YFP mice. Mice were killed at weeks 2, 4, and 8. Quantitative polymerase chain reaction was performed to determine the expression of NTs and RAGs in the corneas after lamellar transection. Nerve growth factor (Ngf), brain-derived neurotrophic factor (Bdnf), glial cell-derived neurotrophic factor (Gdnf), neurotrophin 3, neurotrophin 5, small proline-rich repeat protein 1A (Sprr1a), growth-associated protein 43 (Gap43), and beta III tubulin (Tubb3) gene expressions were analyzed. Whole-mount confocal immunofluorescence and Western analyses were performed for localization and abundance of robustly expressed genes. RESULTS: Sprouts of fine YFP-positive fronds emanating from transected (injured) nerve bundles were seen in the flap area at 2 weeks onward. Bdnf and Sprr1a were robustly and significantly expressed at 2 weeks postoperatively (>2-fold increase in expression; P<0.05). Bdnf localized to thy1-YFP+ cells in operated corneas. Sprr1a localized to corneal epithelial cell membranes. At 8 weeks, none of the NTs and RAGs had increased expression. Bdnf (ρ=0.73, P=0.001) and Sprr1a (ρ=0.76, P=0.001) showed a significant positive correlation with beta III tubulin. CONCLUSIONS: The neurotrophin Bdnf and RAG Sprr1a are robustly and significantly expressed during corneal nerve regeneration in vivo.
PURPOSE: To determine the in vivo expression of neurotrophins (NTs) and nerve regeneration-associated genes (RAGs) after surgically creating a hinged lamellar corneal flap in thy1-YFP mice. METHODS:Lamellar corneal flaps with multiple hinges were created in thy1-YFP mice. Mice were killed at weeks 2, 4, and 8. Quantitative polymerase chain reaction was performed to determine the expression of NTs and RAGs in the corneas after lamellar transection. Nerve growth factor (Ngf), brain-derived neurotrophic factor (Bdnf), glial cell-derived neurotrophic factor (Gdnf), neurotrophin 3, neurotrophin 5, small proline-rich repeat protein 1A (Sprr1a), growth-associated protein 43 (Gap43), and beta III tubulin (Tubb3) gene expressions were analyzed. Whole-mount confocal immunofluorescence and Western analyses were performed for localization and abundance of robustly expressed genes. RESULTS: Sprouts of fine YFP-positive fronds emanating from transected (injured) nerve bundles were seen in the flap area at 2 weeks onward. Bdnf and Sprr1a were robustly and significantly expressed at 2 weeks postoperatively (>2-fold increase in expression; P<0.05). Bdnf localized to thy1-YFP+ cells in operated corneas. Sprr1a localized to corneal epithelial cell membranes. At 8 weeks, none of the NTs and RAGs had increased expression. Bdnf (ρ=0.73, P=0.001) and Sprr1a (ρ=0.76, P=0.001) showed a significant positive correlation with beta III tubulin. CONCLUSIONS: The neurotrophin Bdnf and RAG Sprr1a are robustly and significantly expressed during corneal nerve regeneration in vivo.
Authors: Michael Yamakawa; Samuel M Santosa; Neeraj Chawla; Evguenia Ivakhnitskaia; Matthew Del Pino; Sebastian Giakas; Arnold Nadel; Sneha Bontu; Arjun Tambe; Kai Guo; Kyu-Yeon Han; Maria Soledad Cortina; Charles Yu; Mark I Rosenblatt; Jin-Hong Chang; Dimitri T Azar Journal: Biochim Biophys Acta Gen Subj Date: 2020-03-12 Impact factor: 3.770