Topical application of calcitonin gene-related peptide as a regenerative, antifibrotic, and immunomodulatory therapy for corneal injury

Abstract

Calcitonin gene-related peptide (CGRP) is a multifunctional neuropeptide abundantly expressed by corneal nerves. Using a murine model of corneal mechanical injury, we found CGRP levels in the cornea significantly reduced after injury. Topical application of CGRP as an eye drop accelerates corneal epithelial wound closure, reduces corneal opacification, and prevents corneal edema after injury in vivo. CGRP promotes corneal epithelial cell migration, proliferation, and the secretion of laminin. It reduces TGF-β1 signaling and prevents TGF-β1-mediated stromal fibroblast activation and tissue fibrosis. CGRP preserves corneal endothelial cell density, morphology, and pump function, thus reducing corneal edema. Lastly, CGRP reduces neutrophil infiltration, macrophage maturation, and the production of inflammatory cytokines in the cornea. Taken together, our results show that corneal nerve-derived CGRP plays a cytoprotective, pro-regenerative, anti-fibrotic, and anti-inflammatory role in corneal wound healing. In addition, our results highlight the critical role of sensory nerves in ocular surface homeostasis and injury repair.

Calcitonin Gene-Related Peptide Ameliorates Experimental Dry Eye Disease

“… 14 , 37 Additionally, topical CGRP treatment improved clinical markers of DED, such as CFS and TBUT. These observations are consistent with previous studies demonstrating the protective role of CGRP in promoting epithelial integrity in the cornea after mechanical injury 20 and enhancing bronchial epithelium healing in vitro . 38 While we found no significant effect of CGRP on tear secretion, our current dry eye model of using environmentally induced desiccating stress does not cause long-term tear volume reduction.…”

“…This is consistent with earlier findings from non-DED model systems demonstrating anti-apoptotic and pro-proliferative properties of CGRP on different epithelial cell types including corneal, bronchial, and alveolar epithelium. 20 , 38 , 51 Our study extends this by suggesting that CGRP promotes corneal epithelial health by suppressing apoptosis and enhancing proliferation, potentially through its anti-inflammatory effects, a relationship not fully explored in the literature. Corneal epithelial cell damage is well established in DED, 52 , 53 and our study supports this notion and that CGRP reduced cell death in the corneal epithelium in DED.…”

“…Our results demonstrate the anti-inflammatory properties of CGRP, substantiated by reduced immune cell infiltration into the cornea and suppression of pro-inflammatory cytokines (TNF-α, IL-1β) during DED, consistent with previous studies showing anti-inflammatory activity of CGRP in other model systems. 20 , 32 Importantly, our study revealed that anti-inflammatory effects of CGRP were localized to the cornea, with no significant changes in immune responses in the conjunctiva. This may be attributed to the conjunctiva's rich vasculature.…”

Calcitonin Gene-Related Peptide Ameliorates Experimental Dry Eye Disease

“… 14 , 37 Additionally, topical CGRP treatment improved clinical markers of DED, such as CFS and TBUT. These observations are consistent with previous studies demonstrating the protective role of CGRP in promoting epithelial integrity in the cornea after mechanical injury 20 and enhancing bronchial epithelium healing in vitro . 38 While we found no significant effect of CGRP on tear secretion, our current dry eye model of using environmentally induced desiccating stress does not cause long-term tear volume reduction.…”

“…This is consistent with earlier findings from non-DED model systems demonstrating anti-apoptotic and pro-proliferative properties of CGRP on different epithelial cell types including corneal, bronchial, and alveolar epithelium. 20 , 38 , 51 Our study extends this by suggesting that CGRP promotes corneal epithelial health by suppressing apoptosis and enhancing proliferation, potentially through its anti-inflammatory effects, a relationship not fully explored in the literature. Corneal epithelial cell damage is well established in DED, 52 , 53 and our study supports this notion and that CGRP reduced cell death in the corneal epithelium in DED.…”

“…Our results demonstrate the anti-inflammatory properties of CGRP, substantiated by reduced immune cell infiltration into the cornea and suppression of pro-inflammatory cytokines (TNF-α, IL-1β) during DED, consistent with previous studies showing anti-inflammatory activity of CGRP in other model systems. 20 , 32 Importantly, our study revealed that anti-inflammatory effects of CGRP were localized to the cornea, with no significant changes in immune responses in the conjunctiva. This may be attributed to the conjunctiva's rich vasculature.…”

Clusterin reverses epitheliopathy, reduces inflammation, and restores goblet cells and corneal nerves in a mouse model of autoimmune dry eye

“…A reduced proportion of CGRP-positive corneal nerves in Thbs1 -deficient mice is consistent with the reported wound-healing and anti-inflammatory properties of this neuropeptide 17,51 . To determine if CLU-driven repair of corneal epithelial damage also helps restore CGRP-positive corneal nerves, we performed further quantification of immunoreactive CGRP in beta-3-tubulin-stained corneal nerves.…”

Protective Effect of Calcitonin Gene-Related Peptide on Corneal Epithelial Barrier in Experimental Dry Eye Models

“…Previous studies have demonstrated that CGRP alleviates ocular surface inflammation and promotes corneal wound healing. 13 , 14 Our study further reveals its direct role in maintaining epithelial barrier integrity by preserving the localization of tight junction proteins, confirming that CGRP's cross-organ barrier protective function 6 – 9 is preserved in the cornea. However, it should be noted that CGRP is not a unidirectional protective factor.…”

Calcitonin Gene-Related Peptide Ameliorates Experimental Dry Eye Disease

“… 14 , 37 Additionally, topical CGRP treatment improved clinical markers of DED, such as CFS and TBUT. These observations are consistent with previous studies demonstrating the protective role of CGRP in promoting epithelial integrity in the cornea after mechanical injury 20 and enhancing bronchial epithelium healing in vitro . 38 While we found no significant effect of CGRP on tear secretion, our current dry eye model of using environmentally induced desiccating stress does not cause long-term tear volume reduction.…”

“…This is consistent with earlier findings from non-DED model systems demonstrating anti-apoptotic and pro-proliferative properties of CGRP on different epithelial cell types including corneal, bronchial, and alveolar epithelium. 20 , 38 , 51 Our study extends this by suggesting that CGRP promotes corneal epithelial health by suppressing apoptosis and enhancing proliferation, potentially through its anti-inflammatory effects, a relationship not fully explored in the literature. Corneal epithelial cell damage is well established in DED, 52 , 53 and our study supports this notion and that CGRP reduced cell death in the corneal epithelium in DED.…”

“…Our results demonstrate the anti-inflammatory properties of CGRP, substantiated by reduced immune cell infiltration into the cornea and suppression of pro-inflammatory cytokines (TNF-α, IL-1β) during DED, consistent with previous studies showing anti-inflammatory activity of CGRP in other model systems. 20 , 32 Importantly, our study revealed that anti-inflammatory effects of CGRP were localized to the cornea, with no significant changes in immune responses in the conjunctiva. This may be attributed to the conjunctiva's rich vasculature.…”

Clusterin reverses epitheliopathy, reduces inflammation, and restores goblet cells and corneal nerves in a mouse model of autoimmune dry eye

“…A reduced proportion of CGRP-positive corneal nerves in Thbs1 -deficient mice is consistent with the reported wound-healing and anti-inflammatory properties of this neuropeptide 17,51 . To determine if CLU-driven repair of corneal epithelial damage also helps restore CGRP-positive corneal nerves, we performed further quantification of immunoreactive CGRP in beta-3-tubulin-stained corneal nerves.…”

Protective Effect of Calcitonin Gene-Related Peptide on Corneal Epithelial Barrier in Experimental Dry Eye Models

“…Previous studies have demonstrated that CGRP alleviates ocular surface inflammation and promotes corneal wound healing. 13 , 14 Our study further reveals its direct role in maintaining epithelial barrier integrity by preserving the localization of tight junction proteins, confirming that CGRP's cross-organ barrier protective function 6 – 9 is preserved in the cornea. However, it should be noted that CGRP is not a unidirectional protective factor.…”

Calcitonin Gene-Related Peptide Ameliorates Experimental Dry Eye Disease

“… 14 , 37 Additionally, topical CGRP treatment improved clinical markers of DED, such as CFS and TBUT. These observations are consistent with previous studies demonstrating the protective role of CGRP in promoting epithelial integrity in the cornea after mechanical injury 20 and enhancing bronchial epithelium healing in vitro . 38 While we found no significant effect of CGRP on tear secretion, our current dry eye model of using environmentally induced desiccating stress does not cause long-term tear volume reduction.…”

“…This is consistent with earlier findings from non-DED model systems demonstrating anti-apoptotic and pro-proliferative properties of CGRP on different epithelial cell types including corneal, bronchial, and alveolar epithelium. 20 , 38 , 51 Our study extends this by suggesting that CGRP promotes corneal epithelial health by suppressing apoptosis and enhancing proliferation, potentially through its anti-inflammatory effects, a relationship not fully explored in the literature. Corneal epithelial cell damage is well established in DED, 52 , 53 and our study supports this notion and that CGRP reduced cell death in the corneal epithelium in DED.…”

“…Our results demonstrate the anti-inflammatory properties of CGRP, substantiated by reduced immune cell infiltration into the cornea and suppression of pro-inflammatory cytokines (TNF-α, IL-1β) during DED, consistent with previous studies showing anti-inflammatory activity of CGRP in other model systems. 20 , 32 Importantly, our study revealed that anti-inflammatory effects of CGRP were localized to the cornea, with no significant changes in immune responses in the conjunctiva. This may be attributed to the conjunctiva's rich vasculature.…”

Clusterin reverses epitheliopathy, reduces inflammation, and restores goblet cells and corneal nerves in a mouse model of autoimmune dry eye

“…A reduced proportion of CGRP-positive corneal nerves in Thbs1 -deficient mice is consistent with the reported wound-healing and anti-inflammatory properties of this neuropeptide 17,51 . To determine if CLU-driven repair of corneal epithelial damage also helps restore CGRP-positive corneal nerves, we performed further quantification of immunoreactive CGRP in beta-3-tubulin-stained corneal nerves.…”

Protective Effect of Calcitonin Gene-Related Peptide on Corneal Epithelial Barrier in Experimental Dry Eye Models

“…Previous studies have demonstrated that CGRP alleviates ocular surface inflammation and promotes corneal wound healing. 13 , 14 Our study further reveals its direct role in maintaining epithelial barrier integrity by preserving the localization of tight junction proteins, confirming that CGRP's cross-organ barrier protective function 6 – 9 is preserved in the cornea. However, it should be noted that CGRP is not a unidirectional protective factor.…”