Browsing by Subject "Limbal stem cells (LSCs)"

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    07. The Mechanism of NGF Signaling In Innervation-dependent Corneal Epithelial Renewal: New Topical Treatment For Neurotrophic Keratopathy
    (Wolters Kluwer, 2024-04-19) Hussain, Arif; Mirmoeini, Kaveh; Mulenga, Chilando; Crabtree, Jordan; Tajdaran, Kiana; Henrique, Mario; Blum, Noam; Shalom-Feuerstein, Ruby; Borschel, Gregory; Feinberg, Konstantin; Surgery, School of Medicine
    Background: Corneal clarity is essential for vision. Limbal stem cells (LSCs), the source of transparent corneal epithelial cells, are located in the basal epithelium of the limbus at the corneal-conjunctival interface, where they interact with corneal sensory nerves. Besides protection, corneal nerves may stimulate LSC activity. Pathological corneal denervation can lead to ulcers, scarring, and opacification due to impaired healing from repetitive wounds. This condition, termed Neurotrophic Keratopathy (NK), a major cause of corneal blindness, and lacks a definitive cure. Corneal nerves release various trophic factors that regulate epithelial renewal. Nerve growth factor (NGF) has shown positive effects on corneal healing and maintenance in vivo. Topical recombinant human NGF is the only FDA-approved treatment for NK. However, NGF is not efficacious in 30% of cases, requires very frequent dosing, and costs $100k per course. Moreover, NGF’s ability to heal corneal ulcers is limited. Prior studies showed NGF stimulates proliferation and maintenance of cultured human LSCs, which express TrkA and p75NTR receptors, but didn’t establish a link between NGF signaling and corneal sensory innervation-mediated trophic regulation of epithelial renewal. Furthermore, the role and molecular mechanism of NGF signaling in LSC activity remains unidentified. We hypothesize that NGF, locally expressed in its mature and premature (proNGF) forms, regulates LSC activity-dependent homeostatic and wound-induced corneal epithelial renewal via differential activation of its receptors TrkA and p75NTR, a process dependent on corneal sensory innervation. Methods: A) We conducted in vivo experiments in wild-type and mutant mice and rats to elucidate the NGF signaling mechanism in corneal innervation-dependent epithelial renewal. We examined the effect of combinations of TrkA and p75NTR agonists and antagonists on the healing of experimentally wounded corneas, both intact and surgically denervated. B) To understand the role and mechanism of NGF signaling in LSC activity and its relevance to humans, we assessed the clonogenicity of cultured human LSCs (hLSCs) by pharmacologically modulating NGF receptors, as described in (A). Results: A) While inactivation of TrkA completely prevents healing of normally innervated cornea, inactivation of p75NTR combined with a single daily dose of NGF induces complete and rapid healing of denervated de-epithelialized corneas. B) NGF or specific p75NTR inhibitor THX-B supported colonies’ formation by hLSCs that were further augmented by combination of the two compounds. Conclusions: Corneal sensory nerve-associated expression of NGF in its both forms (NGF and proNGF) regulates the proliferation and differentiation of LSCs by differentially stimulating the activity of the NGF receptors. Combined pharmacological activation and inhibition of TrkA and p75NTR, respectively, will be applied in the development of a superior NGF-based treatment of NK.
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    72. The Role of TrkA And P75NTR NGF Receptors in Corneal Wound Healing
    (Wolters Kluwer, 2022) Tajdaran, Kiana; Feinberg, Konstantin; Mirmoeini, Seyed K.; Zhang, Jennifer; Gordon, Tessa; Ali, Asim; Borschel, Gregory; Surgery, School of Medicine
    Purpose: The cornea is the window through which we see the world and is one of the most densely innervated structures in the body. Besides providing protective sensory input, corneal nerves may also stimulate limbal stem cells (LSCs), governing corneal epithelial maintenance and recovery. Loss of corneal innervation, through injury, diabetes, tumors, infections, and even improper contact lens use, leads to neurotrophic keratopathy (NK), a degenerative corneal disease that is characterized by corneal epithelial breakdown, scarring, and permanent vision loss1. The only non-invasive treatment option for NK is human recombinant nerve growth factor (rhNGF), but the short half-life of exogenous neurotrophins-based therapies limits their effecacy2. Development of small molecule ligands for neurotrophin receptors that have more favorable pharmacokinetics and plasma stability showed promising results in the treatment of several neurodegenerative conditions in recent years3. In this study, we investigated the molecular mechanism of NK and the role of the NGF receptors, TrkA and p75NTR, in corneal healing. We hypothesized that TrkA inhibition would delay corneal wound healing and p75NTR inhibition would accelerate corneal healing. Establishing the roles of these receptors may enable novel topical therapeutics for NK. Methods: We used commercially available Ntrk1 mutant mice, whose TrkA receptors are inhibited by a mammalian kinase inhibitor (1-NM-PP1)4. Ntrk1 mice (n=20) were divided into three groups, which received saline injection as a control. In one experimental group animals received TrkA inhibitor and the other group received both TrkA and p75 inhibitor for 5 days. On day six we removed the corneal epithelium with a 0.5 mm rotating brush. To measure epithelial healing, we performed digital imaging of fluorescein staining daily for four days after injury. We then harvested the corneas for immunofluorescent and biochemical analyses. Results: We observed a significant delay in corneal epithelial healing following TrkA inhibition. Further, we observed that topical p75NTR inhibition accelerated corneal wound healing. Conclusion: A selective TrkA agonist or p75NTR inhibition could represent new topical therapeutics for NK.
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    QS8: The Roles of the TrkA and p75NTR NGF Receptors in Corneal Wound Healing
    (Wolters Kluwer, 2021-07-26) Tajdaran, Kiana; Feinberg, Konstantin; Mirmoeini, Seyed Kaveh; Zhang, Jennifer; Gordon, Tessa; Borschel, Gregory; Surgery, School of Medicine
    Purpose: The cornea is the window through which we see the world and is one of the most densely innervated structures in the body. Besides providing protective sensory input, corneal nerves have been postulated to stimulate limbal stem cells (LSCs), hence governing corneal epithelial maintenance and recovery. Loss of corneal innervation, through injury, diabetes, tumors, infections, and even improper contact lens use, leads to neurotrophic keratopathy (NK), a degenerative corneal disease that is characterized by corneal epithelial breakdown, scarring, and permanent vision loss1. The only non-invasive treatment option for NK is the human recombinant nerve growth factor (rhNGF), but the short half-life of exogenous neurotrophins-based therapies make this therapeutic approach less effective2. Development of the small molecule ligands for neurotrophins receptors that have better pharmacokinetics and plasma stability showed promising results in the treatment of several neurodegenerative conditions in the recent years3. In this study, we were prompted to investigate the molecular mechanism of NK and the role of NGF receptors, TrkA and p75NTR, in corneal healing. We hypothesized that TrkA inhibition would delay corneal wound healing and p75NTR inhibition accelerates corneal healing. This knowledge will lay the basis for a new non-invasive approach for NK. Methods: For this experiment, we took advantage of commercially available Ntrk1 mutant mice, which allow for pharmacological inhibition of TrkA receptor with an inhibitor known as not mammalian kinase inhibitor (1-NM-PP1)4. Ntrk1 mice (n=20) were divided into three groups, which received saline injection as a control. In one experimental group animals were received TrkA inhibitor and the other group received both TrkA and p75 inhibitor for 5 days. On day six we removed the corneal epithelium with a 0.5 mm rotating brush. To measure epithelial healing, we performed digital imaging of fluorescein staining daily for four days after injury. We then harvested the corneas for immunofluorescent and biochemical analyses. Results: Our results show a significant delay in corneal epithelial healing following TrkA inhibition and acceleration in corneal healing after p75NTR inhibition. Conclusion: A selective TrkA agonist or p75NTR inhibitors could be a new therapeutic approach for NK.