Expression Profiles of Neuropeptides, Neurotransmitters, and Their Receptors in Human Keratocytes In Vitro and In Situ

Abstract

Keratocytes, the quiescent cells of the corneal stroma, play a crucial role in corneal wound healing. Neuropeptides and neurotransmitters are usually associated with neuronal signaling, but have recently been shown to be produced also by non-neuronal cells and to be involved in many cellular processes. The aim of this study was to assess the endogenous intracellular and secreted levels of the neuropeptides substance P (SP) and neurokinin A (NKA), and of the neurotransmitters acetylcholine (ACh), catecholamines (adrenaline, noradrenaline and dopamine), and glutamate, as well as the expression profiles of their receptors, in human primary keratocytes in vitro and in keratocytes of human corneal tissue sections in situ. Cultured keratocytes expressed genes encoding for SP and NKA, and for catecholamine and glutamate synthesizing enzymes, as well as genes for neuropeptide, adrenergic and ACh (muscarinic) receptors. Keratocytes in culture produced SP, NKA, catecholamines, ACh, and glutamate, and expressed neurokinin-1 and -2 receptors (NK-1R and NK-2R), dopamine receptor D2, muscarinic ACh receptors, and NDMAR1 glutamate receptor. Human corneal sections expressed SP, NKA, NK-1R, NK-2R, receptor D2, choline acetyl transferase (ChAT), M3, M4 and M5 muscarinic ACh receptors, glutamate, and NMDAR1, but not catecholamine synthesizing enzyme or the α1 and β2 adrenoreceptors, nor M1 receptor. In addition, expression profiles assumed significant differences between keratocytes from the peripheral cornea as compared to those from the central cornea, as well as differences between keratocytes cultured under various serum concentrations. In conclusion, human keratocytes express an array of neuropeptides and neurotransmitters. The cells furthermore express receptors for neuropeptides/neurotransmitters, which suggests that they are susceptible to stimulation by these substances in the cornea, whether of neuronal or non-neuronal origin. As it has been shown that neuropeptides/neurotransmitters are involved in cell proliferation, migration, and angiogenesis, it is possible that they play a role in corneal wound healing.

Skin matters! The role of keratinocytes in nociception: a rational argument for the development of topical analgesics

“…This has also been explored in a different model and activation of keratinocytes expressing channel rhodopsin via blue light can produce action potentials 12. Keratinocytes express a number of receptors and synthesize neuropeptides, neurotransmitters, and autacoids13 and are also involved in localized inflammatory processes. It is to be acknowledged that these cells may have a key role in the pathogenesis of neuropathic pain 14,15.…”

Antiapoptotic Effect of Acetylcholine in Fas-Induced Apoptosis in Human Keratocytes

“…We think that the reason behind muscarinic receptors being responsible for transducing the ACh antiapoptotic signal is the fact that the muscarinic receptors are abundantly expressed by keratocytes. In our previous study, 38 we showed that cultured keratocytes express M1, M3, M4, and M5 subtypes of muscarinic receptors (lacking only M2). We also observed that the M3 receptor was most probably responsible for transducing the ACh-enhanced proliferation signal.…”

“…38 Donated corneas were scraped with a scalpel in order to remove epithelial and endothelial cells. Next, the central part of the cornea was separated from the peripheral part, and both were minced with a scalpel.…”

“…33 Interestingly, stimulation of both mAChRs and nAChRs has been shown to protect many cell types from apoptosis, 32,34,35 and moreover, it has been shown that ACh can prevent apoptosis of embryonic stem cells and cardiomyocytes through regulation of Bcl-2. 36,37 Therefore, of relevance for apoptosis of keratocytes in the corneal stroma, and its role in corneal wound healing, is the fact that ACh and its receptors have been found in human keratocytes both in situ in corneal tissue sections and in vitro in primary human cell cultures, 38 and that ACh also enhances proliferation of human keratocytes in vitro. 39 In the present study, we hypothesize that ACh reduces Fasinduced apoptosis of keratocytes, through stimulation of either nAChRs or mAChRs and possibly via regulation of initiator caspases as well as regulation of mitochondrial proteins involved in apoptosis.…”

The role of sensory innervation in cornea‐lens regeneration

“…Additionally, Substance P has been implicated in tissue regeneration/repair, having been found in newt forelimb blastemas . Finally, the cutaneous nerves in the cornea may also provide supportive factors for stem cells residing in the limbal stem cell niche …”