Functional and Morphologic Alterations in Mechanical, Polymodal, and Cold Sensory Nerve Fibers of the Cornea Following Photorefractive Keratectomy

Bech, Federico; González-González, Omar; Artime, Enol; Serrano, Joana; Alcalde, Ignacio; Gallar, Juana; Merayo‐Lloves, Jesús; Belmonte, Carlos

doi:10.1167/iovs.18-24007

Cited by 30 publications

(39 citation statements)

References 54 publications

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“…The pattern of regenerative growth shown by identified TRPM8(ϩ) thermoreceptor fibers after surgical axotomy is essentially the same as that recently reported for corneal fibers of all sensory modalities after excimer laser ablation of the central cornea (Bech et al, 2018). This suggests that TG neurons of different sensory modalities share the general cellular mechanisms governing the anatomical distribution plan of regenerating axons after injury.…”

Section: Discussionsupporting

confidence: 83%

“…Regenerating cold thermoreceptors displayed an enhanced ongoing activity 3 weeks after mechanical stromal parent axon's axotomy, a procedure that preserved only a few intact stromal branches in the hinge. This result contrasts with the low background activity in the majority of corneal cold terminals after a superficial excimer laser corneal lesion, where only the epithelium nerve branches and a fraction of the superficial stromal nerve trunks were destroyed (Bech et al, 2018). In this case, many parent axons maintained part of their complex branching tree intact, and only 20% of the recorded, regenerating cold nerve terminals exhibited abnormal background firing and lower threshold (i.e., higher sensitivity) to cooling stimuli, possibly corresponding to more severely injured axons.…”

Section: Discussioncontrasting

confidence: 67%

“…The remainder of the cold-sensitive neurons, named low-background high-threshold cold thermoreceptors (LB-HTCs), exhibit a low background impulse activity at 34°C and require large temperature reductions to increase their firing rate. It has been proposed that HB-LTCs tonically modulate basal tearing and blinking rates, whereas LB-HTCs are mainly implicated in evoking sensations associated to corneal dryness (Parra et al, 2010;Hirata et al, 2012;Quallo et al, 2015;Bech et al, 2018).…”

Section: Introductionmentioning

confidence: 99%

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Role of TRPM8 Channels in Altered Cold Sensitivity of Corneal Primary Sensory Neurons Induced by Axonal Damage

et al. 2019

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The cornea is extensively innervated by trigeminal ganglion cold thermoreceptor neurons expressing TRPM8 (transient receptor potential cation channel subfamily M member 8). These neurons respond to cooling, hyperosmolarity and wetness of the corneal surface. Surgical injury of corneal nerve fibers alters tear production and often causes dry eye sensation. The contribution of TRPM8-expressing corneal cold-sensitive neurons (CCSNs) to these symptoms is unclear. Using extracellular recording of CCSNs nerve terminals combined with in vivo confocal tracking of reinnervation, Ca 2ϩ imaging and patch-clamp recordings of fluorescent retrogradely labeled corneal neurons in culture, we analyzed the functional modifications of CCSNs induced by peripheral axonal damage in male mice. After injury, the percentage of CCSNs, the cold-and menthol-evoked intracellular [Ca 2ϩ ] rises and the TRPM8 current density in CCSNs were larger than in sham animals, with no differences in the brake K ϩ current I KD. Active and passive membrane properties of CCSNs from both groups were alike and corresponded mainly to those of canonical low-and high-threshold cold thermoreceptor neurons. Ongoing firing activity and menthol sensitivity were higher in CCSN terminals of injured mice, an observation accounted for by mathematical modeling. These functional changes developed in parallel with a partial reinnervation of the cornea by TRPM8(ϩ) fibers and with an increase in basal tearing in injured animals compared with sham mice. Our results unveil key TRPM8-dependent functional changes in CCSNs in response to injury, suggesting that increased tearing rate and ocular dryness sensation derived from deep surgical ablation of corneal nerves are due to enhanced functional expression of TRPM8 channels in these injured trigeminal primary sensory neurons.

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Section: Discussionsupporting

confidence: 83%

Section: Discussioncontrasting

confidence: 67%

Section: Introductionmentioning

confidence: 99%

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Role of TRPM8 Channels in Altered Cold Sensitivity of Corneal Primary Sensory Neurons Induced by Axonal Damage

et al. 2019

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“…Surgical injury was performed using an Allegretto WaveLight excimer laser (PR-020407, Wavelight GmbH, Alcon, Erlangen, Germany) as previously described. [26][27][28][29][30] Right eyes were subjected to PRK surgery with a 2.0-mm ablation zone on the central cornea and a depth of 45 μm. The procedure included the simultaneous ablation of the epithelium (using a defined epithelial thickness profile of 25 μm centrally) and 20 μm of the anterior stroma (about 20% of the total stromal thickness) in a single step.…”

Section: Prk Mouse Modelmentioning

confidence: 99%

Development and Optimization of Freeze-Dried Eye Drops Derived From Plasma Rich in Growth Factors Technology

Anitua

Fuente

Alcalde

et al. 2020

Trans. Vis. Sci. Tech.

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To investigate whether plasma rich in growth factors (PRGF) eye drops maintain their biological potential after a freeze drying process. The addition of a lyoprotectant like trehalose was also evaluated. Methods: Blood from three healthy donors was collected to obtain eye drops by PRGF technology. The resultant eye drops were divided in four groups: PRGF, freeze-dried PRGF (PRGF lyo), and PRGF lyophilized mixed with 2,5% trehalose (PRGF lyo+2.5T) or 5% trehalose (PRGF lyof+5T). Chemical and biological characteristics were evaluated. Photorefractive keratectomy was performed on C57BL/6 mice which were divided in three treatment groups: control, PRGF, and PRGF lyo. Corneal wound healing and haze formation were evaluated macroscopically. Eyes were collected at 1, 2, 3, and 7 days after surgery, and were processed for histologic studies. Results: The pH values of PRGF samples increased significantly after the lyophilization process. Osmolarity levels increased significantly in PRGF samples mixed with trehalose in comparison with PRGF samples without protectants. The freeze drying process maintained growth factors levels as well as the biological properties of PRGF eye drops even without the use of lyoprotectants. PRGF lyo treatment significantly decreased the re-epithelialization time and haze formation in photorefractive keratectomy-treated corneas regarding PRGF and control groups. Furthermore, the PRGF lyo group significantly decreased the number of smooth muscle actin-positive cells in comparison with the control group at each time of the study and at days 2 and 3 in the PRGF group. Conclusions: The freeze drying process preserves the protein and growth factor content as well as the biological properties of PRGF eye drops, even without the use of protectants. Freeze-dried PRGF eye drops accelerate corneal tissue regeneration after photorefractive keratectomy in comparison with the control group. Translational Relevance: Our study shows the feasibility to preserve the biological capability of PRGF eye drops as freeze-dried formulation, avoiding the addition of protectants. Resumen (Foreign Language Abstract): Objetivo: Investigar si el colirio de plasma rico en factores de crecimiento (PRGF) mantiene su potencial biológico después de un proceso de liofilización. También se evaluó la adición de un lioprotector como la trehalosa. Métodos: Se recogió sangre de tres donantes sanos para obtener colirio mediante la tecnología PRGF. El colirio resultante se dividió en cuatro grupos: PRGF, PRGF liofilizado (PRGF lyo) y PRGF liofilizado mezclado con trehalosa al 2,5% (PRGF lyo + 2.5T) o trehalosa al 5% (PRGF lyof + 5T). Se evaluaron las características químicas y biológicas de cada uno de los colirios. Se realizó una queratectomía fotorrefractiva (PRK) en ratones C57BL/6 que se dividieron en tres grupos de tratamiento: Control, PRGF y PRGF lyo. La curación de la herida corneal y la formación de "haze"se evaluaron macroscópicamente.

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“…As a result, in recent times, there has been a proliferation of research, especially using mouse animal models of corneal injury and nerve regeneration (e.g. Bech et al, 2018;Harris et al, 2018;Ozaki et al, 2018;Stepp et al, 2018;Zhang et al, 2018). Several of these studies have shown an interplay between immune cells and corneal nerves (e.g.…”

Section: Introductionmentioning

confidence: 99%

Impact of topical anti-fibrotics on corneal nerve regeneration in vivo

Hindman

DeMagistris

Callan

et al. 2019

Experimental Eye Research

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Recent work in vitro has shown that fibroblasts and myofibroblasts have opposing effects on neurite outgrowth by peripheral sensory neurons. Here, we tested a prediction from this work that dampening the fibrotic response in the early phases of corneal wound healing in vivo could enhance reinnervation after a large, deep corneal injury such as that induced by photorefractive keratectomy (PRK). Since topical steroids and Mitomycin C (MMC) are often used clinically for mitigating corneal inflammation and scarring after PRK, they were ideal to test this prediction. Twenty adult cats underwent bilateral, myopic PRK over a 6mm optical zone followed by either: (1) intraoperative MMC (n=12 eyes), (2) intraoperative prednisolone acetate (PA) followed by twice daily topical application for 14 days (n=12 eyes), or (3) no post-operative treatment (n=16 eyes). Anti-fibrotic effects of MMC and PA were verified optically and histologically. First, optical coherence tomography (OCT) performed pre-operatively and 2, 4 and 12 weeks post-PRK was used to assess changes in corneal backscatter reflectivity. Post-mortem immunohistochemistry was then performed at 2, 4 and 12 weeks post-PRK, using antibodies against α-smooth muscle actin (α-SMA). Finally, immunohistochemistry with antibodies against βIII-tubulin (Tuj-1) was performed in the same corneas to quantify changes in nerve distribution relative to unoperated, control cat corneas. Two weeks after PRK, untreated corneas exhibited the greatest amount of staining for α-SMA, followed by PA-treated and MMC-treated eyes. This was matched by higher OCT-based stromal reflectivity values in untreated, than PA-and MMC-treated eyes. PA treatment appeared to slow epithelial healing and although normal epithelial thickness was restored by 12 weeks-post-PRK, intra-epithelial nerve length only reached ~1/6 normal values in PA-treated eyes. Even peripheral cornea (outside the ablation zone) exhibited depressed intra-epithelial nerve densities after PA treatment. Stromal nerves were abundant under the α-SMA zone, but appeared to largely avoid it, creating an area of sub-epithelial stroma devoid of nerve trunks. In turn, this may have led to the lack of sub-basal and intra-epithelial nerves in the ablation zone of PA-treated eyes 4 weeks after PRK, and their continuing paucity 12 weeks after PRK. Intra-operative MMC,

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Functional and Morphologic Alterations in Mechanical, Polymodal, and Cold Sensory Nerve Fibers of the Cornea Following Photorefractive Keratectomy

Cited by 30 publications

References 54 publications

Role of TRPM8 Channels in Altered Cold Sensitivity of Corneal Primary Sensory Neurons Induced by Axonal Damage

Role of TRPM8 Channels in Altered Cold Sensitivity of Corneal Primary Sensory Neurons Induced by Axonal Damage

Development and Optimization of Freeze-Dried Eye Drops Derived From Plasma Rich in Growth Factors Technology

Impact of topical anti-fibrotics on corneal nerve regeneration in vivo

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