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      Novel Artificial Tears Containing Cross-Linked Hyaluronic Acid: An In Vitro Re-Epithelialization Study

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          Abstract

          Dry eye syndrome is a common disease which can damage the corneal epithelium. It is treated with eye drops to stimulate tear production and hydrate the corneal surface. The most prescribed artificial tear remedies contain hyaluronic acid (HA), which enhances epithelial wound healing, improving tissue health. To the best of our knowledge, only a few recent studies have investigated cross-linked HA (HA-CL) in eye drops for human applications. This work consists in an in vitro evaluation of the re-epithelialization ability of two different preparations containing a recently synthetized HA cross-linked with urea: 0.02% ( w/ v) HA-CL (solution 1, S1), and 0.4% ( w/ v) HA-CL (solution 2, S2). The study was conducted on both 2D human corneal cells (HCEpiC) and 3D reconstructed tissues of human corneal epithelium (HCE). Viability by 3(4,5-dimethylthiazol-2)2,5-diphenyltetrazolium bromide (MTT) test, pro-inflammatory cytokine release (interleukin-8, IL-8) by ELISA, and morphology by hematoxylin and eosin (HE) staining were evaluated. In addition, to understand the molecular basis of the re-epithelialization properties, cyclin D1 levels were assessed by western blot. The results showed no cellular toxicity, a slight decrease in IL-8 release, and restoration of epithelium integrity when the wounded 3D model was treated with S1 and S2. In parallel, cyclin D1 levels increased in cells treated with both S1 and S2.

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          Sodium hyaluronate (hyaluronic acid) promotes migration of human corneal epithelial cells in vitro.

          Lydia Powell, Harminder Dua, R Amankwah … (2004)
          Sodium hyaluronate (hyaluronic acid) is known to promote corneal epithelial wound healing in vivo and in vitro, in animal experiments. Sodium hyaluronate is the ligand for CD44, a cell surface adhesion molecule which has been found on normal human corneal epithelial cells. The purpose of this study was to investigate the effect of sodium hyaluronate on human corneal epithelial cell migration, proliferation, and CD44 receptor expression. Human corneal epithelial cell cultures were established from 32 donor corneoscleral rims and maintained separately in three different culture conditions: (1) culture medium only, (2) sodium hyaluronate enriched (0.6 mg/ml) medium, and (3) hydroxypropylmethylcellulose enriched (2.5 mg/ml) medium. The total area of migrating epithelial cell sheets in each case was measured by planimetry on days 4, 8, 12, and 16. Cytospin preparations of cells cultured in the different culture conditions were examined immunohistochemically for proliferation and CD44 receptor expression using antibodies directed against Ki67 and CD44 respectively. Cells cultured in the presence of sodium hyaluronate showed significantly increased migration at days 12 and 16 (Friedmen test: p = 0.0012, day 16; p = <0.001, day 12) compared with cells cultured in the other media. There was no difference in cell proliferation (Ki67) or CD44 expression on cells cultured in the different culture conditions. Sodium hyaluronate promotes migration but not proliferation or CD44 expression on human corneal epithelial cells in vitro. The beneficial effect of sodium hyaluronate in corneal wound healing is likely to be related to rapid migration of cells leading to rapid wound closure. This may be facilitated by the adhesion between CD44 on the cells and hyaluronic acid, which coats the surface of the denuded cornea.
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            Hyaluronan: Preparation, Structure, Properties, and Applications.

            Lubomír Lapcík L Jr and, Lubomír Lapčík, Stefaan De Smedt … (1998)
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              Effectiveness of sodium hyaluronate eyedrops in the treatment of dry eye.

              Michael E. Johnson, Paul Murphy, Mike Boulton (2006)
              Dry eye is a common condition, affecting approximately 10-20% of the adult population. Artificial tears are often effective in relieving symptoms in mild and moderate dry eye by replenishing deficient tear volume. Sodium hyaluronate has been proposed as a component in artificial tears, due to its viscoelastic rheology. This paper reports on a study carried out to assess the efficacy of two recently developed eyedrops containing 0.1% and 0.3% sodium hyaluronate (SH) in the treatment of moderate dry eye. Thirteen subjects were recruited with moderate dry eye. Forty microlitres of 0.1% SH, 0.3% SH, or 0.9% saline were instilled in both eyes, and the subjects' symptom intensity and non-invasive break-up time (NIBUT) were measured at 5, 15, 30, 45, and 60 min, and then hourly, until 6 h after drop instillation. This was repeated twice following an interval of 7(+/-1) days, but with a different treatment so that at the end of the final visit each subject had trialled all products. Drop allocation was randomized and double-masked. Both symptoms and NIBUT improved with all treatments. These changes were of a larger magnitude and longer duration with the SH containing eyedrops than with saline. SH of 0.3% tended to perform better than 0.1% SH and achieved statistical significance (P=0.04) for NIBUT when considered over the whole 6-h study period. Sodium hyaluronate of 0.1% and 0.3% reduces symptoms of ocular irritation and lengthens NIBUT in subjects with moderate dry eye more effectively than saline, in terms of peak effect and duration of action.
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                Author and article information

                Journal
                Journal ID (nlm-ta): Molecules
                Journal ID (iso-abbrev): Molecules
                Journal ID (publisher-id): molecules
                Title: Molecules : A Journal of Synthetic Chemistry and Natural Product Chemistry
                Publisher: MDPI
                ISSN (Electronic): 1420-3049
                Publication date (Electronic): 30 November 2017
                Publication date Collection: December 2017
                Volume: 22
                Issue: 12
                Electronic Location Identifier: 2104
                Affiliations
                [1 ]Department of Life Sciences and Biotechnology, Master Course in Cosmetic Science and Technology (COSMAST), University of Ferrara, Via L. Borsari 46, 44121 Ferrara, Italy; arianna.fallacara@ 123456unife.it (A.F.); mv9@ 123456unife.it (S.M.)
                [2 ]Ophthalmology Unit, Azienda ULSS n.22, 37012 Bussolengo, Italy; g.panozzo@ 123456iol.it
                [3 ]Department of Animal Sciences, Plants for Human Health Institute, North Carolina State University, NC Research Campus, 600 Laureate Way, Kannapolis, NC 28081, USA; apecore@ 123456ncsu.edu (A.P.); valacc@ 123456ncsu.edu (G.V.)
                Author notes
                [* ]Correspondence: vrs@ 123456unife.it ; Tel.: +39-0532-455294; Fax: +39-0532-455378
                Author information
                https://orcid.org/0000-0002-2434-4727
                https://orcid.org/0000-0003-1348-4422
                Article
                Publisher ID: molecules-22-02104
                DOI: 10.3390/molecules22122104
                PMC ID: 6149675
                PubMed ID: 29189737
                SO-VID: 48a508ab-2f83-49db-a4a0-e7a5a0d4d835
                Copyright © © 2017 by the authors.
                License:

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

                History
                Date received : 01 November 2017
                Date accepted : 27 November 2017
                Categories
                Subject: Article

                Keywords: anti-inflammatory,artificial tears,corneal epithelium,cyclin d1,dry eye syndrome,ha,ha-cl,il-8,re-epithelialization
                Data availability:
                Keywords: anti-inflammatory, artificial tears, corneal epithelium, cyclin d1, dry eye syndrome, ha, ha-cl, il-8, re-epithelialization

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