Combined hyperosmolarity and inflammatory conditions in stressed human corneal epithelial cells and macrophages to evaluate osmoprotective agents as potential DED treatments

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López Cano, José Javier and Gonzalez-Cela Casamayor, Miriam and Andrés Guerrero, Vanessa and Herrero Vanrell, Rocío and Benítez del Castillo, José Manuel and Molina Martínez, Irene Teresa (2021) Combined hyperosmolarity and inflammatory conditions in stressed human corneal epithelial cells and macrophages to evaluate osmoprotective agents as potential DED treatments. Experimental Eye Research, 211 . p. 108723. ISSN 0014-4835

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Official URL: https://doi.org/10.1016/j.exer.2021.108723




Abstract

Purpose: To develop an easy-to-perform combined model in human corneal epithelial cells (HCECs) and Balb/c mice macrophages J774.A1 (MP) for preliminary screening of potential ophthalmic therapeutic substances. Methods: HCECs were exposed to different osmolarities (350–500 mOsm/L) and MTT assay was employed for cell survival and flow cytometry to assess apoptosis-necrosis and relative cell size (RCS) distribution. Effectiveness of Betaine, L-Carnitine, Taurine at different concentrations (ranging from 20 mM to 200 mM) was studied. Also, mucoadhesive polymers such as Hyaluronic acid (HA) and Hydroxypropylmethylcellulose (HPMC) (0.4 and 0.8%) were evaluated. Cells were pre-incubated with the compounds (8h) and then exposed to hyperosmotic stress (470 mOsm/L) for 16h. Moreover, anti-inflammatory activity was performed in LPS-stimulated MP. Results: Exposure to hyperosmotic solutions between 450 and 500 mOsm/L promoted the highest cell death after 16h exposures (p < 0.0001) with a drop in viability to 34.96% ± 11.77 for 470 mOsm/L. Pre-incubation with Betaine at 150 mM and 200 mM provided the highest cell survival against hyperosmolarity (66.01% ± 3.65 and 65.90% ± 0.78 respectively) while HA 0.4% was the most effective polymer in preventing cell death (42.2% ± 3.60). Flow cytometry showed that Betaine and Taurine at concentrations between 150-200 mM and 20–80 mM respectively presented the highest anti-apoptotic activity. Also, HA and HPMC polymers reduced apoptoticinduced cell death. All osmoprotectants modified RCS, and polymers increased their value over 100%. LCarnitine 50 mM, Taurine 40 mM and HA 0.4% presented the highest TNF-α inhibition activity (60%) albeit all of them showed anti-inflammatory inhibition percentages higher than 20%.
Conclusions: HCECs hyperosmolar model combined with inflammatory conditions in macrophages allows the screening of osmoprotectants by simulating chronic hyperosmolarity (16h) and inflammation (24h).


Item Type:Article
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CRUE-CSIC (Acuerdos Transformativos 2021)

Uncontrolled Keywords:Hyperosmolarity; Osmoprotectants; DED; Inflammation; Screening; Osmoprotection model
Subjects:Medical sciences > Pharmacy > Pharmaceutical technology
ID Code:70414
Deposited On:14 Feb 2022 09:46
Last Modified:21 Feb 2022 15:03

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