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      Synergistic Effect of Artificial Tears Containing Epigallocatechin Gallate and Hyaluronic Acid for the Treatment of Rabbits with Dry Eye Syndrome

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          Abstract

          Dry eye syndrome (DES) is a common eye disease. Artificial tears (AT) are used to treat DES, but they are not effective. In this study, we assessed the anti-inflammatory effect of AT containing epigallocatechin gallate (EGCG) and hyaluronic acid (HA) on DES. Human corneal epithelial cells (HCECs) were used in the WST-8 assay to determine the safe dose of EGCG. Lipopolysaccharide-stimulated HCECs showing inflammation were treated with EGCG/HA. The expression of IL-1ß, IL-6, IL-8, and TNF-α was assessed by real-time PCR and AT physical properties such as the viscosity, osmolarity, and pH were examined. AT containing EGCG and HA were topically administered in a rabbit DES model established by treatment with 0.1% benzalkonium chloride (BAC). Tear secretion was assessed and fluorescein, H&E, and TUNEL staining were performed. Inflammatory cytokine levels in the corneas were also examined. The non-toxic optimal concentration of EGCG used for the treatment of HCECs in vitro was 10 μg/mL. The expression of several inflammatory genes, including IL-1ß, IL-6, IL-8, and TNF-α, was significantly inhibited in inflamed HCECs treated with 10 μg/mL EGCG and 0.1% (w/v) HA (E10/HA) compared to that in inflamed HCECs treated with either EGCG or HA alone. AT containing E10/HA mimic human tears, with similar osmolarity and viscosity and a neutral pH. Fluorescence examination of the ocular surface of mouse eyes showed that HA increased drug retention on the ocular surface. Topical treatment of DES rabbits with AT plus E10/HA increased tear secretion, reduced corneal epithelial damage, and maintained the epithelial layers and stromal structure. Moreover, the corneas of the E10/HA-treated rabbits showed fewer apoptotic cells, lower inflammation, and decreased IL-6, IL-8, and TNF-α levels. In conclusion, we showed that AT plus E10/HA had anti-inflammatory and mucoadhesive properties when used as topical eye drops and were effective for treating DES in rabbits.

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          Tea polyphenols for health promotion.

          People have been consuming brewed tea from the leaves of the Camellia sinensis plant for almost 50 centuries. Although health benefits have been attributed to tea, especially green tea consumption since the beginning of its history, scientific investigations of this beverage and its constituents have been underway for less than three decades. Currently, tea, in the form of green or black tea, next to water, is the most widely consumed beverage in the world. In vitro and animal studies provide strong evidence that polyphenols derived from tea may possess the bioactivity to affect the pathogenesis of several chronic diseases. Among all tea polyphenols, epigallocatechin-3-gallate has been shown to be responsible for much of the health promoting ability of green tea. Tea and tea preparations have been shown to inhibit tumorigenesis in a variety of animal models of carcinogenesis. However, with increasing interest in the health promoting properties of tea and a significant rise in scientific investigation, this review covers recent findings on the medicinal properties and health benefits of tea with special reference to cancer and cardiovascular diseases.
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            Experimental dry eye stimulates production of inflammatory cytokines and MMP-9 and activates MAPK signaling pathways on the ocular surface.

            To evaluate whether experimentally induced dry eye in mice activates mitogen-activated protein kinase (MAPK) signaling pathways, c-Jun N-terminal kinases (JNK), extracellular-regulated kinases (ERK), and p38 and stimulates ocular surface inflammation. 129SvEv/CD-1 mixed mice aged 6 to 8 weeks were treated with systemic scopolamine and exposure to an air draft for different lengths of time, from 4 hours to 10 days. Untreated mice were used as the control. The concentrations of IL-1beta and TNF-alpha in tear fluid washings and in corneal and conjunctival epithelia were measured by ELISA. MMP-9 in tear washings was evaluated by zymography, and gelatinase activity in the cornea and conjunctiva was determined by in situ zymography. Corneal and conjunctival epithelia were lysed in RIPA buffer for Western blot with MAPK antibodies, or they were lysed in 4 M guanidium thiocyanate solution for extraction of total RNA, which was used to determine gene expression by semiquantitative RT-PCR, real-time PCR, and gene array. Compared with those in age-matched control subjects, the concentrations of IL-1beta and MMP-9 in tear fluid washings and the concentrations of IL-1beta and TNF-alpha and gelatinolytic activity in the corneal and conjunctival epithelia were significantly increased in mice receiving treatments to induce dry eye after 5 or 10 days. The expression of IL-1beta, TNF-alpha, and MMP-9 mRNA by the corneal and conjunctival epithelia was also stimulated in mice treated for 5 or 10 days. The levels of phosphorylated JNK1/2, ERK1/2, and p38 MAPKs in the corneal and conjunctival epithelia were markedly increased as early as 4 hours after treatment, and they remained elevated up to 5 days. Experimental dry eye stimulates expression and production of IL-1beta, TNF-alpha, and MMP-9 and activates MAPK signaling pathways on the ocular surface. MAPKs are known to stimulate the production of inflammatory cytokines and MMPs, and they could play an important role in the induction of these factors that have been implicated in the pathogenesis of dry eye disease.
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              Polymeric nanoparticulate system: a potential approach for ocular drug delivery.

              Various efforts in ocular drug delivery have been made to improve the bioavailability and to prolong the residence time of drugs applied topically onto the eye. The potential use of polymeric nanoparticles as drug carriers has led to the development of many different colloidal delivery vehicles. Drug loaded polymeric nanoparticles (DNPs) offer several favorable biological properties, such as biodegradability, nontoxicity, biocompatibility and mucoadhesiveness. These submicron particles are better than conventional ophthalmic dosage forms to enhance bioavailability without blurring the vision. DNPs have been shown to be amenable to targeting of the drug to the site of action, leading to a decrease in the dose required and a decrease in side effects. Additionally, DNPs can be fabricated by simple techniques with better physical stability than liposomes. This unique combination of properties makes DNPs a novel polymeric drug delivery device, which fulfils the requirements for ophthalmic application. This review discusses the polymeric nanoparticles, physiochemical characterization, fabrication techniques, therapeutic significances, patented technology of nanoparticles and future possibility in the field of ocular drug delivery.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                23 June 2016
                2016
                : 11
                : 6
                : e0157982
                Affiliations
                [1 ]Graduate Institute of Biomedical Materials and Tissue Engineering, College of Biomedical Engineering, Taipei Medical University, No. 250, Wu-Hsing Street, Taipei City 110, Taiwan
                [2 ]Department of Chemical Engineering and Biotechnology, National Taipei University of Technology, No.1, Section 3, Chung-Hsiao E. Road, Taipei City 106, Taiwan
                [3 ]Institute of Biomedical Engineering and Nanomedicine Research, National Health Research Institutes, Taiwan, No. 35, Keyan Rd., Zhunan Town, Miaoli County 350, Taiwan
                [4 ]Department of Ophthalmology, Taipei Veterans General Hospital, No. 201, Sec. 2, Shipai Rd., Beitou District, Taipei City 112, Taiwan
                [5 ]Department of Ophthalmology, School of Medicine, College of Medicine, Taipei Medical University, No. 250, Wu-Hsing Street, Taipei City 110, Taiwan
                Cedars-Sinai Medical Center; UCLA School of Medicine, UNITED STATES
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Conceived and designed the experiments: C-LT H-WF K-HC. Performed the experiments: Y-JH Z-YC C-LT. Analyzed the data: Y-JH Z-YC C-LT. Contributed reagents/materials/analysis tools: H-WF K-HC C-LT. Wrote the paper: C-LT.

                Author information
                http://orcid.org/0000-0001-9446-4030
                Article
                PONE-D-15-40478
                10.1371/journal.pone.0157982
                4919017
                27336157
                7b1ce62c-b946-42d2-8e20-e7a982780691
                © 2016 Tseng et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 2 October 2015
                : 8 June 2016
                Page count
                Figures: 9, Tables: 1, Pages: 18
                Funding
                Funded by: funder-id http://dx.doi.org/10.13039/501100004663, Ministry of Science and Technology, Taiwan;
                Award ID: NSC 102-2221-E-038 -007
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/501100004737, National Health Research Institutes;
                Award ID: NHRI-EX103- 10334EI
                Award Recipient :
                Funded by: Southern Taiwan Science Park
                Award ID: CZ-01-1-01-105
                Award Recipient :
                This work was supported by grants from Ministry of Science and Technology (MOST), Taiwan under grant number at MOST 102-2221-E-038 -007; and an integrated research grant in health and medical sciences from National Health Research Institute (NHRI), Taiwan at grant number of NHRI-EX103-10334EI awarded to CL Tseng. An Innovation grant of Southern Taiwan Science Park at grant number of CZ-01-01-01-105 was awarded to HW Fang.
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