DOI QR코드

DOI QR Code

Antioxidant potential of silk protein sericin against hydrogen peroxide-induced oxidative stress in skin fibroblasts

  • Dash, Rupesh (Department of Biotechnology, Indian Institute of Technology) ;
  • Acharya, Chitrangada (Department of Biotechnology, Indian Institute of Technology) ;
  • Bindu, P.C. (Department of Biotechnology, Indian Institute of Technology) ;
  • Kundu, S.C. (Department of Biotechnology, Indian Institute of Technology)
  • Received : 2007.02.28
  • Accepted : 2007.09.14
  • Published : 2008.03.31

Abstract

The antioxidant potential of silk protein sericin from the non-mulberry tropical tasar silkworm Antheraea mylitta cocoon has been assessed and compared with that of the mulberry silkworm, Bombyx mori. Skin fibroblast cell line (AH927) challenged with hydrogen peroxide served as the positive control for the experiment. Our results showed that the sericin obtained from tasar cocoons offers protection against oxidative stress and cell viability is restored to that of control on pre-incubation with the sericin. Fibroblasts pre-incubated with non-mulberry sericin had significantly lower levels of catalase; lactate dehydrogenase and malondialdehyde activity when compared to untreated ones. This report indicates that the silk protein sericin from the non-mulberry tropical tasar silkworm, A. mylitta can serve as a valuable antioxidant.

Keywords

References

  1. Harman, D. (1956) Aging: a theory based on free radical and radiation chemistry. J. Gerontol. 11, 298-300 https://doi.org/10.1093/geronj/11.3.298
  2. Datta, K., Babbar, P., Srivastava, T., Sinha, S. and Chattopadhyay, P. (2002) p53 dependent apoptosis in glioma cell lines in response to hydrogen peroxide induced oxidative stress. Int. J. Biochem. Cell Biol. 34, 148-157 https://doi.org/10.1016/S1357-2725(01)00106-6
  3. Arora, S., Kaur, K. and Kaur, S. (2003) Indian medicinal plants as a reservoir of protective phytochemicals. Teratog. Carcinog. Mutagen. 1, 295-300
  4. Hashim, M. S., Lincy, V., Remya, V., Teena, M. and Anila, L. (2005) Effect of polyphenolic compounds from Coriandrum sativum on $H_2O_2$-induced oxidative stress in human lymphocytes. Food Chem. 92, 63-65 https://doi.org/10.1016/j.foodchem.2004.04.048
  5. Verhoeyen, M. E., Bovy, A., Collins, G., Muir, S., Robinson, S., de Vos, C. H. and Colliver, S. (2002) Increasing antioxidant levels in tomatoes through modification of the flavonoid biosynthetic pathway. J. Exp. Bot. 53, 2099-2106 https://doi.org/10.1093/jxb/erf044
  6. Yang, Y., Wang, J., Xu, C., Pan, H. and Zhang, Z. (2006) Maltol inhibits apoptosis of human neuroblastoma cells induced by hydrogen peroxide. J. Biochem. Mol. Biol. 39, 145-149 https://doi.org/10.5483/BMBRep.2006.39.2.145
  7. Ghosh, A., Sarkar, K., Sil, P. C. (2006) Protective Effect of a 43 kD Protein from the Leaves of the Herb, Cajanus indicus L on Chloroform Induced Hepatic-disorder. J. Biochem. Mol. Biol. 39, 197-207 https://doi.org/10.5483/BMBRep.2006.39.2.197
  8. Han, Y. T., Han, Z. W., Yu, G. Y., Wang, Y. J., Cui, R. U. and Wang, C. B. (2004) Inhibitory effect of polypeptide from Chlamys farreri on ultraviolet A-induced oxidative damage on human skin fibroblasts in vitro. Pharmacol. Res. 49, 265-274 https://doi.org/10.1016/j.phrs.2003.09.009
  9. Altman, G. H., Jakuba, C., Calabro, T., Horan, R. L., Chen, J., Lu, H., Richmond, J. and Kaplan, D. L (2003) Silk-based biomaterials. Biomaterials 24, 401-416 https://doi.org/10.1016/S0142-9612(02)00353-8
  10. Jin, H. J., Chen, J., Karageorgiou, V., Altman, G. H. and Kaplan, D. L. (2004) Human bone marrow stromal cell responses on electrospun silk fibroin mats. Biomaterials 25, 1039-1047 https://doi.org/10.1016/S0142-9612(03)00609-4
  11. Meinel, L., Hofmann, S., Karageorgiou, V., Kirker-Head, C., McCool, J., Gronowicz, G., Zichner, L., Langer, R., Novakovic, G. V. and Kaplan, D. L. (2005) The inflammatory responses to silk films in vitro and in vivo. Biomaterials 26, 147-155 https://doi.org/10.1016/j.biomaterials.2004.02.047
  12. Vepari, C. and Kaplan, D. L. (2007) Silk as a biomaterial. Prog. Polym. Sci. 32, 991-997 https://doi.org/10.1016/j.progpolymsci.2007.05.013
  13. Wang, Y., Kim, H. J., Vunjak-Novakovic, G. and Kaplan, D. L. (2006) Stem cell-based tissue engineering with silk biomaterials. Biomaterials 27, 6064-6082 https://doi.org/10.1016/j.biomaterials.2006.07.008
  14. Zhang, Y. Q. (2002) Applications of natural silk protein sericin in biomaterials. Biotechnol. Adv. 20, 91-100 https://doi.org/10.1016/S0734-9750(02)00003-4
  15. Kato, N., Sato, S., Yamanaka, A., Yamada, H., Fuwa, N. and Nomura, M. (1998) Silk protein sericin inhibits lipid peroxidation and tyrosinase activity. Biosci. Biotechnol. Biochem. 62, 145-147 https://doi.org/10.1271/bbb.62.145
  16. Zhaorigetu, S., Yanaka, N., Sasaki, M., Watanabe, H. and Kato, N. (2003) Inhibitory effects of silk protein, sericin on UVB-induced acute damage and tumor promotion by reducing oxidative stress in the skin of hairless mouse. J. Photochem. Photobiol. B, Biol. 71, 11-17 https://doi.org/10.1016/S1011-1344(03)00092-7
  17. Zhang, Y. Q., Tao, M. L., Shen, W. D., Mao, J. P. and Chen, Y. H. (2006) Synthesis of silk sericin peptides-L-asparaginase (SS-ASNase) bioconjugates and their characterization. J. Chem. Technol. Biotechnol. 81, 136-145 https://doi.org/10.1002/jctb.1370
  18. Chen, Q. and Ames, B. N. (1994) Senescence-like growth arrest induced by hydrogen peroxide in human diploid fibroblast F65 cells. Proc. Natl. Acad. Sci. USA 91, 4130-4134 https://doi.org/10.1073/pnas.91.10.4130
  19. Imlay, J. A., Chin, S. M. and Linn, S. (1988) Toxic DNA damage by hydrogen peroxide through the Fenton reaction in vivo and in vitro. Science 240, 640-642 https://doi.org/10.1126/science.2834821
  20. Zuliani, T., Denis, V., Noblesse, E., Schnebert, S., Andre, P., Dumas, M. and Ratinaud, M. H. (2005) Hydrogen peroxide-induced cell death in normal human keratinocytes is differentiation dependent. Free Radic. Biol. Med. 38, 307-316 https://doi.org/10.1016/j.freeradbiomed.2004.09.021
  21. Tesco, G., Latorraca, P., Piersanti, S., Sorbi, S., Piacentini, S. and Amaducci, L. (1992) Free radical injury in skin cultured fibroblasts from Alzheimer's disease patients. Ann. N. Y. Acad. Sci. 673, 149-153 https://doi.org/10.1111/j.1749-6632.1992.tb27446.x
  22. Ahmad, S. and Pardini, R. S. (2000) Mechanisms for regulating oxygen toxicity in phytophagous insects. Free Radic. Biol. Med. 8, 401-413 https://doi.org/10.1016/0891-5849(90)90107-T
  23. Ennamany, R., Marzetto, S., Saboureau, D. and Creppy, E. E. (1995) Lipid peroxidation induced by Boletus satanas: implication in $m^{5}dC$ variation in Vero cells related to inhibition of cell growth. Cell Biol. Toxicol. 11, 347-354 https://doi.org/10.1007/BF01305906
  24. Sasaki, M., Kato, N., Watanabe, H. and Yamada, H. (2000) Silk protein, sericin, suppresses colon carcinogenesis induced by 1, 2-dimethylhydrazine in mice. Oncol. Rep. 7, 1049-1052
  25. Takasu, Y., Yamada, H. and Tsubouchi, K. (2002) Isolation of three main sericin components from the cocoon of the silkworm, Bombyx mori. Biosci. Biotechnol. Biochem. 66, 2715-2718 https://doi.org/10.1271/bbb.66.2715
  26. Dash, R., Mukherjee, S. and Kundu, S. C. (2006) Isolation, purification and characterization of silk protein sericin from cocoon peduncles of tropical tasar silkworm, Antheraea mylitta. Int. J. Biol. Macromol. 38, 255-258 https://doi.org/10.1016/j.ijbiomac.2006.03.001
  27. Dash, R., Ghosh, S. K., Kaplan, D. L. and Kundu, S. C. (2007) Purification and biochemical characterization of a 70 kDa sericin from tropical tasar silkworm, Antheraea mylitta. Comp. Biochem. Physiol. B Biochem. Mol. Biol. 147, 129-134 https://doi.org/10.1016/j.cbpb.2007.01.009
  28. Darr, D., Combs, S., Dunston, S., Manning, T. and Pinnell, S. (1992) Topical vitamin C protects porcine skin from ultraviolet radiation-induced damage. Br. J. Dermatol. 127, 247-253 https://doi.org/10.1111/j.1365-2133.1992.tb00122.x
  29. Jagetia, G. C., Rajanikant, G. K. and Rao, S. K. (2003) Evaluation of the effect of ascorbic acid treatment in the artificially wounded mouse exposed to different doses of fractionated gamma radiation. Radiat. Res. 159, 371-380 https://doi.org/10.1667/0033-7587(2003)159[0371:EOTEOA]2.0.CO;2
  30. Katiyar, S. K., Afaq, F., Perez, A. and Mukhtar, H. (2001) Green tea polyphenol (3)-epigallocatechin-3-galate treatment of human skin inhibits ultraviolet radiation-induced oxidative stress. Carcinogenesis 22, 287-294 https://doi.org/10.1093/carcin/22.2.287
  31. Vaxman, F., Olender, S., Lambert, A., Nisand, G., Aprahamian, M., Bruch, J. F. (1995) Effect of pantothenic acid and ascorbic acid supplementation on human skin wound healing process. A double-blind, prospective and randomized trial. Eur. Surg. Res. 27, 158-166 https://doi.org/10.1159/000129395
  32. Sofia, S., McCarthy, M. B., Gronowicz, G. and Kaplan, D. L. (2001) Functionalized silk-based biomaterials for bone formation. J. Biomed. Mater. Res. 54, 139-148 https://doi.org/10.1002/1097-4636(200101)54:1<139::AID-JBM17>3.0.CO;2-7
  33. Mossman, T. (1983) Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J. Immunol. Methods 65, 55-63 https://doi.org/10.1016/0022-1759(83)90303-4
  34. Bergmeyer, H. U. and Bernt, E. (1963) Lactate dehydrogenase; in Methods of Enzymatic Analysis, Bergmeyer, H. U. (ed.), pp 574-579, Academic Press, London, UK
  35. Maehly, A. C. and Chance, B. (1954) The assay of catalase and peroxidase. Methods Biochem. Anal. 1, 357-364
  36. Niehaus, W. and Samuelson, B. (1968) Formation of malonaldehyde from phospholipids arachidonate during microsomal lipid peroxidation. Eur. J. Biochem. 6, 126-130 https://doi.org/10.1111/j.1432-1033.1968.tb00428.x

Cited by

  1. Induction of apoptosis and inhibition of telomerase activity in human lung carcinoma cells by the water extract of Cordyceps militaris vol.47, pp.7, 2009, https://doi.org/10.1016/j.fct.2009.04.014
  2. Sustained-release of protein from biodegradable sericin film, gel and sponge vol.407, pp.1-2, 2011, https://doi.org/10.1016/j.ijpharm.2011.01.006
  3. Emulsifying Properties of Sericin Obtained from Hot Water Degumming Process vol.40, pp.1, 2017, https://doi.org/10.1111/jfpe.12267
  4. Water Soluble Bioactives of Nacre Mediate Antioxidant Activity and Osteoblast Differentiation vol.8, pp.12, 2013, https://doi.org/10.1371/journal.pone.0084584
  5. Construction and Application of an Electronic Spatiotemporal Expression Profile and Gene Ontology Analysis Platform Based on the EST Database of the Silkworm,Bombyx mori vol.10, pp.114, 2010, https://doi.org/10.1673/031.010.11401
  6. Wound healing property of isolated compounds from Boesenbergia kingii rhizomes vol.184, 2016, https://doi.org/10.1016/j.jep.2016.03.001
  7. Anti-oxidant and Anti-aging Activities of Sericinjam Gland Hydrolysate Extract in Human Dermal Fibroblasts vol.39, pp.1, 2013, https://doi.org/10.15230/SCSK.2013.39.1.009
  8. Rice panicles: New promising unconventional cereal product for health benefits vol.66, 2015, https://doi.org/10.1016/j.jcs.2015.09.001
  9. Combination of physiologically balanced growth factors with antioxidants for reversal of facial photodamage vol.12, pp.1, 2010, https://doi.org/10.3109/14764170903449786
  10. An injectable silk sericin hydrogel promotes cardiac functional recovery after ischemic myocardial infarction vol.41, 2016, https://doi.org/10.1016/j.actbio.2016.05.039
  11. Antioxidant activity of chemically and enzymatically modified sericin extracted from cocoons of Bombyx mori vol.5, 2016, https://doi.org/10.1016/j.bcab.2016.01.010
  12. Effects of matured silkworm hemolymph on suppressing melanin synthesis vol.51, pp.2, 2013, https://doi.org/10.7852/jses.2013.51.2.207
  13. A Study on the Development of Baby Powder Using Silk gland Powder of Silkworm vol.50, pp.1, 2012, https://doi.org/10.7852/jses.2012.50.1.20
  14. A chitosan–glutathione based injectable hydrogel for suppression of oxidative stress damage in cardiomyocytes vol.34, pp.36, 2013, https://doi.org/10.1016/j.biomaterials.2013.08.031
  15. Non-immunogenic, porous and antibacterial chitosan and Antheraea mylitta silk sericin hydrogels as potential dermal substitute vol.167, 2017, https://doi.org/10.1016/j.carbpol.2017.02.098
  16. Anti-wrinkle effect of bone morphogenetic protein receptor 1a-extracellular domain (BMPR1a-ECD) vol.46, pp.9, 2013, https://doi.org/10.5483/BMBRep.2013.46.9.238
  17. Maturation and fertilisation of sheep oocytes cultured in serum-free medium containing silk protein sericin vol.63, pp.1, 2015, https://doi.org/10.1556/AVet.2015.009
  18. Comparison of protective effect of ascorbic acid on redox and endocannabinoid systems interactions in in vitro cultured human skin fibroblasts exposed to UV radiation and hydrogen peroxide vol.309, pp.4, 2017, https://doi.org/10.1007/s00403-017-1729-0
  19. A Study on the Development of Functional Cosmetics Using Silk-gland Powder of Silkworm vol.38, pp.2, 2012, https://doi.org/10.15230/SCSK.2012.38.2.163
  20. Photoprotection by Silk Cocoons vol.14, pp.10, 2013, https://doi.org/10.1021/bm401023h
  21. Effect of Sericin Supplementation DuringIn VitroMaturation on the Maturation, Fertilization and Development of Porcine Oocytes vol.49, pp.2, 2014, https://doi.org/10.1111/rda.12274
  22. Processing and characterization of silk sericin from Bombyx mori and its application in biomaterials and biomedicines vol.61, 2016, https://doi.org/10.1016/j.msec.2015.12.082
  23. In vivo bone regeneration ability of different layers of natural silk cocoon processed using an eco-friendly method vol.25, pp.8, 2017, https://doi.org/10.1007/s13233-017-5085-x
  24. Anti-invasive activity of sanguinarine through modulation of tight junctions and matrix metalloproteinase activities in MDA-MB-231 human breast carcinoma cells vol.179, pp.2-3, 2009, https://doi.org/10.1016/j.cbi.2008.11.009
  25. Sponge-Like Dressings Based on the Association of Chitosan and Sericin for the Treatment of Chronic Skin Ulcers. I. Design of Experiments–Assisted Development vol.105, pp.3, 2016, https://doi.org/10.1016/j.xphs.2015.11.047
  26. Potential Wound Healing Activities of Galla Rhois in Human Fibroblasts and Keratinocytes vol.43, pp.08, 2015, https://doi.org/10.1142/S0192415X15500925
  27. The Influence of Silkworm Species on Cellular Interactions with Novel PVA/Silk Sericin Hydrogels vol.12, pp.3, 2012, https://doi.org/10.1002/mabi.201100292
  28. Buffalo casein derived peptide can alleviates H 2 O 2 induced cellular damage and necrosis in fibroblast cells vol.69, pp.7, 2017, https://doi.org/10.1016/j.etp.2017.04.009
  29. The characterization of protein release from sericin film in the presence of an enzyme: Towards fibroblast growth factor-2 delivery vol.414, pp.1-2, 2011, https://doi.org/10.1016/j.ijpharm.2011.05.033
  30. Non-mulberry silk sericin/poly (vinyl alcohol) hydrogel matrices for potential biotechnological applications vol.49, pp.2, 2011, https://doi.org/10.1016/j.ijbiomac.2011.03.015
  31. Silk sericin loaded alginate nanoparticles: Preparation and anti-inflammatory efficacy vol.80, 2015, https://doi.org/10.1016/j.ijbiomac.2015.07.018
  32. Wound healing activity of ent-kaura-9(11),16-dien-19-oic acid isolated from Wedelia trilobata (L.) leaves vol.19, pp.13, 2012, https://doi.org/10.1016/j.phymed.2012.07.014
  33. Use of Silk Protein, Sericin, as a Sustained-Release Material in the Form of a Gel, Sponge and Film vol.58, pp.11, 2010, https://doi.org/10.1248/cpb.58.1480
  34. A novel nanocomposite for bone tissue engineering based on chitosan–silk sericin/hydroxyapatite: biomimetic synthesis and its cytocompatibility vol.5, pp.69, 2015, https://doi.org/10.1039/C5RA08216A
  35. Potential of 2D crosslinked sericin membranes with improved biostability for skin tissue engineering vol.347, pp.3, 2012, https://doi.org/10.1007/s00441-011-1269-4
  36. Effect of sericin on preimplantation development of bovine embryos cultured individually vol.78, pp.4, 2012, https://doi.org/10.1016/j.theriogenology.2012.03.021
  37. Sericins exhibit ROS-scavenging, anti-tyrosinase, anti-elastase, and in vitro immunomodulatory activities vol.58, 2013, https://doi.org/10.1016/j.ijbiomac.2013.03.054
  38. Para Rubber Seed Oil: New Promising Unconventional Oil for Cosmetics vol.63, pp.7, 2014, https://doi.org/10.5650/jos.ess14015
  39. Silk sericin ameliorates wound healing and its clinical efficacy in burn wounds vol.305, pp.7, 2013, https://doi.org/10.1007/s00403-013-1371-4
  40. Phenolic composition and antioxidant activity of Thai and Eri silk sericins vol.21, pp.2, 2012, https://doi.org/10.1007/s10068-012-0050-0
  41. Non-Bioengineered Silk Fibroin Protein 3D Scaffolds for Potential Biotechnological and Tissue Engineering Applications vol.8, pp.9, 2008, https://doi.org/10.1002/mabi.200800113
  42. Promoting Cell Survival and Proliferation in Degradable Poly(vinyl alcohol)-Tyramine Hydrogels vol.15, pp.10, 2015, https://doi.org/10.1002/mabi.201500121
  43. Antioxidant activities of two sericin proteins extracted from cocoon of silkworm (Bombyx mori) measured by DPPH, chemiluminescence, ORAC and ESR methods vol.2, pp.3, 2014, https://doi.org/10.3892/br.2014.244
  44. Sericin from Bombyx mori cocoons. Part I: Extraction and physicochemical-biological characterization for biopharmaceutical applications vol.61, 2017, https://doi.org/10.1016/j.procbio.2017.06.019
  45. The development of non-toxic ionic-crosslinked chitosan-based microspheres as carriers for the controlled release of silk sericin vol.17, pp.5, 2015, https://doi.org/10.1007/s10544-015-9991-4
  46. Silk sericin–alginate–chitosan microcapsules: Hepatocytes encapsulation for enhanced cellular functions vol.65, 2014, https://doi.org/10.1016/j.ijbiomac.2014.01.042
  47. Protective effects of ‘Khamira Abresham Hakim Arshad Wala’, a unani formulation against doxorubicin-induced cardiotoxicity and nephrotoxicity vol.21, pp.1, 2011, https://doi.org/10.3109/15376516.2010.529188
  48. Apoptosis Induction of Human Lung Carcinoma Cells by Chan Su (Venenum Bufonis) Through Activation of Caspases vol.2, pp.3, 2009, https://doi.org/10.1016/S2005-2901(09)60057-1
  49. Fabrication of cellulose nanofiber/chitin whisker/silk sericin bionanocomposite sponges and characterizations of their physical and biological properties vol.96, 2014, https://doi.org/10.1016/j.compscitech.2014.03.006
  50. Effects of Silk Sericin on Incision Wound Healing in a Dorsal Skin Flap Wound Healing Rat Model vol.22, 2016, https://doi.org/10.12659/MSM.897981
  51. Ultrastructure changes in buffalo (Bubalus bubalis ) oocytes before and after maturation in vitro with sericin 2017, https://doi.org/10.1111/asj.12839
  52. Biological activity assessment and phenolic compounds characterization from the fruit pericarp ofLitchi chinensisfor cosmetic applications vol.50, pp.11, 2012, https://doi.org/10.3109/13880209.2012.675342
  53. Sericin supplementation improves semen freezability of buffalo bulls by minimizing oxidative stress during cryopreservation vol.152, 2015, https://doi.org/10.1016/j.anireprosci.2014.11.015
  54. Evaluation of the wound healing potential of Wedelia trilobata (L.) leaves vol.141, pp.3, 2012, https://doi.org/10.1016/j.jep.2012.03.019
  55. Drug loading and release on tumor cells using silk fibroin–albumin nanoparticles as carriers vol.24, pp.3, 2013, https://doi.org/10.1088/0957-4484/24/3/035103
  56. Sericin consumption suppresses development and progression of colon tumorigenesis in 1,2-dimethylhydrazine-treated rats vol.67, pp.5, 2012, https://doi.org/10.2478/s11756-012-0093-y
  57. Free radical scavenging injectable hydrogels for regenerative therapy vol.71, 2017, https://doi.org/10.1016/j.msec.2016.09.087
  58. Non-mulberry silk gland fibroin protein 3-D scaffold for enhanced differentiation of human mesenchymal stem cells into osteocytes vol.5, pp.7, 2009, https://doi.org/10.1016/j.actbio.2009.02.033
  59. Enhanced in vitro developmental competence of sheep embryos following sericin supplementation of the in vitro maturation and in vitro culture media vol.136, 2016, https://doi.org/10.1016/j.smallrumres.2016.01.019
  60. The potential of silk sericin protein as a serum substitute or an additive in cell culture and cryopreservation vol.49, pp.6, 2017, https://doi.org/10.1007/s00726-017-2396-3
  61. Study on the Thixotropy of Sericin vol.07, pp.02, 2017, https://doi.org/10.12677/MS.2017.72032
  62. Inhibits Matrix Metalloproteinase Activity and Regulates Tight Junction Related Protein Expression in Hep3B Human Hepatocarcinoma Cells vol.13, pp.1, 2010, https://doi.org/10.1089/jmf.2009.1233
  63. Cryoprotective effect of sericin supplementation in freezing and thawing media on the outcome of cryopreservation in human sperm pp.1473-0790, 2018, https://doi.org/10.1080/13685538.2018.1529156
  64. Sericin enhances the developmental competence of heat-stressed bovine embryos vol.85, pp.8-9, 2018, https://doi.org/10.1002/mrd.23038
  65. Polyphenols from Acorn Leaves (Quercus liaotungensis) Protect Pancreatic Beta Cells and Their Inhibitory Activity against α-Glucosidase and Protein Tyrosine Phosphatase 1B vol.23, pp.9, 2018, https://doi.org/10.3390/molecules23092167
  66. Silk Protein-Based Membrane for Guided Bone Regeneration vol.8, pp.8, 2018, https://doi.org/10.3390/app8081214
  67. Instillation of Sericin Enhances Corneal Wound Healing through the ERK Pathway in Rat Debrided Corneal Epithelium vol.19, pp.4, 2018, https://doi.org/10.3390/ijms19041123