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浙江大学学报(医学版)  2015, Vol. 44 Issue (1): 15-23    DOI: 10.3785/j.issn.1008-9292.2015.01.003
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表没食子儿茶素没食子酸酯对小鼠角膜碱烧伤的治疗作用
吴联群1, 卢敏2
1. 第二军医大学附属长征医院眼科, 上海 200003;
2. 广东医学院附属医院 佛山市三水区人民医院眼科, 广东 佛山 528100
Efficacy of epigallocatechin gallate in treatment of alkali burn injury of murine cornea
WU Lian-qun1, LU Min2
1. Department of Ophthalmology, Shanghai Changzheng Hospital, Second Military Medical University, Shanghai 200003, China;
2. Department of Ophthalmology, Sanshui People's Hospital of Foshan, Affiliated Hospital of Guangdong Medical College, Foshan 528100, China
全文: PDF(4002 KB)  
摘要: 

目的:研究表没食子儿茶素没食子酸酯(EGCG)对角膜碱烧伤的治疗作用. 方法:制作C57BL/6J小鼠角膜碱烧伤模型,实验分为EGCG组和磷酸盐缓冲液(PBS)组,分别给予腹腔注射EGCG溶液或者等量PBS,裂隙灯显微镜和组织病理学观察和评价小鼠角膜上皮修复、新生血管生长以及炎症反应程度,免疫组织化学染色和实时定量PCR法检测血管内皮生长因子(VEGF)的表达,髓过氧化物酶定量测定评价中性粒细胞的浸润程度. 结果:EGCG组小鼠角膜上皮修复速率显著大于PBS组,碱烧伤后第1、3、7天的差异有统计学意义(均P <0.05).EGCG组和PBS组小鼠均见新生血管生长,在碱烧伤后第3、7、14天EGCG组新生血管评分和角膜切片中新生血管数量均显著低于PBS组,EGCG组的VEGF蛋白表达量在碱烧伤后第3、7天显著低于PBS组,EGCG组VEGFmRNA表达量在碱烧伤后第1、3、7天均低于PBS组,差异均有统计学意义(均P <0.05).碱烧伤后第7、14天EGCG组的炎症指数低于PBS组,第3、7、14天EGCG组角膜组织切片中中性粒细胞浸润数量和髓过氧化物酶检测值均低于PBS组,差异均有统计学意义(均P <0.05). 结论:腹腔注射EGCG可有效促进碱烧伤后小鼠角膜上皮修复,抑制新生血管形成和炎症细胞浸润.

关键词 儿茶素/药理学没食子酸/药理学角膜/药物作用烧伤, 化学/病理角膜新生血管化炎症    
Abstract

Objective: To evaluate the efficacy of epigallocatechin gallate (EGCG) in treatment of corneal alkali burn injury in mice. Methods: Corneal alkali burn injury was induced by sodium hydroxide method in C57BL/6J mice. The mice with cornea burns were treated intraperitoneally with EGCG solution or phosphate buffer solution (PBS) respectively. The healing of corneal epithelium, the formation of corneal neovascularization (CNV) and the inflammation reaction were assessed by slit-lamp microscopy and histological examination. Expression of vascular endothelial growth factor (VEGF) mRNA and protein in cornea was evaluated by real-time reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry, respectively. Myeloperoxidase (MPO) assay was used to quantitatively evaluate the polymorphonuclear neutrophils (PMNs) infiltration in the corneas. Results: The healing rate of corneal epithelium in EGCG group was significantly higher than that of PBS group at d1, d3 and d7 after treatment (d1: 41.0%±13.0% vs 23.8%±7.6%; d3: 76.6%±7.5% vs 61.2%±6.8%; d7: 87.8%±8.5% vs 74.0%±9.1%; all P <0.05). The CNV scores and the number of CNV in the corneal sections of EGCG group were significantly lower than those of PBS group at d3, d7 and d14 after treatment (CNV score: d3: 1.1±0.5 vs 6.6±1.0; d7: 1.3±0.3 vs 8.1±1.0; d14: 0.9±0.2 vs 9.2±1.1; CNV number: d3: 1.68±0.61 vs 2.92±0.95; d7: 4.80±1.36 vs 7.92±1.28; d14: 3.64±0.71 vs 5.88±0.76; all P <0.05). The expression of VEGF protein at d3 (0.19±0.05 vs 0.45±0.08) and d7 (0.42±0.07 vs 0.84±0.09), the expression of VEGF mRNA at d1, d3 and d7 in EGCG group were significantly lower than those in PBS group (all P <0.05). Compared to PBS group, the inflammatory index at d3 (3.2±0.4 vs 3.7±0.5) and d7 (2.3±0.5 vs 4.0±0.0), the number of PMNs in the corneal sections and the MPO values at d3, d7 and d14 in EGCG group were significantly decreased (PMNs: d3: 34.5±15.7 vs 90.0±28.8; d7: 17.1±11.4 vs 54.9±25.9; d14: 12.8±4.6 vs 39.0±17.9; all P <0.05). Conclusion: In the murine corneal alkali burn model, intraperitoneal injection of EGCG solution can promote the healing of corneal epithelium, inhibit the formation of CNV and reduce the inflammatory cell infiltration in the corneas.

Key wordsCatechin/pharmacology    Gallic acid/pharmacology    Cornea/drug effects    Alkalies    Burns, chemical/pathology    Corneal neovascularization    Inflammation
收稿日期: 2014-06-27
CLC:  R772.2  
基金资助:

浙江省中医药科学研究基金(2011ZA047);浙江省自然科学基金(LY13H120003).

通讯作者: 卢 敏(1969-),男,学士,主任医师,从事眼科工作;E-mail: drlumin@126.com     E-mail: drlumin@126.com
作者简介: 吴联群(1981-),女,博士,主治医师,从事眼科工作;E-mail: wulianqun19@aliyun.com
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引用本文:

吴联群, 卢敏. 表没食子儿茶素没食子酸酯对小鼠角膜碱烧伤的治疗作用[J]. 浙江大学学报(医学版), 2015, 44(1): 15-23.
WU Lian-qun, LU Min. Efficacy of epigallocatechin gallate in treatment of alkali burn injury of murine cornea. Journal of ZheJiang University(Medical Science), 2015, 44(1): 15-23.

链接本文:

http://www.zjujournals.com/xueshu/med/CN/10.3785/j.issn.1008-9292.2015.01.003      或      http://www.zjujournals.com/xueshu/med/CN/Y2015/V44/I1/15

[1] DONÀM, DELL'AICA I, CALABREASE F, et al. Neutrophil restraint by green tea: inhibition of inflammation, associated angiogenesis, and pulmonary fibrosis[J]. J Immunol, 2003, 170(8):4335-4341.
[2] AHMAD N, CHENG P, MUKHTAR H. Cell cycle dysregulation by green tea polyphenol epigallocatechin-3-gallate[J]. Biochem Biophys Res Commun, 2000, 275(2):328-334.
[3] ALBINI A. Tumor and endothelial cell invasion of basement membranes. The matrigel chemoinvasion assay as a tool for dissecting molecular mechanisms [J]. Pathol Oncol Res,1998,4(3):230-241.
[4] PIYAVIRIYAKUL S, SHIMIZU K, ASAKAWA T, et al. Anti-angiogenic activity and intracellular distribution of epigallocatechin-3-gallate analogs[J]. Biol Pharm Bull, 2011,34(3):396-400.
[5] TOSETTI F, FERRARI N, DE FLORA S, et al. Angioprevention: angiogenesis is a common and key target for cancer chemopreventive agents[J]. FASEB J, 2002, 16(1):2-14.
[6] ALBINI A, IWAMOTO Y, KLEINMAN H K, et al. A rapid in vitro assay for quantitating the invasion potential of tumor cells[J]. Cancer Res, 1987, 47(12):3239-3245.
[7] BALASUBRAMANIAN S, EFIMOVA T, ECKERT R L. Green tea polyphenol stimulates a Ras, MEKK1, MEK3, and p38 cascade to increase activator protein 1 factor-dependent involucrin gene expression in normal human keratinocytes[J]. J Biol Chem, 2002, 277(3):1828-1836.
[8] CAO Y, CAO R. Angiogenesis inhibited by drinking tea[J]. Nature, 1999, 398(6726):381.
[9] SÁNCHEZ-HUERTA V, GUTIÉRREZ-SÁNCHEZ L, FLORES-ESTRADE J. (-)-Epigallocatechin 3-gallate (EGCG) at the ocular surface inhibits corneal neovascularization[J]. Med Hypotheses, 2011,76(3):311-313.
[10] YUAN X, WILHELMUS K R. Corneal neovascularization during experimental fungal keratitis[J]. Mol Vis, 2009,15: 1988-1996.
[11] OZTVRK F, KURT E, CERI M, et al. The effect of propolis extract in experimental chemical corneal injury[J]. Ophthalmic Res, 2000,32(1):13-18.
[12] YAMADA J, DANA MR, SOTOZONO C, et al. Local suppression of IL-6 by receptor antagonist in the rat model of corneal alkali injury[J]. Exp Eye Res, 2003, 76(2):161-167.
[13] MEKKO G R, PIZZOLATTI M L, WASILEWSKI D, et al. The effect of subconjunctival bevacizumab on corneal neovascularization, inflammation and reepithelization in a rabbit model[J]. Clinics (Sao Paulo), 2011,66(8):1443-1450.
[14] CHAN K H, CHAN S C, YEUNG S C, et al. Inhibitory effect of Chinese green tea on cigarette smoke-induced up-regulation of airway neutrophil elastase and matrix metalloproteinase-12 via antioxidant activity[J]. Free Radic Res, 2012, 46(9):1123-1129.
[15] KATIYAR S K, MATSUI M S, ELMETS C A, et al. Polyphenolic antioxidant (-)-epigallocatechin-3-gallate from green tea reduces UVB-induced inflammatory responses and infiltration of leukocytes in human skin[J]. Photochem Photobiol, 1999,69(2):148-153.
[16] LEE H S, CHANHAN S K, OKANOBO A, et al. Therapeutic efficacy of topical epigallocatechin gallate in murine dry eye[J]. Cornea, 2011,30(12):1465-1472.
[17] OAK M H, CHATAIGNEAU M, KERAVIS T, et al. Red wine polyphenolic compounds inhibit vascular endothelial growth factor expression in vascular smooth muscle cells by preventing the activation of the p38 mitogen-activated protein kinase pathway[J]. Arterioscler Thromb Vasc Biol, 2003,23(6):1001-1007.
[18] SARTIPPOUR M R, SHAO Z M, HEBER D, et al. Green tea inhibits vascular endothelial growth factor(VEGF) induction in human breast cancer cells[J]. J Nutr, 2002,132(8):2307-2311.
[19] OAK M H, EL BEDOUI J, ANGLARD P, et al. Red wine polyphenolic compounds strongly inhibit pro-matrix metalloproteinase-2 expression and its activation in response to thrombin via direct inhibition of membrane type1-matrix metalloproteinase in vascular smooth muscle cells[J]. Circulation, 2004, 110(13):1861-1867.
[20] ANNABI B, LACHAMBRE M P, BOUSQUET-GAGNON N, et al. Green tea polyphenol (-)-epigallocatechin 3-gallate inhibits MMP-2 secretion and MT1-MMP-driven migration in glioblastoma cells[J]. Biochim Biophys Acta, 2002,1542(1-3):209-220.
[21] KOJIMA-YUASA A, HUA J J, KENNEDY D O, et al. Green tea extract inhibits angiogenesis of human umbilical vein endothelial cells through reduction of expression of VEGF receptors[J]. Life Sci, 2003,73(10):1299-1313.
[22] IIJIMA K, YOSHIZUMI M, HASHIMOTO M, et al. Red wine polyphenols inhibit vascular smooth muscle cell migration through two distinct signaling pathways[J]. Circulation, 2002,105(20):2404-2410.

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