Multifunctional roles of human cathelicidin (LL-37) at the ocular surface
Multifunctional roles of human cathelicidin (LL-37) at the ocular surface
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DOI：
10.1167/iovs.05-1649
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发表时间：
2006-06-01
影响因子：
4.4
通讯作者：
McDermott, Alison M.
中科院分区：
文献类型：
--
作者：
Huang, Ling C.;Petkova, Tihomira D.;McDermott, Alison M.
目的。本研究的目的是检测伤口愈合过程中角膜上皮中抗菌肽 LL-37 的表达，并研究 LL-37 是否刺激人角膜上皮细胞 (HCEC) 迁移、增殖和细胞因子产生。方法。通过 RT-PCR 和免疫染色测定组织切片和再上皮化之前（原始）和之后（再生）从角膜刮取的 HCEC 中 LL-37 的表达。 LL-37 对铜绿假单胞菌 (PA) 的抗菌活性是在 NaCl 和眼泪存在的情况下测定的。进行盲孔室测定以研究 LL-37 对迁移的影响。使用钙黄绿素-AM测定增殖，并通过MTT测定评估细胞毒性。进行ELISA以评估LL-37刺激HCEC细胞因子分泌的能力。结果。 LL-37 肽存在于整个角膜上皮中 (n = 4)。所有原始角膜上皮样本均表达低水平的 LL-37（it = 10）。受伤后 24（n = 3，共 5）或 48（n = 4，共 5）小时收集的再生上皮样本显示 LL-37 表达上调。 LL-37 在 NaCl 存在下杀死 PA（EC50 = 10.3 +/- 2.5 mu g/mL）并在泪液中保留其活性 (n = 3)。 LL-37 诱导 HCEC 迁移 (n = 5) 和 IL-8、IL-6、IL-1 beta 和 TNF-α 的分泌（2 至 23 倍，n = 4-7）。抑制剂研究表明，LL-37 的作用是通过多种途径介导的，涉及 G 蛋白偶联受体（迁移中的甲酰基肽受体样 1）和表皮生长因子受体（n = 2 至 5）。 LL-37 不刺激 HCEC 增殖 (n = 3)，高浓度 (> 10 μg/mL) 具有细胞毒性 (n = 3)。结论。 LL-37 表达在再生人角膜上皮中上调，在生理相关条件下对眼部病原体具有抗菌活性，并刺激 HCEC 迁移和细胞因子产生。这些发现表明 LL-37 作为一种多功能介质，有助于保护角膜免受感染并调节伤口愈合。
PURPOSE. The goals of this study were to examine the expression of the antimicrobial peptide LL-37 in the corneal epithelium during wound healing and to investigate whether LL-37 stimulates human corneal epithelial cell (HCEC) migration, proliferation, and cytokine production.METHODS. Expression of LL-37 was determined by RT-PCR and immunostaining in tissue sections and HCECs scraped from corneas before (original) and after (regrown) re-epithelialization. The antimicrobial activity of LL-37 against Pseudomonas aeruginosa (PA) was determined in the presence of NaCl and tears. Blind-well chamber assays were performed to study the effect of LL-37 on migration. Proliferation was determined using calcein-AM, and cytotoxicity was evaluated by MTT assay. ELISA was performed to assess the ability of LL-37 to stimulate HCEC cytokine secretion.RESULTS. LL-37 peptide was present throughout the corneal epithelium (n = 4). All original corneal epithelial samples expressed a low level of LL-37 (it = 10). Regrown epithelial samples collected 24 (n = 3 of 5) or 48 (n = 4 of 5) hours after wounding showed upregulated expression of LL-37. LL-37 killed PA in the presence of NaCl (EC50 = 10.3 +/- 2.5 mu g/mL) and retained its activity in tears (n = 3). LL-37 induced HCEC migration (n = 5) and secretion of IL-8, IL-6, IL-1 beta, and TNF-alpha (2- to 23-fold, n = 4-7). Inhibitor studies indicated that LL-37's effects are mediated through multiple pathways involving a G protein-coupled receptor (formyl peptide receptor-like 1 in migration) and the epidermal growth factor receptor (n = 2 to 5). LL-37 did not stimulate HCEC proliferation (n = 3) and high concentrations (> 10 mu g/mL) were cytotoxic (n = 3).CONCLUSIONS. LL-37 expression is upregulated in regenerating human corneal epithelium, has antibacterial activity against ocular pathogens under physiologically relevant conditions, and stimulates HCEC migration and cytokine production. These findings suggest that LL-37 acts as a multifunctional mediator that helps protect the cornea from infection and modulates wound healing.