Institute of Applied Ophthalmobiology (IOBA), Universidad de Valladolid, Valladolid, Spain.
Institute of Applied Ophthalmobiology (IOBA), Universidad de Valladolid, Valladolid, Spain; Networking Research Center on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Carlos III National Institute of Health, Spain.
Exp Eye Res. 2024 Apr;241:109854. doi: 10.1016/j.exer.2024.109854. Epub 2024 Mar 5.
Mucosal chemokines have antimicrobial properties and play an important role in mucosal immunity. However, little is known about their expression on the ocular surface. This study aimed to analyze the expression of the mucosal chemokines CCL28, CXCL14 and CXCL17 in corneal and conjunctival epithelial cells under in vitro dry eye (DE) conditions, and in conjunctival samples from healthy subjects and DE patients. Human corneal epithelial cells (HCE) and immortalized human conjunctival epithelial cells (IM-HConEpiC) were incubated under hyperosmolar (400-500 mOsM) or inflammatory (TNF-α 25 ng/mL) conditions for 6 h and 24 h to measure CCL28, CXCL14, and CXCL17 gene expression by RT-PCR and their secretion by immunobead-based analysis (CCL28, CXCL14) and ELISA (CXCL17). Additionally, twenty-seven DE patients and 13 healthy subjects were included in this study. DE-related questionnaires (OSDI, mSIDEQ and NRS) evaluated symptomatology. Ocular surface integrity was assessed using vital staining. Tactile sensitivity was measured with Cochet-Bonnet esthesiometer, and mechanic and thermal (heat and cold) sensitivity using Belmonte's non-contact esthesiometer. Subbasal nerve plexus and dendritic cell density were analyzed by in vivo confocal microscopy. Conjunctival cells from participants were collected by impression cytology to measure mucosal chemokines gene expression by RT-PCR. Our results showed that HCE and IM-HConEpiC cells increased CCL28, CXCL14, and CXCL17 secretion under hyperosmolar conditions. The gene expression of CCL28 was significantly upregulated in conjunctival samples from DE patients. CCL28 expression correlated positively with symptomatology, corneal staining, heat sensitivity threshold, and dendritic cell density. CXCL14 expression correlated positively with age, ocular pain, conjunctival staining, tactile sensitivity, and image reflectivity. CXCL17 expression correlated positively with corneal staining. These results suggest that corneal and conjunctival epithelial cells could be a source of CCL28, CXCL14, and CXCL17 on the ocular surface and that CCL28 might be involved in DE pathogenesis.
黏膜趋化因子具有抗菌特性，并在黏膜免疫中发挥重要作用。然而，人们对其在眼表面的表达知之甚少。本研究旨在分析体外干燥眼 (DE) 条件下和健康受试者及 DE 患者结膜样本中角膜和结膜上皮细胞中黏膜趋化因子 CCL28、CXCL14 和 CXCL17 的表达。将人角膜上皮细胞 (HCE) 和永生化人结膜上皮细胞 (IM-HConEpiC) 在高渗 (400-500 mOsM) 或炎症 (TNF-α 25 ng/mL) 条件下孵育 6 小时和 24 小时，通过 RT-PCR 测量 CCL28、CXCL14 和 CXCL17 的基因表达，并通过免疫珠分析 (CCL28、CXCL14) 和 ELISA (CXCL17) 测量其分泌。此外，本研究纳入了 27 名 DE 患者和 13 名健康受试者。使用 OSID、mSIDEQ 和 NRS 等 DE 相关问卷评估症状。使用活体染色评估眼表面完整性。使用 Cochet-Bonnet 触觉计测量触觉敏感度，使用 Belmonte 的非接触式触觉计测量机械和热 (热和冷) 敏感度。使用体内共聚焦显微镜分析基底神经丛和树突状细胞密度。通过印迹细胞学收集参与者的结膜细胞，通过 RT-PCR 测量黏膜趋化因子的基因表达。我们的结果表明，HCE 和 IM-HConEpiC 细胞在高渗条件下增加 CCL28、CXCL14 和 CXCL17 的分泌。DE 患者结膜样本中 CCL28 的基因表达显著上调。CCL28 的表达与症状、角膜染色、热敏感度阈值和树突状细胞密度呈正相关。CXCL14 的表达与年龄、眼痛、结膜染色、触觉敏感度和图像反射率呈正相关。CXCL17 的表达与角膜染色呈正相关。这些结果表明，角膜和结膜上皮细胞可能是眼表面 CCL28、CXCL14 和 CXCL17 的来源，CCL28 可能参与 DE 的发病机制。
