Kolar Satya Sree, Baidouri Hasna, McDermott Alison M
The Ocular Surface Institute, University of Houston, College of Optometry, Houston, Texas, United States.
Invest Ophthalmol Vis Sci. 2017 May 1;58(5):2463-2472. doi: 10.1167/iovs.16-20658.
Fusarium solani (F. solani) keratitis is a potentially sight-threatening fungal infection of the cornea. Antimicrobial peptides (AMPs), such as human β-defensins (hBDs) and cathelicidins, essential components of the immune system, likely have a protective role against F. solani keratitis. We examined the role of pattern recognition receptors (PRRs), Dectin-1, and TLR2 in F. solani-induced modulation of AMP expression in vitro.
Human corneal epithelial cells (HCECs) were exposed to heat-inactivated F. solani or pathogen-associated molecular patterns (PAMPs) of F. solani (Zymosan or Zymosan Depleted) for 6, 12, or 24 hours following which AMP mRNA and protein levels were determined. Involvement of TLR2 and Dectin-1 was confirmed by using siRNA knock-down (TLR2 and Dectin-1) or chemical inhibitor BAY 61-3606 (Dectin-1). The functional significance of AMP upregulation was tested using culture supernatant from F. solani or PAMP-treated HCECs against F. solani in the presence of hBD2 or LL37 neutralizing antibody.
We confirm that HCECs express Dectin-1 and TLR2. HCECs demonstrated upregulation of AMPs hBD2 and cathelicidin LL37 following exposure to heat-inactivated F. solani or PAMPs. TLR2 and Dectin-1 knockdown and BAY 61-3606 treatment decreased AMP mRNA upregulation confirming PRR involvement. The culture supernatant from F. solani or PAMP-treated HCECs showed substantial killing of F. solani and hBD2 or LL37 neutralizing antibody significantly decreased this effect implicating involvement of these AMPs.
These findings demonstrate that Dectin-1 and TLR2 have an important role in regulating F. solani-induced AMP expression in corneal epithelial cells.
茄病镰刀菌角膜炎是一种可能威胁视力的角膜真菌感染。抗菌肽（AMPs），如人类β-防御素（hBDs）和cathelicidins，作为免疫系统的重要组成部分，可能对茄病镰刀菌角膜炎具有保护作用。我们在体外研究了模式识别受体（PRRs）、Dectin-1和TLR2在茄病镰刀菌诱导的AMPs表达调节中的作用。
将人角膜上皮细胞（HCECs）暴露于热灭活的茄病镰刀菌或茄病镰刀菌的病原体相关分子模式（PAMPs）（酵母聚糖或去糖酵母聚糖）6、12或24小时，然后测定AMPs的mRNA和蛋白质水平。通过使用小干扰RNA敲低（TLR2和Dectin-1）或化学抑制剂BAY 61-3606（Dectin-1）来证实TLR2和Dectin-1的参与。在存在hBD2或LL37中和抗体的情况下，使用来自茄病镰刀菌或PAMP处理的HCECs的培养上清液对抗茄病镰刀菌，测试AMPs上调的功能意义。
我们证实HCECs表达Dectin-1和TLR2。HCECs在暴露于热灭活的茄病镰刀菌或PAMPs后，AMPs hBD2和cathelicidin LL37表现出上调。TLR2和Dectin-1敲低以及BAY 61-3606处理降低了AMPs mRNA上调，证实了PRR的参与。来自茄病镰刀菌或PAMP处理的HCECs的培养上清液显示出对茄病镰刀菌的显著杀伤作用，hBD2或LL37中和抗体显著降低了这种作用，表明这些AMPs参与其中。
这些发现表明Dectin-1和TLR2在调节角膜上皮细胞中茄病镰刀菌诱导的AMPs表达中起重要作用。
