LL-37 via EGFR transactivation to promote high glucose-attenuated epithelial wound healing in organ-cultured corneas.

With rapid increases in the prevalence of diabetes mellitus (DM) worldwide, ocular complications have become a leading cause of blindness in the world.1 In addition to abnormalities of the retina (diabetic retinopathy) and the lens (cataract), various types of corneal epithelial disorders are also relatively common in persons with DM.2 Abnormalities of the cornea include defects in epithelium-basement membrane adhesion3–8 and altered epithelial functions such as basal cell degeneration,9 superficial punctate keratitis,10 breakdown of barrier function,11 fragility,12 recurrent erosions, and persistent epithelial defects.13 Furthermore, a considerable delay in corneal reepithelialization is often observed in patients with diabetes who undergo vitrectomy.2,13–15 Delayed healing of the epithelial defect may also result in sight-threatening complications, such as stromal opacification, surface irregularity, and microbial keratitis.15 Hence, accelerating epithelial healing is of great importance for effectively treating these complications. We previously demonstrated that epithelial wounding induces the ectodomain shedding of heparin-binding EGF-like growth factor (HB-EGF), subsequent activation of epidermal growth factor receptor (EGFR), and its two major downstream effectors, phosphoinositide 3-kinase (PI3K) and extracellular signal-regulated kinase (ERK).16 Wound-induced EGFR activation plays a critical role in corneal epithelial wound healing.16–18 More recently, we discovered that, consistent with attenuating epithelial wound closure, high glucose also suppresses the EGFR-PI3K-AKT signaling pathway ex vivo.19 Importantly, hyperglycemia was found to disturb the expression and distribution of phospho-AKT in diabetic human corneas, suggesting an essential role of EGFR signaling in maintaining a healthy cornea.19 Echoing this concept is the finding that anti-EGFR cancer treatments resulted in diffuse punctuate keratitis and corneal erosion in patients.20,21 Thus, maintaining a proper level of EGFR signaling is critical for the physiologic state of an epithelium in tissue, and long-term exposure to hyperglycemia may inevitably affect the EGFR signaling apparatus, leading to cell dysfunction, including compromised barrier function and delayed wound healing. LL-37, an epithelium and immune cell–secreted polypeptide from gene cathelicidin, belongs to a group of antimicrobial peptides that play an important role in host defense against infection.22,23 LL-37 plays a key role in innate immunity against a broad spectrum of microbes in a number of tissues, including the cornea,24 urinary tract,25 and skin.26,27 In addition to acting as a natural antibiotic for innate immune defense, emerging evidence suggests that LL-37 is an important cell-signaling molecule, especially during tissue damage.28–30 Moreover, LL-37 has been documented to interact with the G protein-coupled receptor and to transactivate EGFR, leading to keratinocyte migration.31 In human corneas, LL-37 peptide was detected throughout the epithelium, albeit at a low level, and its expression was upregulated in regrown human CECs.24 Wound-induced cathelicidin expression was also observed in B6 mice (Kumar A, Yu FS, unpublished data, 2009). To date, the process and release of LL-37 from cathelicidin in HCECs remains elusive. A recent report30 revealed that adenoviral transfer of LL-37 effectively promotes reepithelialization of excisional skin wounds in diabetic mice. In the present study, we reported that LL-37 induces HB-EGF shedding, EGFR activation, and corneal epithelial wound closure in vitro. Moreover, we demonstrated that LL-37 partially restores high glucose–delayed wound healing ex vivo via EGFR signaling. Our results indicate that LL-37, both antimicrobial and regenerative, is a potential therapeutic for hastening epithelial wound healing and preventing infection in diabetic keratopathy.