Estrogen Regulation of the 15-LOX/LXA4 Protective Lipid Circuit in the Corneal Inflammatory-Reparative Response

The cornea is a transparent, avascular tissue at the ocular surface that refracts light and protects the eye from microbial infection. Inflammation, while a necessary and beneficial part of wound repair, can be detrimental if left unchecked especially in the cornea where transparency is necessary for vision. The cornea has thus evolved tightly regulated circuits to promote wound healing, maintain host defense, and control inflammation. One such pathway that plays a key role in the cornea is the 15-lipoxygenase(LOX)/lipoxin(LX) A4 lipid circuit that is present endogenously in the uninjured cornea. LXA4 is a lipid mediator derived from dietary fatty acids that displays potent anti-inflammatory and pro-resolving bioactions. Previous results have demonstrated a key role for LXA4 in a murine model of corneal epithelial abrasion where 15-LOX (and subsequent LXA4) deficiency impairs rate of wound repair while topical administration of LXA4 restores corneal wound healing./nSex-specific differences have long been observed in an inflammatory and/or autoimmune setting. Females have a higher incidence of many inflammatory diseases including rheumatoid arthritis, systemic lupus erythematosus, and Sjogren's syndrome. Furthermore, there is a sex-specific difference in rate of dermal wound repair that changes post-menopause, showing that inflammation and wound healing are intimately linked. In the eye, Dry Eye Syndrome is a highly widespread inflammatory-based disease that affects predominantly post-menopausal women. Androgen deficiency has been implicated in Dry Eye disease progression. On the other hand, estrogen's role in the eye has been largely ignored despite the presence of estrogen and its two traditional receptors in leukocytes and most major ocular tissues. An epidemiological study of forty thousand female health professionals revealed a correlation between estrogen replacement therapy and an increase in reported dry eye symptoms, implicating estrogen in inflammatory regulation./nThe overarching aim of this dissertation was to determine if sex-specific differences are driven by estrogen regulation of the corneal inflammatory reparative response; more specifically, to establish a link, if any, between estrogen signaling and the 15-LOX/LXA4 lipid circuit. Given the potent protective bioactions of LXA4 in corneal wound healing, an understanding of sex-steroid regulation of the 15-LOX/LXA4 circuit may have important implications in ocular therapy and inflammatory disease progression. The following hypothesis was tested: is there a sex-specific difference in the corneal inflammatory reparative response and, if so, what role does estrogen play in the establishment of such differences?/nAn estrogen-driven sex-specific difference in corneal wound healing was demonstrated for the first time. Estrogen receptor (ER) β signaling downregulated corneal epithelial 15-LOX expression and subsequent LXA4 formation, the first study to link sex-specific differences and the action of estradiol to the protective anti-inflammatory 15-LOX/LXA4 circuit. Furthermore, a sex-specific difference in corneal inflammatory tone and leukocyte population upon re-injury was established. In vitro assays of phagocytosis demonstrated for the first time that phagocytosis is a potent stimulus for LXA4 formation, establishing a positive feedforward loop for LXA4 action in leukocytes whereby LXA4 stimulates phagocytosis and subsequent phagocytosis stimulates LXA4 production. Estrogen signaling via ERβ was also shown to upregulate neutrophil retention and delay clearance by inhibiting the LXA4-induced stimulation of phagocytosis. Taken together, these results provide evidence for an ERβ-driven upregulation of the inflammatory response and downregulation of the protective 15-LOX/LXA4 lipid pathway in the corneal wound healing response. Elucidation of these sex-specific and sex steroid-driven differences provides insights into the high female-dominated incidence of inflammatory diseases. Estrogen primes females for an amplified inflammatory response upon a disruption to the ocular surface. This has important ramifications in the cornea where chronic inflammation can lead to blindness.

Advisor: Gronert, Karsten