Ultraviolet B (UVB) irradiation causes skin inflammation, photodamage, and photoaging, all major medical problems for Armed Forces personnel and veterans. Because UVB is absorbed mostly by the epidermis, with just 10% reaching the dermis, we seek to investigate likely signals from epidermis to mediate inflammation and damage in the dermis in response to UVB. Extracellular vesicles (EVs), which are small lipid bilayer membrane structures released by many cells, carry lipids, proteins, and nucleic acids to mediate cell-cell communication. Data from us and others indicate that UVB irradiation of keratinocytes (KCs) triggers the release of biologically active EVs. Our preliminary data suggest that these UVB-induced KC-derived EVs (KC-EVs) activate several key pro-inflammatory pathways in dermal cells, including the stimulator of interferon genes (STING), the NLRP3 inflammasome, and NF-KB. Our central hypothesis is that UVB provokes KCs in the epidermis to release biologically active EVs that then leave the epidermis to act on dermal fibroblasts, immune cells in the dermis, and even systemically, to mediate photoaging and other harms. There are three Aims. Aim 1. To characterize KC-EVs induced by UVB irradiation and the immune cells they act upon in vivo in the dermis and in vitro. Experimental approaches include UVB-irradiation of mice followed by isolation and characterization of KC-EVs from dermis (1a); use in vivo of GW4869, an agent that blocks EV generation, to determine which immune cell types are no longer recruited by UVB to dermis, or are recruited but not activated (1b); and identification of target immune cell types that respond to KC-EVs in vitro (1c) and in vivo (1d), including after intradermal injection of UVB- induced KC-EVs into mice. Aim 2. To characterize molecular mechanisms by which KC-EVs mediate UVB-induced dermal inflammation. We will test the roles of STING, the inflammasome, and NF-KB (2a) using experimental systems in vivo and in vitro from Aim 1. We will also test the roles of specific biologically active molecules carried by KC- EVs, such as dsDNA, peroxidized lipids known to function as danger signals, and specific proteins, such as integrins (2b). Aim 3. To assess potential therapeutic strategies that ameliorate KC-EV-mediated dermal inflammation under UVB irradiation. We will test whether melatonin, an endogenous inhibitor of STING and inflammasomes, suppresses UVB -induced KC-EV production and dermal inflammation in vivo (3a). We will also assess the mechanism by which melatonin blocks the ability of KC-EVs to mediate dermal inflammation, i.e., by interfering with STING, the inflammasome, and/or NF-KB (3b), using experimental systems in vivo and in vitro from Aim 1. Overall, these aims will reveal a better understanding of the role of epidermis, especially KC-EVs, in UVB-induced photoaging and skin inflammation, and provide potential novel therapeutic strategies to prevent and treat these conditions through targeting ...