UVB radiation-generated microvesicle particles as effectors for photosensitivity

摘要 紫外线B(UVB)辐射对皮肤有深远的影响，并会因发烧而产生全身后果 从免疫抑制到维生素D的产生。由于UVB只穿透了表皮，一个主要的未回答 光生物学中的问题是UVB治疗的表皮皮肤如何发出全身信号。最近的研究表明 表明从细胞中释放的被称为微囊泡颗粒(MVP)的膜结合的小囊泡 对各种应激源的反应可以作为强有力的信号媒介，因为它们能够携带核和 细胞质成分。我们已经证明UVB从上皮细胞产生MVP释放，并 皮肤，这可能为UVB介导的系统信号提供一种潜在的机制。我们的团队和其他人 已经证明UVB辐射会产生高水平的脂质介质血小板激活因子 酶促生产(PAF)和非酶反应生产PAF受体(PAFR)激动剂 氧气物种。我们正在进行的研究使用抗氧化剂和PAFR表达/空细胞系和 酸性鞘磷脂酶(ASMase)的药理/遗传抑制与此有关 在UVB产生的MVP(UVB-MVP)中，PAFR信号导致aSMase激活。最后，我们提供 UVB-MVP不携带大量蛋白质细胞因子，但携带生物活性PAF的证据 激动剂。我们有证据表明代谢不稳定的PAF激动剂在MVP和这些 生物活性脂质参与了UVB的急性促炎和延迟性免疫抑制作用。还没有 关于UVB-MVP是如何产生的，以及这种新的途径是否可以被利用来治疗，还存在着知识空白 光敏性疾病。为了验证UVB在人体内产生MVP的假设，设计了两个目标 皮肤以依赖PAF的方式参与aSMase，并通过其传递局部和全身效应 携带PAF激动剂。AIM 1将使用体外细胞系和小鼠遗传和药理学模型来 确定UVB-MVP的产生机制。这一目标将验证定义UVB角色的工具- MVP在UVB急性促炎作用中的作用，使用我们已有的小鼠光敏模型 先前证明的是PAF依赖和单独的光敏性小鼠狼疮模型。AIM 2将使用 体外皮肤外植体和活体人体受试者，以测试人体皮肤产生UVB-MVP的能力。 此外，我们将确定口服抗氧化剂和外用aSMase抑制剂治疗是否会阻断UVB-MVP 在人类中产生和UVB介导的急性炎症。最后，我们将测试人类受试者是否表现出 临床上对UVB的光敏反应与UVB-MVP的增加有关，如果局部使用aSMase抑制剂阻断 UVB-MVP和夸张的皮肤反应。该项目的成功完成将(I)解决 光生物学中的一个重要问题，即角质形成细胞特异性刺激如何产生系统信号 作用，(Ii)提供阻断UVB局部和全身效应的药理机制。这些研究还包括 对理解包括电离辐射在内的其他促氧化应激源的影响的影响。

Abstract Ultraviolet B (UVB) radiation has profound effects upon skin and generates systemic consequences from fever to immunosuppression to vitamin D production. As UVB only penetrates the epidermis, a major unanswered question in photobiology is how UVB-treated epidermal skin sends systemic signals. Recent studies have indicated that small membrane-bound vesicles known as microvesicle particles (MVP) released from cells in response to various stressors can act as potent signaling agents due to their ability to carry nuclear and cytoplasmic components. We have demonstrated that UVB generates MVP release from epithelial cells and skin, which could provide a potential mechanism for UVB-mediated systemic signaling. Our group and others have demonstrated that UVB radiation generates high levels of the lipid mediator Platelet-activating factor (PAF) produced enzymatically and PAF-receptor (PAFR) agonists produced non-enzymatically via reactive oxygen species. Our ongoing studies using antioxidants and PAFR-expressing/null cell lines and pharmacologic/genetic inhibition of the enzyme acid sphingomyelinase (aSMase) have implicated involvement of PAFR signaling resulting in aSMase activation in UVB generated MVP (UVB-MVP). Finally, we provide evidence that UVB-MVP do not carry significant amounts of protein cytokines, yet carry bioactive PAF agonists. We have evidence that metabolically labile PAF agonists are protected traveling in MVP and these bioactive lipids are involved in acute pro-inflammatory and delayed immunosuppressive effects of UVB. Yet knowledge gaps exist as to how UVB-MVP are generated and if this new pathway can be exploited to treat photosensitivity diseases. Two aims are designed to test the hypothesis that UVB generates MVP in human skin in a PAF-dependent manner involving aSMase and transfers both local and systemic effects via their carried PAF agonists. Aim 1 will use in vitro cell lines and murine genetic and pharmacologic models to determine the mechanisms of UVB-MVP generation. This aim will validate tools to define the roles of UVB- MVP in acute pro-inflammatory effects of UVB, using a murine model of photosensitivity that we have previously demonstrated is PAF-dependent and a separate photosensitive murine lupus model. Aim 2 will use both ex vivo skin explants and in vivo human subjects to test the ability of human skin to generate UVB-MVP. Moreover, we will define if oral antioxidants and topical aSMase inhibitor treatments will block UVB-MVP generation and UVB-mediated acute inflammation in humans. Finally, we will test if human subjects exhibiting clinical photosensitivity respond to UVB with increased UVB-MVP and if a topical aSMase inhibitor blocks the UVB-MVP and the exaggerated skin reactions. Successful completion of this project will (i) address an important question in photobiology as to how a keratinocyte-specific stimulus can generate systemic signaling effects, (ii) offer pharmacologic mechanisms to block UVB local and systemic effects. These studies also have implications for understanding the effects of other pro-oxidative stressors including ionizing radiation.