miR-183/ALOX15信号通路调控促炎症消退介质在银屑病中的作用及机制研究

Neutrophils chemotaxis, abnormal proliferation of keratinocytes and differentiation imbalances of lymphocyte form the pathological basis of psoriasis, which well correspond to the effect of anti and pro-resolving inflammation of pro-resolving mediators, and it has been confirmed that they can inhibit the formation of psoriasis. We demonstrated that the expression of pro-resolving mediators and ALOX15, the key enzyme in pro-resolving mediator production, were down-regulated in psoriasis skin lesion. It was reported that the expression of miR-183 was obviously increased psoriasis lesions, we also found the high level expression of miR-183 and enhanced cell proliferation in lipopolysaccharide induced activated-keratinocytes. Through gene sequence comparison, we found that it exists the binding site of miR-183 in the 3’-UTR of ALOX15. We propose a hypothetical, miR-183 regulate the expression of pro-resolving mieditors through ALOX15 pathway, and promote the development of psoriasis. This project intends to clarify the effect and molecular mechanism of the regulation of the expression of pro-resolving mieditors through miR-183/ALOX15 pathway, and investigate the role of miR-183 on the formation of psoriasis in cellular and animal level. By the interference with key molecules such as miR-183 and ALOX15, to inhibit the formation of psoriasis, this study could provide new theoretical basis and new target to the therapy of psoriasis.

银屑病的病理基础是中性粒细胞趋化、角质形成细胞（KC）异常增殖以及淋巴细胞分化失衡，这与促炎症消退介质（脂氧素和消退素等）的抗炎促消退效应很好的对应，并已证实其能抑制银屑病发病。我们发现银屑病皮损区促炎症消退介质和及其关键酶ALOX15表达均明显下降。有报道，人银屑病皮损中miR-183明显上调，我们前期也发现脂多糖活化的KC中，miR-183表达显著升高，且KC增殖加强。通过基因序列对比发现 ALOX15的3’-UTR区存在miR-183的结合位点。我们提出假想，miR-183可通过ALOX15调控促炎症消退介质，并促进银屑病发展。本项目拟在细胞和动物水平上，明确miR-183通过ALOX15对多种促炎症消退介质的调控作用和机制；阐明miR-183对银屑病发病的影响。通过对关键分子miR-183、ALOX15进行干预，抑制银屑病皮损的发展，为治疗银屑病提供新的靶点和理论基础。

银屑病的发病率在包括中国在内的世界各地均呈上升趋势，其高复发性及主要累及青壮年劳动者的特点给患者及其家庭、社会带来沉重的负担。银屑病的发病基础涉及角质形成细胞（KC）过度增殖、中性粒细胞浸润、淋巴细胞的异常分化和浸润以及大量炎性因子的合成等多个环节。我们的前期研究已经证实，由花生四烯酸衍生的、被喻为炎症“刹车信号”的促炎症消退介质--脂氧素在银屑病发病中发挥着明确的抗炎促消退效应。同时我们发现，银屑病皮损区促炎症消退介质—脂氧素和及其关键酶15-LOX表达均明显下降。有报道，人银屑病皮损中miR-183明显上调，我们前期也发现脂多糖活化的KC中miR-183表达显著升高，且KC增殖加强。通过基因序列对比发现 15-LOX的3’-UTR区存在miR-183的结合位点。在本项目中，我们探讨了miR-183是否可通过15-LOX调控脂氧素合成，最终参与银屑病发病。我们发现，与正常人相比，在进展期寻常性银屑病患者的血浆中miR-183的含量明显上升，而在咪喹莫特诱导的银屑病小鼠的血浆中，miR-183的含量同样也较正常组小鼠显著上升。通过给与小鼠尾静脉注射合成的miR-183抑制剂（anti-183）下调miR-183的表达。我们发现，下调miR-183可以明显从形态及病理上改善咪喹莫特诱导的银屑病小鼠的皮损，同时模型小鼠血清中IL-1β， TNF-a，IL-17和IL-23蛋白的表达明显减少。在LPS刺激后体外分离培养的正常人角质形成细胞（NHEKs）中，我们发现anti-183可以抑制NHEKs的细胞增殖以及下调炎性因子（IL-1β，TNF-a，IL-6和IL-8）mRNAs和蛋白含量的表达。而anti-183组小鼠皮损中银屑病相关信号因子TRAF6及STAT3的蛋白表达比anti-NC组小鼠明显减少。对模型小鼠皮损匀浆后ELISA检测发现，anti-183上调了皮损中促炎症消退介质—脂氧素A4（LXA4）和脂氧素B4（LXB4）的含量。通过使用双荧光素酶法，我们发现，15-LOX是miR-183的直接靶点。上述结果表明，miR-183 可能通过负向调控 15-LOX 表达来影响LXA4及LXB4等促炎症消退脂质介质的表达，从而炎症细胞和炎性因子不能被适时地清除，炎症反应不能被终止，最终导致银屑病的发生和迁延不愈。