Lysosomal phospholipase A2 in autoimmune disease

描述（由申请人提供）：系统性红斑狼疮历来是且仍然是一种知之甚少的自身免疫性疾病。最近在识别狼疮方面取得了进展，促进了人类狼疮许多方面的单基因改变。尽管取得了这些进展，但仍没有统一的假设，也没有基于新机制的定向免疫治疗这种疾病。近年来，一些临床和实验研究指出，凋亡细胞清除障碍是狼疮的一个共同特征。由于巨噬细胞识别、结合和摄取凋亡细胞的分子基础已经被更好地定义，与吞噬受体“吃我”信号表达受损相关的基因改变与狼疮小鼠模型有关。例子包括乳脂球egf因子8和c-mer原癌基因酪氨酸激酶。磷脂溶酶-磷脂酰胆碱已被确定为巨噬细胞募集参与凋亡细胞清除的重要“找我”信号。然而，lyso-磷脂酰胆碱是如何作为发现信号产生的尚不清楚，编码酶或参与lyso-PC形成的酶的基因的可比改变的鉴定将导致狼疮表型尚未报道。我们的实验室发现、克隆并鉴定了一种新的磷脂酶A2，命名为溶酶体磷脂酶A2。这种酶在磷脂酶A2s家族中是不同的。溶酶体磷脂酶A2的特点是酸性pH值最佳，神经酰胺转酰基化的能力，以及巨噬细胞对吞噬刺激（如酶酶酶）的分泌。缺乏溶酶体磷脂酶A2表达的敲除小鼠的特征是迟发性自身免疫表型，模仿狼疮的大多数方面。这些小鼠出现淋巴增生伴肝脾肿大、消瘦和肾功能衰竭。小鼠表现出明显的ANAs阳性，抗dsdna滴度，高循环免疫球蛋白水平和肾小球肾炎。脾脏和淋巴结的特点是与巨噬细胞相关的高水平凋亡小体的持续存在，这与凋亡细胞清除的缺陷一致。基于这些观察，提出以下主要假设。溶酶体磷脂酶A2是巨噬细胞消化和清除凋亡细胞所必需的。我们提出以下具体目的来验证这一假设：1)对LPLA2缺失的小鼠进行免疫表型分析。2)确定溶酶体磷脂酶A2缺失抑制或消除巨噬细胞对凋亡细胞清除的机制。3)确定溶酶体磷脂酶A2是否足以或必须挽救LPLA2缺失小鼠的狼疮表型。4)确定肼嗪、奎尼丁、普鲁卡因酰胺对溶酶体磷脂酶A2的抑制是否为药物性狼疮的基础。

DESCRIPTION (provided by applicant): Systemic lupus erythematosus historically has been and still remains a poorly understood autoimmune disorder. Recent advances have been made in identifying lupus promoting single gene alterations that phenocopy many aspects of lupus in humans. Despite these advances there remains no unifying hypothesis and no new mechanism based, directed immunotherapy for this disease. In recent years several clinical and experimental studies have pointed to the impairment in the clearance of apoptotic cells as a common feature of lupus. As the molecular basis for the recognition, binding, and uptake of apoptotic cells by macrophages has become better defined, gene alterations associated with the impaired expression of phagocytic receptors for "eat me" signals are associated with murine models of lupus. Examples include the milk fat globule-EGF factor 8 and the c-mer proto-oncogene tyrosine kinase. The phospholipid lyso- phosphatidylcholine has been identified as an important "find me" signal for the recruitment of macrophages involved in the clearance of apoptotic cells. However, how lyso-phosphatidylcholine is generated as a find me signal is unknown and the identification of comparable alterations in the genes encoding the enzyme or enzymes involved in lyso-PC formation that would result in a lupus phenotype have not been reported. Our laboratory discovered, cloned, and characterized a novel phospholipase A2 named lysosomal phospholipase A2. This enzyme is distinct among the family of phospholipase A2s. Lysosomal phospholipase A2 is characterized by an acidic pH optimum, the ability to transacylate ceramide, and it secretion from macrophages in response to phagocytic stimuli such as zymosan. A knockout mouse, lacking the expression of lysosomal phospholipase A2 is characterized by a late onset autoimmune phenotype that mimics most aspects of lupus. These mice develop lymphoproliferation with hepatosplenomegaly, wasting, and renal failure. The mice exhibit markedly positive ANAs, anti-dsDNA titers, high circulating immunoglobulin levels, and glomerulonephritis. The spleens and lymph nodes are characterized by the persistence of high levels of apoptotic bodies in association with resident macrophages, consistent with a defect in the clearance of apoptotic cells. Based on these observations, the following primary hypothesis is proposed. Lysosomal phospholipase A2 is necessary for the digestion and clearance of apoptotic cells by macrophages. The following specific aims are proposed to test this hypothesis: 1) To immunologically phenotype LPLA2 null mice. 2) To determine the mechanism by which the absence of lysosomal phospholipase A2 inhibits or eliminates the clearance of apoptotic cells by macrophages. 3) To determine lysosomal phospholipase A2 is either sufficient or necessary to rescue the lupus phenotype in the LPLA2 null mice. 4) To determine whether inhibition of lysosomal phospholipase A2 by hydralazine, quinidine, and procainamide is the basis for drug induced lupus. PUBLIC HEALTH RELEVANCE: We have discovered and characterized a new lysosomal phospholipase A2. Knockout mice, deficient in the activity of this enzyme, develop a late onset autoimmune phenotype with features similar to systemic lupus erythematosus. Understanding the mechanisms whereby impaired degradation of phospholipids result in the autoimmune phenotype may provide new insights into the pathogenesis of lupus and potential new targets for therapy.