STRUCTURE/FUNCTIONS OF HUMAN TEAR LIPOPHILIN

我们发现了一种新的16.5 kDa分子(脂磷脂)，它是正常人类眼泪的重要组成部分。亲脂素分子是杂二聚体，整个亚基通过分子内的3个半胱氨酸二硫键共价连接。我们的初步序列和结构数据表明，脂磷脂与前列腺素同源，前列腺素是一种由前列腺率分泌的类固醇结合蛋白。较大的脂蛋白亚基也与某些其他类固醇结合和PLA2抑制蛋白(乳巨球蛋白、子宫球蛋白和人CC-10)显示出很强的同源性，这些蛋白由乳腺、子宫和肺上皮细胞分泌。我们召集了一个经验丰富的多学科合作研究小组，以确定脂磷脂的结构和功能性质。考虑到它的同系物和泪液中相当大的脂磷脂浓度，我可能在角膜和结膜的营养、润滑和抗炎需求方面发挥关键作用。具体目标是：1)确定两个脂磷脂成分的完整氨基酸序列；确定它们的半胱氨酸如何配对，以及异源二聚体是否以非共价结合成四聚体。2)从泪腺cDNA文库中克隆每一种脂磷脂成分，确定是否有非眼组织表达脂磷素相关多肽，并确定两种脂磷脂成分基因的染色体位置。3)用免疫组织化学方法确定泪腺和其他组织中脂磷脂的储存部位。4)建立脂磷脂的酶联免疫吸附测定方法，并测定正常男女泪液中脂磷脂的浓度。5)检测人泪液脂磷脂结合具有生物学意义的脂类的能力，包括维甲酸、选定的类固醇激素和磷脂；鉴定内源性脂类，并检测其抑制正常泪液中发现的II型分泌型磷脂酶A2的能力。这项研究的长期目标是了解脂磷脂如何对角膜和结膜健康做出贡献，并探索其创造新的诊断和治疗方式的潜力。

We discovered a new 16.5 kDa molecular (lipophilin) that is a prominent component of normal human tears. Lipophilin molecules are heterodimers whole subunits are linked covalently by 3 intramolecular cystine disulfide bonds. Our preliminary sequences and structure data suggest that lipophilin is homologous to prostatein, a steroid-binding protein secrete by the rate prostate gland. The larger subunit of lipophilin also shows strong homology to certain other steroid-binding and PLA2-inhibitory proteins (mammaglobin, uteroglobin and human CC-10) that are secreted by mammary, uterine, and lung epithelial cells. We have assembled an experienced, multi-disciplinary group of co-investigators to determine the structural and functional properties of lipophilin. Both a consideration of its homologues and the considerable concentration of lipophilin in tears suggest that I may play a crucial role with respect to the nutritional lubricating and anti-inflammatory needs of the cornea and conjunctiva. The Specific Aims are: 1) To determine the complete amino acid sequences of both lipophilin components; define how their cysteines pair, and if the heterodimers associate non0covalently into tetramers. 2) To clone each lipophilin component from a lacrimal gland cDNA library; determine if any non-ocular tissues express lipophilin-related peptides; and identify the chromosomal locations of the genes for both lipophilin components. 3) To identify sites of lipophilin storage in the lacrimal gland and other tissues by immunohistochemistry. 4) To establish an ELISA assay for lipophilin and measure its concentration in the tears of normal men and women. 5) To examine the ability of human tear lipophilin to bind biologically significant lipids, including retinoids, selected steroid hormones and phospholipids; to identify endogenous lipids and to examine its ability to inhibit the Type II secretory phospholipase A2 found in normal human tears. The long-term objectives for this research are to learn how lipophilin contributes to corneal and conjuctival health, and to explore its potential for creating new diagnostic and therapeutic modalities.