CORNEAL ARACHIDONIC ACID METABOLISM

这项研究的长期目标是确定大鼠如何 角膜代谢花生四烯酸（AA）及其病理生理作用 AA 代谢物和蛋白激酶 C (PKC) 在角膜炎症中的作用， 新血管形成和再上皮化。 具体目标1将 确定氟比洛芬是否减少角膜新生血管 机制独立于角膜环氧合酶活性的抑制。 PGE2 水平将使用气相色谱/质谱法测量 (GC/MS) 作为角膜环氧合酶活性的反映。 另一个 研究将确定氟比洛芬是否抑制角膜新生血管形成 在白细胞减少的大鼠中。 这些研究将加深我们对 角膜血管生成，由于角膜血管生成减少而成为临床重要问题 视力。 具体目标2将确定是否抑制 花生四烯酸 8-或 9-脂氧合酶活性导致延迟 直径3毫米的再上皮化或器官培养的大鼠角膜 上皮擦伤。 具体目标 3 将检验 PKC 的假设 和脂氧合酶抑制剂通过阻止肌动蛋白来延迟再上皮化 聚合是上皮细胞迁移所必需的。 F-肌动蛋白 将通过上皮细胞染色的图像分析进行评估 罗丹明鬼笔环肽与抑制 PKC 和相关 花生四烯酸脂氧合酶。 具体目标 2 和 3 将有助于我们 了解调节上皮伤口愈合的机制。 具体目标4确定钙是否诱导膜 大鼠角膜12-脂氧合酶的易位。 这可能最终 提出减少角膜炎症的新药理学方法 因为 12(S)-HETE（大鼠角膜中 AA 的主要代谢物）是一种 该组织中中性粒细胞的有效趋化因子。 具体目标 5 将检验以下假设：正常、烧灼和缺氧大鼠 角膜仅合成 12(S)-HETE。 12-HETE的立体异构体是 使用放射性标记的 AA 和 GC/MS 测定。 确定哪个 12-HETE的立体异构体在不同情况下产生 至关重要，因为 12(R)-HETE 可能会影响角膜充盈度，而 12(S)- HETE 是一种白细胞趋化剂。 具体目标6将确定 PKC 是否差异调节胆碱能激活 上皮细胞中的磷脂酶 C 和 D。 这些研究可能有助于澄清 乙酰胆碱如何对角膜上皮伤口发挥有益作用 治愈。

The long-term objectives of this research are to define how the rat cornea metabolizes arachidonic acid (AA) and the pathophysiological roles of AA metabolites and protein kinase C (PKC) in corneal inflammation, neovascularization, and reepithelialization. Specific aim 1 will determine whether flurbiprofen diminishes corneal neovascularization by mechanisms independent of inhibition of corneal cyclooxygenase activity. PGE2 levels will be measured using gas chromatography/mass spectrometry (GC/MS) as a reflection of corneal cyclooxygenase activity. Another study will determine if flurbiprofen inhibits corneal neovascularization in leukopenic rats. These studies will advance our understanding of corneal angiogenesis, a clinically important problem due to reduction in visual acuity. Specific aim 2 will determine whether inhibition of arachidonate 8- or 9-lipoxygenase activity results in delayed reepithelialization or organ-cultured rat corneas with 3 mm diameter epithelial abrasions. Specific aim 3 will test the hypothesis that PKC and lipoxygenase inhibitors delay reepithelialization by preventing actin polymerization which is necessary for epithelial cell migration. F-actin will be assessed by image analysis of epithelial cells stained with rhodamine phalloidin and correlated with inhibition of PKC and arachidonate lipoxygenases. Specific aims 2 and 3 will contribute to our understanding of mechanisms regulating epithelial wound healing. Specific aim 4 will determine whether calcium induces membrane translocation of rat corneal 12-lipoxygenase. This may ultimately suggest new pharmacological approaches to reducing corneal inflammation since 12(S)-HETE, the major metabolite of AA in the rat cornea, is a potent chemotactic factor for neutrophils in this tissue. Specific aim 5 will test the hypothesis that normal, cauterized, and hypoxic rat corneas synthesize only 12(S)-HETE. The stereoisomer of 12-HETE will be determined using radiolabeled AA and by GC/MS. Determining which stereoisomer of 12-HETE is produced under different circumstances is critical since 12(R)-HETE may influence corneal turgidity, while 12(S)- HETE is a leukocyte chemoattractant. Specific aim 6 will determine whether PKC differentially regulates cholinergic activation of phospholipases C and D in the epithelium. these studies may help clarify how acetylcholine exerts a beneficial effect on corneal epithelial wound healing.