ULTRAVIOLET LIGHT INDUCED PHENOMENA IN BIOMEMBRANES

细胞膜中的光化学负责启动许多 细胞对有无紫外线和可见光辐射的反应 添加了光敏分子。质膜和线粒体 膜通常是生物体中主要的光敏化部位 使用染料加可见光辐射治疗肿瘤(光动力疗法， PDT)。这个应用程序长期目标是提供一个 膜系感光染料设计的合理依据 在医学和生物学方面的应用。这项建议的具体目的 包括： 具体目标1：分别生产单线态氧(1O2*)和游离态 来自质膜中相同光敏剂分子的自由基。 单线态氧将使用低强度光产生；自由基将 使用脉冲激光来生产。数学建模的能力 预测光敏化的最佳激光强度和波长 将对单元格进行评估。 具体目标2：确定地点和身份的影响 (1O2*或自由基)的主要反应物种在 血浆中生化变化的光敏作用 薄膜。将使用在不同部位定位的光敏剂 在质膜上或产生不同的活性物种。变化 在膜功能之间将进行定量比较 光敏剂。需要测量的膜功能包括：脯氨酸 摄取，Na，K-ATPase活性，膜通透性，细胞内 Ca2和膜电位。血浆变化的动力学 膜电位和细胞内钙离子将在 用荧光激发激光后的微秒到分钟时间尺度 记者染发。 具体目标3：确定细胞对膜的反应 光敏化反应反映了形成的身份或位置 活性物种。膜相关酶的激活， 磷脂酶A2和C，将在光敏化后用 在质膜中产生1O2*或自由基的染料。 早期反应基因的诱导将被用来测量 活性物种对等离子体早期核反应的影响 膜光敏化。测量将包括诱导c- Jun、c-fos和c-myc基因的表达。光敏剂诱导细胞凋亡的实验研究 将对造成质膜损伤的因素进行评估。之后 光敏作用下，DNA将被分离并裂解成核小体- 大小的碎片将被量化。

Photochemistry in cellular membranes is responsible for initiating many responses of cells to ultraviolet and visible radiation with and without added photosensitizing molecules. The plasma membrane and mitochondrial membranes are frequently the sites of primary photosensitization in the use of dyes plus visible radiation to treat tumors (photodynamic therapy, PDT). The long range objective of this application is to provide a rational basis for design of membrane-photosensitizing dyes for applications in medicine and biology. The specific aims of this proposal are: Specific Aim 1: To produce, separately, singlet oxygen (1O2*) and free radicals from the same photosensitizer molecules in the plasma membrane. Singlet oxygen will be produced using low intensity light; radicals will be produced using pulsed lasers. The ability of mathematical modeling to predict optimal laser intensity and wavelength for photosensitization of cells will be evaluated. Specific Aim 2: To establish the influence of the location and identity (1O2* or free radicals) of the primary reactive species formed during photosensitization on biochemical alterations produced in the plasma membrane. Photosensitizers will be used that localize at different sites in the plasma membrane or generate different reactive species. Changes in membrane functions will be quantitatively compared between photosensitizers. Membrane functions to be measured include proline uptake, activity of Na+,K+-ATPase, membrane permeability, intracellular Ca2+, and membrane potential. The kinetics for alteration of plasma membrane potential and intracellular Ca2+ will be monitored on a microsecond to minute time scale after laser excitation using fluorescent reporter dyes. Specific Aim 3: To determine whether cellular responses to membrane photosensitization reflect the identity or location of formation of the reactive species. The activation of the membrane-associated enzymes, phospholipases A2 and C, will be assessed after photosensitization with dyes producing either 1O2* or free radicals in the plasma membrane. Induction of early response genes will be used as used to measure the effects of reactive species on an early nuclear response to plasma membrane photosensitization. Measurements will include induction of c- jun, c-fos and c-myc mRNA. Induction of apoptosis by photosensitizers that cause plasma membrane damage will be assessed. After photosensitization, DNA will be isolated and cleavage into nucleosome- size fragments will be quantitated.