Intestinal Drug Absorption and Excretion-Detoxication Using Regulation of Membrane Permeation by Oligopeptide-produced Neutrophils

细菌产生的甲酰-甲硫基-亮氨酰-苯丙氨酸刺激中性粒细胞，扩大细胞旁途径，紧密连接(TJ)，提示寡肽可能是一种内源性吸收促进剂。在第一年，我们利用FITC-葡聚糖4000(FD-4)的通透性(Pm)和大鼠空肠、回肠和结肠的跨皮细胞电阻(Rm)的变化来检验中性粒细胞产生的活性氧化合物过氧化氢的增强作用。过氧化氢降低了所有吸收部位的跨皮电阻(Rm)和增加了Pm，表明TJ的增大在空肠和回肠显著，在粘膜侧比在浆膜侧更明显。除过氧化氢外，O2^-和OH^-参与了TJ的开放机制。第二年，以人结肠肾上腺癌细胞(Caco-2)为模型，研究了过氧化氢对TJ开放的影响。此外，在Caco-2细胞中，过氧化氢降低Rm，且在顶端的作用比在基底部的作用更显著，这表明过氧化氢在Caco-2和大鼠的肠道中具有相同的作用。此外，从P-糖蛋白(P-gp)的功能变化来考察过氧化氢对跨细胞途径的影响。P-gp具有底物从细胞到腔的分泌作用。P-gp的底物罗丹明123从粘膜到浆膜的转运增加，反向转运减少，表明P-gp功能减弱。中性粒细胞产生过氧化氢的过程分析，免疫细胞的参与，以及寡肽在口服剂型中的应用仍有待解决。

Formyl-methionyl-leucyl-phenylalanine which is produced by bacteria stimulates neutrophils and enlarges paracellular pathway, tight junction (TJ), suggesting that the oligopeptide can be an endogenous absorption enhancer. In the first year, we examined the enhancing action of the active oxygen compound which is produced by neutrophils, hydrogen peroxide, using changes in permeability (Pm) of the paracellular permeant compounds, FITC-dextran 4000 (FD-4) and the transepithelial electric resistance (Rm) in rat jejunum, ileum and colon.Hydrogen peroxide reduces Rm and increases Pm in all absorption sites, indicating the enlargement of TJ.These actions are remarkable in jejunum and ileum, and greater in the mucosal side than in the serosal side. In addition to hydrogen peroxide, O_2^- and OH^- were found to participate the TJ opening mechanism.In the second year, the effects of hydrogen peroxide were examined using human colon adrenocarcinoma (Caco-2). Also in Caco-2 cells, hydrogen peroxide decreases Rm and the action is more remarkable in the apical side than in the basal side, indicating that hydrogen peroxide has the same action in Caco-2 and the rat intestine. Further, the effects of hydrogen peroxide on the transcellular pathway were examined from the functional changes in P-glycoprotein (P-gp) which has the secretory action of the substrates from the cell to lumen. The transport of Rhodamine 123, which is a substrate of P-gp, from mucosal to serosal direction was increased but that of the reverse direction was decreased, indicating the reduction of P-gp function.The analysis of production process of hydrogen peroxide by neutrophils, the participation of immunological cell, and the application of oligopeptide to the oral dosage formulation remained to be unresolved.