Analysis of Drug Permeation Mechanism across Oral Mucosa Using Cultured Stratified Cell Layers

使用培养的分层细胞层分析药物在口腔粘膜的渗透机制

• 批准号: 15590131
• 负责人: KIMURA Toshikiro
• 金额: $ 2.37万
• 依托单位: OKAYAMA UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15590131/
• 关键词: oral mucosa; permeability evaluation; HaCaT cells; stratified cell layer; optimal culture condition; D-glucose transport

In order to clarify the drug permeation mechanism across oral mucosa, the optimal culture condition for the stratified cell layer using HaCaT cells was determined and the various basic studies were performed on glucose transport systems.As to the culture condition for HaCaT cells, DMEM EGF(+) or DMEM+Supplement EGF(+) was selected from the point of proliferative activity. From histological view point, DMEM EGF(+) was selected. From the transmucosal electrical resistance, DMEM : Ham's F-12 EGF(+) was the best. Taking factors such as cell viability, stratification, transmucosal electrical resistance and glucose transport into consideration, the best condition for HaCaT cell layers was selected to be the culture for 3 to 4 weeks using DMEM EGF(+).The result of glucose transport experiments using HaCaT cells showed that the uptake of D-glucose into cells was much faster than that of L-glucose, suggesting that the expression of stereo-selective glucose transporters in HaCaT cells. Western blot analysis showed that the presence of SGLT, GLUTs 1,2 and 3. Nevertheless, the transport rate of D-glucose across the cell layer was not significantly different from that of L-glucose. The reason for this observation is remaining unsolved.As the passively transported drugs, mannitol, melatonin and estradiol were selected, and the Papp values were measured. The results of Papp values for the three drugs were correlated with their lipophilicity, suggesting that the cultured HaCaT cell layers are effective for the evaluation of the oral-mucosal permeability to passively transported drugs.

为了阐明口服粘膜的药物渗透机制，确定了使用HACAT细胞进行分层细胞层的最佳培养条件，并对葡萄糖传输系统进行了各种基础研究。 +）或DMEM+补充EGF（+）是从增殖活性点中选择的。从组织学角度来看，选择了DMEM EGF（+）。从透射电阻中，DMEM：HAM的F-12 EGF（+）是最好的。考虑使用DMEM EGF（+），选择了诸如细胞活力，分层，透射电阻和葡萄糖转运等因素为3至4周的培养物。 HACAT细胞表明，D-葡萄糖对细胞的摄取比L-葡萄糖的摄取快得多，这表明立体选择性葡萄糖转运蛋白在HACAT细胞中的表达表达。蛋白质印迹分析表明，SGLT，GLUT 1,2和3的存在。然而，跨细胞层的D-葡萄糖的转运速率与L-葡萄糖的转运率没有显着差异。该观察结果的原因仍未解决。选择被动运输的药物，甘露醇，褪黑激素和雌二醇，并测量了PAPP值。这三种药物的PAPP值的结果与它们的亲脂性相关，这表明培养的HACAT细胞层有效评估口腔粘膜渗透性对被动运输的药物。