有机阴离子转运多肽OATP1A2介导芬太尼的转运  

ORGANIC ANION-TRANSPORTING POLYPEPTIDE 1A2 MEDIATES THE TRANSPORT OF FENTANYL

作　　者：杨惠雯[1] 张帆[1] 黄利华[2] 韩睿[1] 欧阳文[1] 向红[2] 廖琴[1] 

机构地区：[1]中南大学湘雅三医院麻醉科,长沙410013 [2]中南大学湘雅三医院实验中心,长沙410013

出　　处：《中国疼痛医学杂志》2017年第7期494-501,共8页Chinese Journal of Pain Medicine

基　　金：湖南省自然科学基金(16JJ2158)

摘　　要：目的:建立稳定表达有机阴离子转运多肽1A2(organic anion-transporting polypeptide 1A2,OATP1A2)的HEK293细胞株,体外研究OATP1A2是否转运芬太尼(Fentanyl)。方法:用脂质体转染法将pIRES2-Zs Green1-OATP1A2质粒转染至HEK293细胞中,G418(600 mg/ml)筛选单克隆阳性细胞,采用Western blot、RT-PCR证实HEK293-OATP1A2构建成功;用不同浓度的探针药物非索非那定(Fexofenadine,FEX)验证HEK293-OATP1A2的转运功能;运用不同浓度的芬太尼孵育细胞,另取两组用相同浓度的芬太尼孵育的细胞,分别加或不加抑制剂柚皮素(Naringenin,100μg/m L)进行HEK293-OATP1A2转运实验。结果:FEX浓度为1、10、100 nM时HEK293-OATP1A2实验组与HEK293-VC对照组,FEX的吸收值均有显著的统计学差异(P<0.05),且FEX浓度为100 nM时,实验组对FEX的吸收值是对照组的2.8倍;加入抑制剂柚皮素组FEX吸收值较不加抑制剂时减少了66.8±0.6%。芬太尼转运结果显示,芬太尼浓度为1、10、100、1000 nM时实验组与对照组比较,芬太尼的吸收值均有显著的统计学差异(P<0.05),且芬太尼浓度为1000 nM时,实验组对芬太尼的吸收值是对照组的2.2倍;加入抑制剂组芬太尼的吸收值较未加抑制剂组减少了86.5±0.5%(P<0.05)。结论:成功建立了稳定表达OATP1A2的HEK293细胞株,体外研究发现OATP1A2能介导芬太尼的转运。Objective： To establish human embryonic kidney （HEK293） cell line stably expressing human OATP1A2, and to explore whether the uptake of fentanyl can be mediated by OATP1A2 in vitro. Methods： HEK293 cells were transfected with the plasmid pIRES2-ZsGreenl-OATP I A2 using a Liposome transfection reagent LipofectamineTM 2000. After geneticin （G-418; 600 mg/ml） treatment, single colonies were selected and characterized. Western blot analysis and real-time PCR were used to verify the success of transfection. Different concentrations of probe drug fexofenadine was used to verify HEK293-OATP1A2 transport function. The cells were incubated with different concentrations of fentanyl. HEK293-OATP 1A2 uptake experiment was performed with inhibitor naringenin. Results： Fexofenadine and fentanyl showed a significantly increased uptake （P 〈 0.05） into the HEK293-OATP1A2 compared with the HEK293-VC cells. The uptake value of Fexofenadine at 100 nM was 2.8-fold to that of control. The OATP1A2 inhibitor naringenin at a concentration of 100 μg/mL significantly reduced the cellular fexofenadine and fentanyl net uptake to 66,8 4-0.6% and 86.5± 0.5% of control （fexofenadine uptake without inhibitor）, respectively （P 〈 0.05）. Conclusion： HEK293 cell line with overexpression of OATP1A2 was successfully constructed, and OATP1A2 mediatinR fentanyl transport in vitro has been confirmed.

关 键 词：有机阴离子转运体多肽1A2 芬太尼 底物 转运功能 过表达 

分 类 号：R614[医药卫生—麻醉学]