Epithelial secretion of vinblastine by human intestinal adenocarcinoma cell (HCT-8 and T84) layers expressing P-glycoprotein.
Epithelial secretion of vinblastine by human intestinal adenocarcinoma cell (HCT-8 and T84) layers expressing P-glycoprotein.
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DOI：
10.1038/bjc.1991.328
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发表时间：
1991-09
影响因子：
8.8
通讯作者：
Simmons, N L
中科院分区：
文献类型：
--
作者：
Hunter, J;Hirst, B H;Simmons, N L
通过用单克隆抗体JSB-1免疫沉淀170-180 kDa蛋白，证明了两种人肠腺癌细胞系（HCT-8，回盲部和T84，结肠）中P-糖蛋白的表达。当在可渗透基质上生长时，HCT-8和T84均形成高跨上皮电阻（大于700 Ω cm 2）的功能性上皮细胞层。这些上皮层表现出载体分泌（在HCT-8和T84细胞层中，从含10 nM长春碱的浴液中，从基底到顶端方向的净长春碱通量分别为0.135和0.452 pmol h-1 cm-2）。这些载体长春碱分泌物对维拉帕米的抑制敏感。长春碱被动经上皮渗透受限于细胞间（紧密）连接的存在，如高经上皮电阻所示，维拉帕米增加了这种被动长春碱渗透，同时降低了电阻。细胞长春碱负荷从基底侧显著更大，这也容易受到基础维拉帕米的抑制。长春碱在人肠结肠腺癌细胞层中的载体转运的证明是支持以下假设的直接证据：胃肠道上皮中mdr 1的功能是通过上皮分泌过程促进解毒。本研究还强调，细胞长春碱蓄积不仅取决于P-糖蛋白功能，还取决于不同的表观膜渗透性和细胞间（紧密）连接的存在，这些连接可能限制药物渗透和细胞蓄积至顶端或基底膜结构域。
P-glycoprotein expression was demonstrated in two human intestinal adenocarcinoma cell-lines (HCT-8, ileocaecal and T84, colonic) by immunoprecipitation of a 170-180 kDa protein with monoclonal antibody JSB-1. Both HCT-8 and T84 formed functional epithelial cell layers of high transepithelial electrical resistance (greater than 700 omega.cm2) when grown on permeable matrices. These epithelial layers demonstrated vectorial secretion (net vinblastine fluxes in the basal-to-apical direction of 0.135 and 0.452 pmol h-1 cm-2 in HCT-8 and T84 cell layers, respectively, from bathing solutions containing 10 nM vinblastine). These vectorial vinblastine secretions were sensitive to inhibition by verapamil. Passive transepithelial vinblastine permeation was limited by the presence of intercellular (tight) junctions, as demonstrated by the high transepithelial electrical resistance, and verapamil increased this passive vinblastine permeation concomitant with a reduction in the electrical resistance. Cellular vinblastine loading was significantly greater from the basal side, and this was also susceptible to inhibition by basal verapamil. The demonstration of vectorial transport of vinblastine in human intestinal colonic adenocarcinoma cell layers is direct evidence in favour of the hypothesis that the function of mdr1 in epithelial from the gastrointestinal tract is to promote detoxification by a process of epithelial secretion. This study also highlights that cellular vinblastine accumulation depends not only upon P-glycoprotein function, but also upon differential apparent membrane permeabilities and the presence of intercellular (tight) junctions that may restrict drug permeation and cellular accumulation to apical or basal membrane domains.