Konings P N, Mania-Farnell B, Beinfeld M C, Oakes M G, Day R, Seidah N G, Davis T P
Department of Pharmacology, College of Medicine, University of Arizona, Tucson 85724.
Neuropeptides. 1993 Jul;25(1):19-30. doi: 10.1016/0143-4179(93)90064-h.
The human cholinergic neuroepithelioma cell line SK-N-MCIXC, which expresses high levels of cholecystokinin (CCK) mRNA and secretes intact CCK into the media, was used to examine CCK processing and metabolism. Our data provide evidence for the existence of specific candidate processing enzymes in SK-N-MCIXC cells which may be involved in processing proCCK in the brain and indicate that SK-N-MCIXC cells provide a model system for studying the regulation of these enzymes. mRNAs for the intracellular processing enzymes, prohormone convertase 1 (PC1), PC2 and furin were present in SK-N-MCIXC cells. PC1 and/or PC2 and/or furin may cleave at the dibasic amino acid pairs Arg-Arg at the C-terminal part of proCCK, and Arg-X-X-Arg at the N-terminal of the CCK-58 sequence in proCCK. The SK-N-MCIXC cell line demonstrated spontaneous and regulated release of CCK and large amounts of CCK-precursors, as measured with region specific radioimmunoassays coupled to high performance liquid chromatography. Storage granules containing glycine-extended CCK were shown in SK-N-MCIXC cells using indirect immunofluorescence. The extracellularly localized CCK-metabolizing enzyme, neutral endopeptidase 24.11 (EC 3.4.24.11), was present in membranes from both SK-N-MCIXC cells and in intact slices of rat cerebral cortex. The rat cerebral cortex is a brain region known to be rich in CCK. The SK-N-MCIXC cell line provides an in vitro model to study the regulation of CCK synthesis and metabolism in neuronal systems since it contains the storage granules, mRNA, intact peptide, and complement of enzymes necessary for biosynthesis and metabolism of CCK.
人胆碱能神经上皮瘤细胞系SK-N-MCIXC表达高水平的胆囊收缩素（CCK）mRNA，并将完整的CCK分泌到培养基中，被用于研究CCK的加工和代谢。我们的数据为SK-N-MCIXC细胞中存在可能参与脑中前CCK加工的特定候选加工酶提供了证据，并表明SK-N-MCIXC细胞为研究这些酶的调节提供了一个模型系统。细胞内加工酶前激素转化酶1（PC1）、PC2和弗林蛋白酶的mRNA存在于SK-N-MCIXC细胞中。PC1和/或PC2和/或弗林蛋白酶可能在proCCK C末端的双碱性氨基酸对Arg-Arg处以及proCCK中CCK-58序列N末端的Arg-X-X-Arg处进行切割。使用与高效液相色谱联用的区域特异性放射免疫测定法测量，SK-N-MCIXC细胞系表现出CCK和大量CCK前体的自发和调节性释放。使用间接免疫荧光法在SK-N-MCIXC细胞中显示出含有甘氨酸延伸型CCK的储存颗粒。细胞外定位的CCK代谢酶中性内肽酶24.11（EC 3.4.24.11）存在于SK-N-MCIXC细胞的膜以及大鼠大脑皮层的完整切片中。大鼠大脑皮层是已知富含CCK的脑区。SK-N-MCIXC细胞系提供了一个体外模型，用于研究神经元系统中CCK合成和代谢的调节，因为它包含储存颗粒、mRNA、完整肽以及CCK生物合成和代谢所需的酶补充。
