Evidence for a Specific Uptake and Retention Mechanism for 25-Hydroxyvitamin D (25OHD) in Skeletal Muscle Cells
Evidence for a Specific Uptake and Retention Mechanism for 25-Hydroxyvitamin D (25OHD) in Skeletal Muscle Cells
复制标题
DOI：
10.1210/en.2012-2245
复制
发表时间：
2013-09-01
期刊：
影响因子：
4.8
通讯作者：
Mason, R. S.
中科院分区：
文献类型：
--
作者：
Abboud, M.;Puglisi, D. A.;Mason, R. S.
关于25-羟基维生素D（25 OHD）在血液中停留时间延长的机制知之甚少。一些证据使我们提出骨骼肌可以作为25 OHD血管外池的部位。在体外研究中，研究了分化的C2小鼠肌肉细胞摄取和释放25 OHD的能力，与其他细胞类型和膜蛋白巨蛋白在这些机制中的参与进行比较。当C2细胞分化为肌管时，标记的25 OHD的时间依赖性摄取比未分化的成肌细胞或非肌成骨细胞MG 63细胞高2-3倍（P <0.001）。在体外释放实验中（25 OHD摄取后），肌管在4小时后仅释放32% +/- 6%的储存25 OHD，而成骨细胞的这一数字为60% +/- 2%（P <0.01）。使用免疫荧光，C2肌管和原代大鼠肌纤维，第一次，显示表达megalin和cubilin，维生素D结合蛋白（DBP），结合几乎所有的25 OHD在血液中的内吞受体。DBP对骨骼肌中的肌动蛋白具有高亲和力。通过共聚焦显微镜观察到Alexandrin或-488标记的DBP进入成熟肌细胞的时间依赖性摄取。将C2肌管与受体相关蛋白（一种巨蛋白抑制剂）孵育（24小时），导致25 OHD摄取减少40%（P <0.01）。这些数据支持的建议，25 OHD，摄取到成熟的肌肉细胞后，是由DBP，它已通过膜巨蛋白内化，并通过结合肌动蛋白保留。
Little is known about the mechanism for the prolonged residence time of 25-hydroxyvitamin D (25OHD) in blood. Several lines of evidence led us to propose that skeletal muscle could function as the site of an extravascular pool of 25OHD. In vitro studies investigated the capacity of differentiated C2 murine muscle cells to take up and release 25OHD, in comparison with other cell types and the involvement of the membrane protein megalin in these mechanisms. When C2 cells are differentiated into myotubes, the time-dependent uptake of labeled 25OHD is 2-3 times higher than in undifferentiated myoblasts or nonmuscle osteoblastic MG63 cells (P < .001). During in vitro release experiments (after 25OHD uptake), myotubes released only 32% +/- 6% stored 25OHD after 4 hours, whereas this figure was 60% +/- 2% for osteoblasts (P < .01). Using immunofluorescence, C2 myotubes and primary rat muscle fibers were, for the first time, shown to express megalin and cubilin, endocytotic receptors for the vitamin D binding protein (DBP), which binds nearly all 25OHD in the blood. DBP has a high affinity for actin in skeletal muscle. A time-dependent uptake of Alexafluor-488-labeled DBP into mature muscle cells was observed by confocal microscopy. Incubation of C2 myotubes (for 24 hours) with receptor-associated protein, a megalin inhibitor, led to a 40% decrease in 25OHD uptake (P < .01). These data support the proposal that 25OHD, after uptake into mature muscle cells, is held there by DBP, which has been internalized via membrane megalin and is retained by binding to actin.