Oates P S, Thomas C, Morgan E H
Department of Physiology, The University of Western Australia, Perth.
J Gastroenterol Hepatol. 1997 Dec;12(12):829-38. doi: 10.1111/j.1440-1746.1997.tb00380.x.
The intestinal mucosa is characterized by cell proliferation, commitment, differentiation, digestion and absorption. These processes occur at specified locations along the crypt to villus axis. A technique is reported for the isolation of cells along this axis which allows the study of any one of these processes in an enriched population of cells. As an example, the uptake of transferrin-bound iron by enterocytes was studied. Rats were fed diets normal, high (30% carbonyl iron) or low in iron for 12 days. Cells from either the duodenum or ileum were isolated by incubating in a Ca(2+)-, Mg2+-free, cation chelating solution for varying periods. The incorporation of thymidine into DNA was measured in these cells as a marker of the crypt region, while alkaline phosphatase and sucrase activities marked mature enterocytes. The in vivo uptake of transferrin-bound 59Fe was measured in cells isolated either 2 or 4 h after intravenous injection. This procedure resulted in the isolation of 10 fractions of viable cells. Earlier fractions were enriched at least 10-fold in villus cells and the last fractions in crypt cells. Cells in intermediate fractions were at various stages of development. Uptake of transferrin-bound iron into enterocytes was highest with feeding an iron-loaded diet compared with control or iron-deficient diets. However, with all diets uptake was highest in crypt cells and this fell at the crypt-villus junction to be only 25%, as high at the villus tip as the crypt. A technique for the reproducible isolation of viable enterocytes along a crypt-villus axis is described. Transferrin receptor activity changes with maturation of the enterocyte.
肠黏膜的特点是细胞增殖、定向分化、消化和吸收。这些过程沿着隐窝至绒毛轴在特定位置发生。本文报道了一种沿此轴分离细胞的技术，该技术可在富集的细胞群体中研究这些过程中的任何一个。例如，研究了肠细胞对转铁蛋白结合铁的摄取。给大鼠喂食正常、高铁（30%羰基铁）或低铁饮食12天。通过在无钙、无镁的阳离子螯合溶液中孵育不同时间，从十二指肠或回肠分离细胞。在这些细胞中测量胸苷掺入DNA的情况，作为隐窝区域的标志物，而碱性磷酸酶和蔗糖酶活性则标志着成熟的肠细胞。在静脉注射后2或4小时分离的细胞中测量转铁蛋白结合的59Fe的体内摄取。该程序导致分离出10个活细胞组分。较早的组分中绒毛细胞富集至少10倍，最后的组分中隐窝细胞富集。中间组分的细胞处于不同的发育阶段。与对照或缺铁饮食相比，喂食高铁饮食时肠细胞对转铁蛋白结合铁的摄取最高。然而，在所有饮食中，隐窝细胞中的摄取最高，在隐窝 - 绒毛交界处下降，仅为绒毛尖端的25%，与隐窝处一样高。描述了一种沿隐窝 - 绒毛轴可重复分离活肠细胞的技术。转铁蛋白受体活性随肠细胞成熟而变化。
