Dische Z, Cremer-Bartels G, Kaye G I
Proc Natl Acad Sci U S A. 1985 Feb;82(3):760-4. doi: 10.1073/pnas.82.3.760.
Glycosaminoglycans (GAG) of bovine cornea were sequentially extracted by 0.15 M NaCl and by 1 M CaCl2, pH 8. The amounts of hexosamine (HexN) and hexuronic acid (HexUA), specific hexoses (Hex), and protein were determined in the extracts. The ultrastructure of the corneal stroma after NaCl and NaCl/CaCl2 extraction was also studied. Approximately 70% of the Hex-HexN extractable by 0.15 M NaCl is removed in the first NaCl extract, with the amount decreasing rapidly to the fifth NaCl extract. Only 20-30% of the extractable HexUA-HexN is removed in the first NaCl extract; subsequent extraction removes successively less HexUA-HexN. There is a sharp increase, however, in both Hex-HexN and HexUA-HexN removed in the first 1 M CaCl2 extract, ranging from 25 to 40% of the total extractable with NaCl. The stroma retained lamellar organization and normal spacing, diameter, and length of collagen fibers after NaCl extraction. Visibility of the 64-nm periodicity was enhanced because of loss of density in the ground substance. CaCl2 extraction led to loss of lamellar organization and apparent disruption of the collagen fibers. Only short thin fibrils remained, embedded in a material having the density and very fine fibrillar organization of basal lamina. Disruption of the collagen fibers is probably due to removal of a specific GAG component necessary to maintain the collagen as an extended morphologic structure but may also be due to a specific degradative effect of CaCl2 on collagen no longer covered by a protective layer of GAG and glycoproteins.
用0.15M NaCl和pH为8的1M CaCl₂依次提取牛角膜的糖胺聚糖（GAG）。测定提取物中己糖胺（HexN）、己糖醛酸（HexUA）、特定己糖（Hex）和蛋白质的含量。还研究了NaCl和NaCl/CaCl₂提取后角膜基质的超微结构。0.15M NaCl可提取的Hex-HexN中约70%在第一次NaCl提取物中被去除，其含量迅速下降至第五次NaCl提取物。第一次NaCl提取物中仅去除20%-30%可提取的HexUA-HexN；随后的提取依次去除较少的HexUA-HexN。然而，在第一次1M CaCl₂提取物中去除的Hex-HexN和HexUA-HexN均急剧增加，占NaCl可提取总量的25%至40%。NaCl提取后，基质保留了板层结构以及胶原纤维的正常间距、直径和长度。由于基质密度降低，64nm周期性的可见性增强。CaCl₂提取导致板层结构丧失和胶原纤维明显破坏。仅残留短而细的纤维，嵌入具有基底层密度和非常细的纤维组织的物质中。胶原纤维的破坏可能是由于去除了维持胶原作为伸展形态结构所需的特定GAG成分，但也可能是由于CaCl₂对不再被GAG和糖蛋白保护层覆盖的胶原的特定降解作用。
