Iozzo R V, Clark C C
Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine, Philadelphia 19104.
Histochemistry. 1987;88(1):23-9. doi: 10.1007/BF00490162.
In addition to containing Type IV collagen, laminin and entactin, basement membranes contain small amounts of proteoglycans substituted primarily with heparan sulfate chains. We have previously shown, however, that parietal yolk sacs in organ culture synthesize predominantly chondroitin sulfate proteoglycan. In the present study, we have used histochemical and immunohistochemical techniques coupled with chondroitinase ABC digestion to provide evidence for the presence of chondroitin sulfate proteoglycan in the basement membrane (Reichert's membrane) of the 14.5-day rat embryo parietal yolk sac. The results revealed numerous cuprolinic blue-positive filaments and granules, 20-30 nm in greater length or diameter, dispersed throughout the thickness of the basement membrane. Both structures were removed by preincubating freshly isolated parietal yolk sacs with chondroitinase ABC. A similar labeling pattern was also obtained with immunoelectron microscopy using gold-labeled monoclonal antibodies directed against the three major isomers of protein-bound chondroitin sulfate. In contrast, coarser cuprolinic blue granules, 40-100 nm in diameter, were neither sensitive to chondroitinase ABC digestion nor labeled by the monoclonal antibodies. These results thus indicate that Reichert's membrane contains chondroitin sulfate proteoglycan in addition to heparan sulfate proteoglycan.
除了含有IV型胶原蛋白、层粘连蛋白和巢蛋白外，基底膜还含有少量主要被硫酸乙酰肝素链取代的蛋白聚糖。然而，我们之前已经表明，器官培养中的壁层卵黄囊主要合成硫酸软骨素蛋白聚糖。在本研究中，我们使用组织化学和免疫组织化学技术并结合软骨素酶ABC消化，来为14.5天大鼠胚胎壁层卵黄囊的基底膜（赖歇特膜）中存在硫酸软骨素蛋白聚糖提供证据。结果显示，在整个基底膜厚度中分散着许多铜蓝蛋白阳性细丝和颗粒，其长度或直径较大，为20 - 30纳米。通过用软骨素酶ABC预孵育新鲜分离的壁层卵黄囊，这两种结构都被去除了。使用针对与蛋白质结合的硫酸软骨素的三种主要异构体的金标记单克隆抗体进行免疫电子显微镜检查，也获得了类似的标记模式。相比之下，直径为40 - 100纳米的较粗铜蓝蛋白颗粒既对软骨素酶ABC消化不敏感，也未被单克隆抗体标记。因此，这些结果表明，赖歇特膜除了含有硫酸乙酰肝素蛋白聚糖外，还含有硫酸软骨素蛋白聚糖。
