Gutowski K A, Hudson J L, Aminoff D
Department of Biological Chemistry, University of Michigan, Ann Arbor 48109-2007.
Exp Gerontol. 1991;26(4):315-26. doi: 10.1016/0531-5565(91)90044-m.
A recent review (Aminoff, 1988) summarized the evidence for and against our hypothesis for the role of glycophorin in the senescence and clearance of mammalian red blood cells (RBC) from circulation. This hypothesis postulates the loss of sialic acid from RBC surface in two forms: (a) as vesicles containing the sialoglycoprotein glycophorin, and (b) as free sialic acid residues from glycophorin molecules remaining on cell surface. In this report we demonstrate the applicability of flow cytometric procedures to explore, at the cellular level, time-dependent changes on RBC surface with change in cell size, and with in vivo age. The RBC are probed with fluorescein isothiocyanate (FITC) labelled lectins and goat anti-human-IgG and -IgM. The relative intensity of fluorescence is correlated to the change in RBC size as measured by forward lightscatter. Reactivity of RBC with FITC-labelled wheat germ agglutinin can be inhibited with either 0.2M N-acetylglucosamine or by removal of sialic acid residues with neuraminidase. The properties of the smallest RBC correspond to those of the oldest RBC in their: (a) decreased reactivity with FITC-labelled lectins that recognize sialic acid residues, wheat germ and Limax flavus agglutinins, and (b) increased reactivity with FITC-labelled goat anti-human-IgG and -IgM. These results are compatible with our glycophorin hypothesis. Moreover, they suggest that the initial loss of sialic acid as glycophorin containing vesicles is gradual, while the subsequent step involving the loss of sialic acid residues is rapid and exposes multiple disaccharide galactose beta(1-3)N-acetylgalacosaminyl residues. These unmasked disaccharide sites are recognized by autoimmune IgG, IgM, and lectin-like receptors on macrophages resulting in the clearance of senescent RBC from circulation.
最近的一篇综述（Aminoff，1988年）总结了支持和反对我们关于血型糖蛋白在哺乳动物红细胞（RBC）衰老及从循环中清除过程中所起作用这一假说的证据。该假说假定红细胞表面唾液酸以两种形式丢失：（a）作为含有唾液酸糖蛋白血型糖蛋白的囊泡，以及（b）作为残留在细胞表面的血型糖蛋白分子上的游离唾液酸残基。在本报告中，我们证明了流式细胞术程序在细胞水平上探索红细胞表面随细胞大小变化以及随体内年龄变化的时间依赖性变化的适用性。用异硫氰酸荧光素（FITC）标记的凝集素以及山羊抗人IgG和IgM对红细胞进行检测。荧光的相对强度与通过前向光散射测量的红细胞大小变化相关。红细胞与FITC标记的麦胚凝集素的反应性可被0.2M N - 乙酰葡糖胺抑制，或通过用神经氨酸酶去除唾液酸残基来抑制。最小的红细胞在以下方面的特性与最老的红细胞相对应：（a）与识别唾液酸残基的FITC标记凝集素、麦胚凝集素和黄斑海蜗牛凝集素的反应性降低，以及（b）与FITC标记的山羊抗人IgG和IgM的反应性增加。这些结果与我们的血型糖蛋白假说相符。此外，它们表明作为含血型糖蛋白囊泡的唾液酸的初始丢失是渐进的，而随后涉及唾液酸残基丢失的步骤是快速发生的，并且暴露出多个二糖半乳糖β(1 - 3)N - N - 乙酰半乳糖胺残基。这些暴露的二糖位点被巨噬细胞上的自身免疫性IgG、IgM和凝集素样受体识别，从而导致衰老红细胞从循环中清除。
