Characterization of a factor that promotes neurite outgrowth: evidence linking activity to a heparan sulfate proteoglycan.
Characterization of a factor that promotes neurite outgrowth: evidence linking activity to a heparan sulfate proteoglycan.
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DOI：
10.1083/jcb.94.3.574
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发表时间：
1982-09
期刊：
影响因子：
--
通讯作者：
Reichardt LF
中科院分区：
文献类型：
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作者：
Lander AD;Fujii DK;Gospodarowicz D;Reichardt LF
大鼠交感神经元，平板培养的牛角膜内皮细胞产生的细胞外基质，迅速延长神经突起的神经生长因子（NGF）的情况下。该反应不受NGF抗血清的影响。快速生长也发生时，交感神经元铺在聚赖氨酸涂层表面已暴露于无血清培养基条件下的角膜内皮细胞（CMSF）。即使在没有血清的情况下培养神经元，也可以看到反应。当铺在用CMSF处理过的多聚赖氨酸包被的培养皿上时，只有处理过的一半上的神经元延长了神经突。CMSF中的活性因子被胰蛋白酶、酸（pH 1.6）、碱（pH 12.7）或加热至80 ℃破坏;它对加热至60 ℃、胶原酶、脱氧核糖核酸酶和神经氨酸酶稳定。该因子在Sepharose 6 B色谱柱的空隙体积后立即洗脱。在缔合氯化铯梯度中，它作为以1.36-1.37的密度为中心的峰沉淀，对应于可以用[35 S]硫酸盐或[3 H]亮氨酸生物合成标记的材料的峰。来自该级分的材料被肝素酶灭活，但不被软骨素酶ABC灭活，这意味着硫酸肝素蛋白聚糖对该因子的活性是必需的。肝素酶制剂中的污染物灭活被排除。进一步纯化表明，活性因子可能以含有硫酸肝素蛋白聚糖和其他分子的聚集体形式存在。CMSF也促进其他类型神经元的轴突生长。此外，多种细胞类型显示出产生与CMSF中类似的因子。
Rat sympathetic neurons, plated onto extracellular matrix produced by cultured bovine corneal endothelial cells, rapidly extended neurites in the absence of nerve growth factor (NGF). The response was unaffected by antiserum to NGF. Rapid outgrowth also occurred when sympathetic neurons were plated onto polylysine-coated surfaces that had been exposed to serum-free medium conditioned by corneal endothelial cells (CMSF). A response was seen even when the neurons were cultured without serum. When plated onto a polylysine-coated dish treated with CMSF over half its surface, only the neurons on the treated half extended neurites. The active factor in CMSF was destroyed by trypsin, acid (pH 1.6), base (pH 12.7), or heating to 80 degrees C; it was stable to heating to 60 degrees C, collagenase, deoxyribonuclease, and neuraminidase. The factor elutes just after the void volume of a Sepharose 6B column. In associative cesium chloride gradients, it sediments as a peak centered at a density of 1.36-1.37, corresponding to a peak of material that can be biosynthetically labeled with [35S]sulfate or [3H]leucine. Material from this fraction was inactivated by heparinase, but not chondroitinase ABC, implying that a heparin sulfate proteoglycan is essential for the factor's activity. Inactivation by contaminants in the heparinase preparation was ruled out. Further purification indicated that the active factor may exist as an aggregate containing a heparin sulfate proteoglycan and other molecules. CMSF also promoted neurite outgrowth by other types of neurons. Furthermore, a variety of cell types were shown to produce factors similar to that in CMSF.