Hussaini I M, Srikumar K, Quesenberry P J, Gonias S L
Department of Pathology, University of Virginia Health Sciences Center, Charlottesville 22908.
J Biol Chem. 1990 Nov 15;265(32):19441-6.
Primary cultures of murine bone marrow macrophages (BMMs) were prepared from marrow cell suspensions. These cells expressed specific receptors that recognized the transformed conformation of human alpha 2-macroglobulin (alpha 2M) generated by reaction with CH3NH2. alpha 2M receptor expression was regulated by colony-stimulating factor-1 (CSF-1). The BMMs were deprived of CSF-1 for 6 h and then treated with different concentrations of the purified cytokine. After 18 h, binding of 125I-alpha 2M-CH3NH2 was examined at 4 degrees C. Analysis of the saturation isotherms and Scatchard transformations indicated that the KD was not affected by CSF-1 (1.9-2.4 nM), whereas the maximum specific radioligand binding capacity (Bmax) was increased from 5.6 x 10(4) receptors/cell in the absence of CSF-1 to 2.2 x 10(5) and 2.6 x 10(5) receptors/cell for BMMs treated with 1,000 and 10,000 units/ml CSF-1, respectively. The difference in total cellular protein after exposure to different levels of CSF-1 for 18 h was small (1.50-1.92 ng/cell) and not statistically significant. A 6-12-h lag phase was identified between the time of CSF-1 exposure and increased alpha 2M receptor expression. Cycloheximide completely blocked the increase in alpha 2M receptor expression when added simultaneously with the CSF-1; greater than 50% inhibition was observed when the cycloheximide was added up to 8 h later. The RNA synthesis inhibitors, actinomycin D and daunomycin, prevented increased alpha 2M receptor expression when added up to 4 h after the CSF-1, but had no effect at 8 h. At 37 degrees C, uptake and digestion of 125I-alpha 2M-CH3NH2 was increased in BMMs treated with 1,000 units/ml CSF-1 for 18 h compared with untreated cells. These studies demonstrate that CSF-1 increases the expression of alpha 2M receptors in BMMs through a pathway that requires new RNA and protein synthesis. We hypothesize that increased alpha 2M receptor expression may play an important role in cellular growth and differentiation.
小鼠骨髓巨噬细胞（BMMs）的原代培养物是从骨髓细胞悬液中制备的。这些细胞表达特定的受体，该受体能识别通过与CH3NH2反应产生的人α2-巨球蛋白（α2M）的转化构象。α2M受体的表达受集落刺激因子-1（CSF-1）调控。将BMMs去除CSF-1 6小时，然后用不同浓度的纯化细胞因子处理。18小时后，在4℃检测125I-α2M-CH3NH2的结合情况。饱和等温线和Scatchard转换分析表明，解离常数（KD）不受CSF-1影响（1.9 - 2.4 nM），而最大特异性放射性配体结合容量（Bmax）从无CSF-1时的5.6×104个受体/细胞，分别增加到用1000和10000单位/ml CSF-1处理的BMMs的2.2×105和2.6×105个受体/细胞。在暴露于不同水平的CSF-1 18小时后，总细胞蛋白的差异很小（1.50 - 1.92 ng/细胞），且无统计学意义。在CSF-1暴露时间和α2M受体表达增加之间确定了6 - 12小时的延迟期。当与CSF-1同时添加时，放线菌酮完全阻断了α2M受体表达的增加；当放线菌酮在8小时后添加时，观察到超过50%的抑制作用。RNA合成抑制剂放线菌素D和柔红霉素在CSF-1后4小时内添加时可阻止α2M受体表达增加，但在8小时时无作用。在37℃，与未处理的细胞相比，用1000单位/ml CSF-1处理18小时的BMMs对125I-α2M-CH3NH2的摄取和消化增加。这些研究表明，CSF-1通过一条需要新的RNA和蛋白质合成的途径增加BMMs中α2M受体的表达。我们假设α2M受体表达增加可能在细胞生长和分化中起重要作用。
