Long E, Huynh H T, Zhao X
Department of Animal Science, McGill University, Ste-Anne-de-Bellevue, Quebec, Canada.
Endocrine. 1998 Oct;9(2):185-92. doi: 10.1385/ENDO:9:2:185.
Insulin-like growth factor 1 (IGF-1) and its binding proteins (IGFBPs) are involved in proliferation and differentiation of many cell types. In the present study, the involvement of IGF-1 and IGFBPs in proliferation and differentiation of murine bone marrow-derived macrophages (BMDM) was investigated. L929-conditioned media (LCM) containing abundant macrophage colony-stimulating factor CSF-1 were used to stimulate BMDM development from their bone marrow precursors. The alteration of IGF-1 and IGFBPs during LCM-induced BMDM proliferation and differentiation was first studied. The cells were cultured in RPMI complete media containing 20% LCM for different time periods and then incubated in serum-free media for 24 h. The supernatants were collected for Western ligand blotting and immunoblotting analyses, and the cell pellets for Northern blotting analyses. The mRNA level of IGF-1 increased in a time-dependent manner. An increase of IGFBP-4 accumulation in the conditioned media was also observed during this process. However the mRNA expression of IGFBP-4 remained constant, indicating a posttranscriptional regulation of IGFBP-4 secretion and/or stability. The effects of exogenous recombinant human IGF-1 (rhIGF-1) on BMDM proliferation and differentiation were further studied. Two IGF-1 analogs (long R3 IGF-1 and des [1-3] IGF-1) were also used in parallel with regular IGF-1 to indicate the involvement of IGFBPs in BMDM development. Cells were cultured in complete media containing 20% LCM for different time periods, and then incubated in serum-free media in the presence of rhIGF-1 or its analogs for 24 h. These three forms of IGF-1 all potentiated the proliferation of freshly isolated BMDM precursors (d 0). rhIGF-1 and long R3 IGF-1, but not des (1-3) IGF-1, continued to stimulate the cell proliferation on d 1. The effects of these three forms of IGF-1 on BMDM differentiation were investigated using mannose receptor expression as a marker. Long R3 IGF-1 and des (1-3) IGF-1, but not rhIGF-1, enhanced BMDM differentiation on d 4. The different effects of rhIGF-1 and its analogs on BMDM differentiation suggest that the accumulation of IGFBP-4 in BMDM development might have an inhibitory effect on IGF-1 actions by sequestering free IGF-1.
胰岛素样生长因子1（IGF-1）及其结合蛋白（IGFBPs）参与多种细胞类型的增殖和分化。在本研究中，研究了IGF-1和IGFBPs在小鼠骨髓来源巨噬细胞（BMDM）增殖和分化中的作用。使用含有丰富巨噬细胞集落刺激因子CSF-1的L929条件培养基（LCM）刺激BMDM从其骨髓前体发育。首先研究了LCM诱导BMDM增殖和分化过程中IGF-1和IGFBPs的变化。将细胞在含有20%LCM的RPMI完全培养基中培养不同时间段，然后在无血清培养基中孵育24小时。收集上清液进行Western配体印迹和免疫印迹分析，收集细胞沉淀进行Northern印迹分析。IGF-1的mRNA水平呈时间依赖性增加。在此过程中还观察到条件培养基中IGFBP-4积累增加。然而，IGFBP-4的mRNA表达保持恒定，表明IGFBP-4分泌和/或稳定性存在转录后调控。进一步研究了外源性重组人IGF-1（rhIGF-1）对BMDM增殖和分化的影响。还将两种IGF-1类似物（长R3 IGF-1和des [1-3] IGF-1）与常规IGF-1并行使用，以表明IGFBPs参与BMDM发育。将细胞在含有20%LCM的完全培养基中培养不同时间段，然后在rhIGF-1或其类似物存在下于无血清培养基中孵育24小时。这三种形式的IGF-1均增强了新鲜分离的BMDM前体（第0天）的增殖。rhIGF-1和长R3 IGF-1，但不是des（1-3）IGF-1，在第1天继续刺激细胞增殖。使用甘露糖受体表达作为标志物研究了这三种形式的IGF-1对BMDM分化的影响。长R3 IGF-1和des（1-3）IGF-1，但不是rhIGF-1，在第4天增强了BMDM分化。rhIGF-1及其类似物对BMDM分化的不同影响表明，BMDM发育过程中IGFBP-4的积累可能通过螯合游离IGF-1对IGF-1的作用产生抑制作用。
