Rios Francisco J O, Gidlund Magnus, Jancar Sonia
Department of Immunology, Institute of Biomedical Sciences, University of São Paulo, São Paulo, Brazil.
Cell Physiol Biochem. 2011;27(3-4):363-72. doi: 10.1159/000327962. Epub 2011 Apr 1.
The uptake of oxLDL by CD36 is not regulated by intracellular levels of cholesterol, leading to macrophage differentiation into foam cells which play a major role in atherosclerosis. Furthermore, oxLDL competes with PAF in macrophages for binding to PAF receptors (PAFR). Here we investigated the involvement of PAFR in CD36 expression and uptake of oxLDL by human monocytes/macrophages. Adherent peripheral blood mononuclear cells were treated with PAFR-antagonists (WEB2170, CV3988); inhibitors of ERK1/2 (PD98059), p38 (SB203580), JNK (SP600125) or diluents, before stimulation with oxLDL or PAF. After 24 h, uptake of FITC-oxLDL and expression of CD36 was determined by flow cytometry and phosphorylation of MAP-kinases by Western blot. It was shown that the uptake of oxLDL was reduced by PAFR antagonists. CD36 expression was up-regulated by oxLDL, an effect reversed by PAFR antagonists. The up-regulation of CD36 and oxLDL uptake both required MAP-kinases activation. The oxLDL-induced ERK1/2 and JNK but not p38 phosphorylation was reversed by PAFR-antagonists suggesting that oxLDL signalling involves PAFR dependent and independent pathways. In macrophages from PAFR(-/-) mice, oxLDL was unable to up-regulate CD36 expression and the oxLDL uptake was reduced compared to wild type. These results suggest that oxLDL interacts with PAFR in macrophages to increase CD36 expression and oxLDL uptake. Whereas pharmacological intervention at the level of PAFR would be beneficial in atherosclerosis remains to be determined.
CD36对氧化型低密度脂蛋白（oxLDL）的摄取不受细胞内胆固醇水平的调节，这会导致巨噬细胞分化为泡沫细胞，而泡沫细胞在动脉粥样硬化中起主要作用。此外，oxLDL在巨噬细胞中与血小板活化因子（PAF）竞争结合血小板活化因子受体（PAFR）。在此，我们研究了PAFR在人单核细胞/巨噬细胞中CD36表达及oxLDL摄取过程中的作用。在用oxLDL或PAF刺激之前，将贴壁的外周血单核细胞用PAFR拮抗剂（WEB2170、CV3988）、细胞外信号调节激酶1/2（ERK1/2）抑制剂（PD98059）、p38抑制剂（SB203580）、c-Jun氨基末端激酶（JNK）抑制剂（SP600125）或稀释剂进行处理。24小时后，通过流式细胞术测定异硫氰酸荧光素标记的oxLDL（FITC-oxLDL）的摄取及CD36的表达，并通过蛋白质印迹法检测丝裂原活化蛋白激酶（MAP激酶）的磷酸化情况。结果显示，PAFR拮抗剂可降低oxLDL的摄取。oxLDL可上调CD36的表达，而PAFR拮抗剂可逆转这一效应。CD36的上调及oxLDL的摄取均需要MAP激酶的激活。PAFR拮抗剂可逆转oxLDL诱导的ERK1/2和JNK磷酸化，但不能逆转p38的磷酸化，这表明oxLDL信号传导涉及PAFR依赖性和非依赖性途径。在来自PAFR基因敲除（PAFR(-/-)）小鼠的巨噬细胞中，oxLDL无法上调CD36的表达，且与野生型相比，oxLDL的摄取减少。这些结果表明，oxLDL在巨噬细胞中与PAFR相互作用，以增加CD36的表达及oxLDL的摄取。而在PAFR水平进行药物干预对动脉粥样硬化是否有益仍有待确定。
