一种血清诱导巨噬细胞转化为泡沫细胞新方法的建立
Establishment of a Novel Method to Induce Formation of the Foam Cells from Macrophages with Fetal Bovine Serum

作者: 赵泓翔 , 缪红 , 商亚珍 :河北省中药研究与开发重点实验室/承德医学院中药研究所,承德;

关键词: 巨噬细胞泡沫细胞血清氧化型低密度脂蛋白体外培养Macrophages Foam Cells Serum Oxidized Low Density Lipoprotein In Vitro Culture

摘要:

目的:利用血清替代氧化型低密度脂蛋白(oxidized low density lipoprotein, ox-LDL)诱导巨噬细胞,建立一种新的诱导泡沫细胞体外培养方法。方法:小鼠腹腔注射RPMI-1640培养液提取巨噬细胞,RPMI-1640无血清培养4 h贴壁,分别加含不同浓度胎牛血清(体积分数5%、10%和20%)的RPMI-1640培养液、无血清氧化型低密度脂蛋白(2 × 10−2 g/L)RPMI-1640培养液培养4 h、8 h和12 h,采用油红O染色,观察巨噬细胞吞噬脂滴转化为泡沫细胞情况。采用酶化学法检测细胞总胆固醇酯和游离胆固醇含量。结果:不同浓度胎牛血清(体积分数5%,10%和20%)作用不同时间(4 h,8 h和12 h)均对巨噬细胞有诱导作用,且随着浓度的增加、时间的延长,巨噬细胞内脂滴增多变大,部分融合成大脂滴。但当血清浓度增加到20%时,作用时间多于8 h,巨噬细胞有部分破裂、脂滴丢失、细胞减少的现象。在浓度为10%的血清作用7 h诱导的泡沫细胞,细胞内的胆固醇酯含量明显高于空白对照的。结论:浓度为10%的血清作用7 h可诱导巨噬细胞转化为泡沫细胞。

Abstract: Objective: To establish a method for forming foam cells from macrophages by serum instead of oxidized low density lipoprotein (ox-LDL) in vitro. Methods: The macrophages were isolated from the mice by intraperitoneal injection RPMI-1640 culture medium. The macrophages were cultured 4 h in serum-free RPMI-1640 medium for adherence. Then, the adherent macrophages were added RPMI-1640 culture medium with different concentrations of fetal bovine serum (volume fraction of 5%, 10% and 20%) or oxidized low density lipoprotein (2 × 10−2 g/L). The cells were stained with oil red O following the above conducts for observing the intracellular lipid droplets. The contents of intracellular total cholesterol and cholesteryl esters were measured by enzymology. Results: Both different concentrations of fetal bovine serum (volume fraction of 5%, 10% and 20%) and different action time (4 h, 8 h and 12 h) can induce the macrophages into foam cells, and the intracellular lipid droplets were bigger and even the cells merged together a bigger lipid droplets. The macrophages membrane was broken and the lipid droplet and cells were lost when the serum was increased to 20% and the action time was more than 8 h. Macrophages cultured by 10% serum and acted 7 h, cellular contents of cholesteroy and cholesteryl ester increased markedly compared with the normal macrophages. Conclusion: A foam cell model has been established by incubating macrophages with 10% fetal-bovine serum amount of RPMI-1640 medium for 7 hours in vitro culture.

文章引用: 赵泓翔 , 缪红 , 商亚珍 (2014) 一种血清诱导巨噬细胞转化为泡沫细胞新方法的建立 。 生物过程, 4, 11-18. doi: 10.12677/BP.2014.42002

参考文献

[1] Badimon, L., Storev, R.F. and Vilahur, G. (2011) Update on lipid, inflammation and atherothrombosis. Thrombosis and Haemostasis, 105, S34-S42.

[2] Saito, A., Fujimura, M., Inoue, T., et al. (2010) Relationship between lectin-like oxidized low-density lipoprotein receptor 1 expression and preoperative echogenic findings of vulnerable carotid plaque. Acta Neurochirurgica, 152, 589- 595.

[3] Li, D.Y. and Mehta, J.L. (2000) Upregulation of endothelial receptor for oxidized LDL (LOX-1) by oxidized LDL and implications in apopotosis of human coronary artery endothelial cells evidence from use of antisense LOX-1 mRNA and chemical inhibitors. Arterosclerosis, Thrombosis, and Vascular Biology, 20, 1116-1122.

[4] Licadtro, F., Candore, G., Lio, D., et al. (2005) Innate immunity and inflammation in ageing: A key for understanding age-related diseases. Immunity & Ageing, 2, 8.

[5] Briley-Saebo, K.C., Shaw, P.X., Mulder, W.J., et al. (2008) Targeted molecular probes for imaging atherosclerotic lesions with magnetic resonance using antibodies that recognize oxidation-specific epitopes. Circulation, 117, 3206- 3215.

[6] 徐佳, 蔡辉 (2011) 氧化低密度脂蛋白与动脉粥样硬化的研究. 安徽医药, 2, 228-230.

[7] Steinberg, D. and Witztum, J.L. (2002) Is the oxidative modification hypothesis relevant to hum an atherosclerosis? Do the antioxidant trials conducted to date refute the hypothesis? Circulation, 105, 2107-2111.

[8] 高琳琳, 李福荣, 姚树桐, 等 (2012) 蚤休薯蓣皂苷对内皮细胞的保护作用涉及TLR4/NF-κB途径. 中国药理学通报, 11, 1598-1602.

[9] Lo, Y.H., Pan, M.H., Li, S., et al. (2010) Nobiletin metabolite, 3’, 4’-dihydroxy-5, 6, 7, 8-tetramethoxyflavone, inhibits LDL oxidation and down-regulates scavenger receptor expression and activity in THP-1 cells. Biochimica et Biophysica Acta, 1801, 114.

[10] 尹美珍, 李世普, 袁琳, 等 (2006) 小鼠腹腔巨噬细胞的分离培养与鉴定. 武汉大学学报, 2, 203-205.

[11] Mander, E. (1994) Apolipoprotein B of oxidized LDL accumulates in the lysosomes of macrophages. Biochemica et Biophysica Acta, 1212, 80-92.

[12] Steinbrecher, U.P. and Lougheed, M. (1992) Scavenger receptor-independent stimulation of cholesterol esterification in macrophages by low density lipoprotein extracted from human aortic intima. Arteriosclerosis and Thrombosis, 12, 608-625.

[13] Asmis, R., Begley, J.G., Jelk, J., et al. (2005) Lipoprotein aggregation protects human monocyte-derived macrophages from Ox-LDL-induced cytotoxicity. Journal of Lipid Research, 46, 1124-1132.

[14] 袁中华, 贾薇, 黄谙非, 等 (2008) 脂肪分化相关蛋白通过酰基辅酶A: 胆固醇酰基转移酶1促进巨噬细胞脂质蓄积. 中国动脉硬化杂志, 2, 101-106.

[15] 王术玲, 潘华新, 王培训 (2009) THP-1巨噬细胞源性泡沫细胞模型的建立及鉴定. 中药新药与临床药理, 3, 281- 284.

[16] 贺全国, 蒋佩, 严鹏科, 等 (2007) 胆固醇流出对氧化性低密度脂蛋白诱导的巨噬细胞源性泡沫细胞凋亡的影响. 国现代医学杂志, 22, 2724-2727.

[17] 何雪峰, 李晓辉, 李淑慧, 等 (2007) 三七总皂苷对小鼠巨噬细胞源性泡沫细胞形成的影响. 中国药房, 30, 2323-2325.

[18] 周云, 沃兴德, 卢德赵 (2010) RAW264.7巨噬细胞源性泡沫细胞模型的建立及鉴定. 中国动脉硬化杂志, 9, 687- 689.

[19] 董关木 (2007) 细胞培养用牛血清现状与进展. 中国生物制品学杂志, 9, 697-700.

分享
Top