EGF和FGF-2对猪皮下脂肪神经嵴干细胞增殖和分化的影响

doi:10.16431/j.cnki.1671-7236.2022.12.035

摘要/Abstract

摘要： 【目的】探究表皮生长因子（epidermal growth factor，EGF）和成纤维细胞生长因子2（fibroblast growth factor 2，FGF-2）对猪皮下脂肪神经嵴干细胞（neural crest stem cells，NCSCs）增殖及分化的影响，以优化猪皮下脂肪神经嵴干细胞的培养条件。【方法】通过体外分离培养原代猪皮下脂肪NCSCs，免疫荧光染色鉴定NCSCs标志物p75 NTR，并用不同浓度的EGF和FGF-2（0和0、10和10、10和20、20和10、20和20、30和30 ng/mL）作用于传代猪皮下脂肪NCSCs，用CCK-8试剂盒测定细胞增殖率，确定细胞生长的最适EGF和FGF-2浓度，将试验分为空白组和最适浓度组，测定两组细胞的生长曲线，成脂化诱导后油红O染色，对比两组细胞的脂滴生成量。【结果】免疫荧光染色结果显示，原代猪皮下脂肪NCSCs经p75 NTR鉴定呈阳性。CCK-8细胞增殖试验结果显示，EGF和FGF-2的浓度均为20 ng/mL时对传代猪皮下脂肪NCSCs的促增殖作用最佳。生长曲线显示，两组细胞均在第5~9天处于对数生长期，第10~15天细胞增殖减缓，逐渐到达停滞期。油红O染色结果显示，最适浓度组胞质内的脂滴生成量远多于对照组。【结论】在培养液中添加20 ng/mL EGF和20 ng/mL FGF-2对猪皮下脂肪NCSCs的增殖和分化有促进作用。

关键词: 猪皮下脂肪; 表皮生长因子(EGF); 成纤维细胞生长因子2(FGF-2); 神经嵴干细胞(NCSCs)

Abstract: 【Objective】 The aim of this study was to investigate the effects of epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF-2) on proliferation and differentiation of porcine subcutaneous adipose neural crest stem cells (NCSCs), so as to optimize the culture conditions of porcine subcutaneous adipose NCSCs.【Method】 The primary porcine subcutaneous adipose NCSCs were isolated and cultured in vitro, and the NCSCs marker p75 NTR was identified by immunofluorescence staining.The porcine subcutaneous adipose NCSCs were treated with different concentrations of EGF and FGF-2 (0 and 0, 10 and 10, 10 and 20, 20 and 10, 20 and 20, 30 and 30 ng/mL).The cell proliferation rate was tested by CCK-8 kit to determine the optimal concentration of EGF and FGF-2 for cell growth.The experiment was divided into control and optimal concentration groups.The cell growth curve of two groups was drawn, and the lipid droplet production of cells in two groups was compared by oil red O staining after the adipogenic induction.【Result】 The results of immunofluorescence staining showed that the primary porcine subcutaneous adipose NCSCs were positive by p75 NTR.The results of CCK-8 cell proliferation showed that EGF and FGF-2 had the best proliferative effect on porcine subcutaneous adipose NCSCs when the concentration of EGF and FGF-2 was 20 ng/mL, respectively.The results of growth curve showed that the cells in two groups were in logarithmic growth phase from 5th to 9th days, and the cell proliferation slowed down from 10th to 15th days, and gradually reached stagnation stage.The results of oil red O staining showed that the amount of lipid droplets in the cytoplasm of the optimal concentration group was much more than that of control group.【Conclusion】 Adding 20 ng/mL EGF and 20 ng/mL FGF-2 in culture medium could promote the proliferation and differentiation of porcine subcutaneous adipose NCSCs.

Key words: porcine subcutaneous adipose; epidermal growth factor (EGF); fibroblast growth factor 2 (FGF-2); neural crest stem cells (NCSCs)

中图分类号:

董博文, 崔小珍, 白瑞, 郑明学, 张黎, 吕晓玲, 任玉红. EGF和FGF-2对猪皮下脂肪神经嵴干细胞增殖和分化的影响[J]. 中国畜牧兽医, 2022, 49(12): 4878-4886.

DONG Bowen, CUI Xiaozhen, BAI Rui, ZHENG Mingxue, ZHANG Li, LYU Xiaoling, REN Yuhong. Effects of EGF and FGF-2 on Proliferation and Differentiation of Porcine Subcutaneous Adipose Neural Crest Stem Cells[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(12): 4878-4886.

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