黑木耳凝集素对RAW264.7细胞免疫调节作用的研究

doi:10.11924/j.issn.1000-6850.casb2020-0228

中国农学通报 ›› 2021, Vol. 37 ›› Issue (9): 125-130.doi: 10.11924/j.issn.1000-6850.casb2020-0228

所属专题：生物技术；食用菌；食用菌；园艺

• 食品·营养·检测·安全 • 上一篇下一篇

黑木耳凝集素对RAW264.7细胞免疫调节作用的研究

房佳敏¹(), 马成瑶², 张彦龙¹()

¹黑龙江大学生命科学学院/黑龙江省普通高校微生物重点实验室,哈尔滨 150080
²黑龙江大学/农业微生物技术教育部工程研究中心,哈尔滨 150500

收稿日期:2020-07-02 修回日期:2020-08-18 出版日期:2021-03-25 发布日期:2021-04-09
通讯作者: 张彦龙
作者简介:房佳敏,女,1996年出生,黑龙江虎林人,在读硕士生,研究方向：天然药物化学。通信地址：150080 黑龙江省哈尔滨市南岗区哈西街道黑龙江大学生命科学学院,E-mail： a369704253@qq.com。
基金资助:
国家星火计划项目(2010GA670002)

The Immunomodulatory Effect of Auricularia curicula Lectin on RAW264.7 Cells

Fang Jiamin¹(), Ma Chengyao², Zhang Yanlong¹()

¹School of Life Sciences, Heilongjiang University/Heilongjiang Provincial Key Laboratory of Microbiology, Harbin 150080
²Engineering Research Center of Agricultural Microbiology Technology, Ministry of Education/ Heilongjiang University, Harbin 150500

Received:2020-07-02 Revised:2020-08-18 Online:2021-03-25 Published:2021-04-09
Contact: Zhang Yanlong

摘要/Abstract

摘要：

为解决黑木耳凝集素对RAW246.7细胞分泌IL-6、IL-1β、TNF-α、NO释放量与免疫调节作用的问题,通过40%(NH₄)₂SO₄沉淀法对黑木耳凝集素进行分离提纯,并伴随凝血活性检测、氨基酸组分检测,得到黑木耳凝集素粗品。经过AKTA蛋白纯化系统得到黑木耳凝集素纯品。使用SDS-PAGE电泳对凝集素分子量进行分析。格里斯法检测巨噬细胞RAW264.7分泌NO的情况。ELISA检测IL-6、IL-1β、TNF-α的释放量。结果表明,黑木耳凝集素分子量约为20 kDa。通过检测发现,黑木耳凝集素对巨噬细胞RAW264.7分泌细胞因子IL-6、IL-1β、TNF-α有促进作用,且呈现剂量依赖,具有免疫调节的作用。

关键词: 黑木耳, 凝集素, 分离纯化, 免疫调节, 细胞因子

Abstract:

The paper aims to solve the problem of the release of IL-6, IL-1β, TNF-α, NO and the immune regulation effect of Auricularia auricula lectin on RAW246.7 cells. The A. auricula lectin was separated and purified by 40% (NH₄)₂SO₄precipitation method, and accompanied by the detection of coagulation activity and amino acid composition, the crude product of A. auricula lectin was obtained. Through the AKTA protein purification system, pure A. auricula lectin was obtained. The molecular weight of the lectin was analyzed by using SDS-PAGE electrophoresis. The Griess method was used to detect the secretion of NO by macrophage RAW264.7. ELISA was used to detect the release of IL-6, IL-1β and TNF-α. The results showed that the molecular weight of the A. auricula lectin was about 20 kDa. Through detection, it was found that A. auricula lectin could promote the secretion of cytokines IL-6, IL-1β and TNF-α by macrophage RAW264.7, and it showed a dose dependent and immunomodulatory effect.

Key words: Auricularia auricula, lectin, extraction and purification, immune regulation, cytokines

中图分类号:

S646.6

房佳敏, 马成瑶, 张彦龙. 黑木耳凝集素对RAW264.7细胞免疫调节作用的研究[J]. 中国农学通报, 2021, 37(9): 125-130.

Fang Jiamin, Ma Chengyao, Zhang Yanlong. The Immunomodulatory Effect of Auricularia curicula Lectin on RAW264.7 Cells[J]. Chinese Agricultural Science Bulletin, 2021, 37(9): 125-130.

图/表 7

图1. SDS-PAGE电泳纯度鉴定结果

图2. 黑木耳凝集素凝血活性测定

氨基酸种类	氨基酸含量/mg	氨基酸种类	氨基酸含量/mg
天冬氨酸	11.83	异亮氨酸	10.70
苏氨酸	5.67	亮氨酸	10.63
丝氨酸	4.79	酪氨酸	7.17
谷氨酸	13.46	苯丙氨酸	6.67
半胱氨酸	10.31	赖氨酸	2.03
丙氨酸	7.45	组氨酸	3.14
蛋氨酸	3.73	精氨酸	2.42

表1. 黑木耳凝集素LAA的氨基酸组成

图3. 黑木耳凝集素LAA对RAW264.7细胞产生IL-6的影响

图4. 黑木耳凝集素LAA对RAW264.7细胞产生IL-1β的影响

图5. LAA对RAW264.7细胞产生TNF-α的影响

图6. 黑木耳凝集素LAA对RAW264.7细胞产生NO的影响

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黑木耳凝集素对RAW264.7细胞免疫调节作用的研究

The Immunomodulatory Effect of Auricularia curicula Lectin on RAW264.7 Cells

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