糖浓度变化对牛肺泡巨噬细胞促炎细胞因子释放的影响

doi:10.3969/j.issn.1004-1524.2022.03.06

浙江农业学报 ›› 2022, Vol. 34 ›› Issue (3): 464-470.DOI: 10.3969/j.issn.1004-1524.2022.03.06

糖浓度变化对牛肺泡巨噬细胞促炎细胞因子释放的影响

谭天宇¹(), 才冬杰¹, 王之盛², 左之才¹^,^*()

1.四川农业大学动物医学院,动物疫病与人类健康四川省重点实验室,四川成都 611130
2.四川农业大学动物营养研究所,四川成都 611130

收稿日期:2020-11-06 出版日期:2022-03-25 发布日期:2022-03-30
通讯作者: 左之才
作者简介:*左之才,E-mail: zzcjl@126.com
谭天宇(1996—),男,四川广安人,硕士研究生,研究方向为中西兽医与临床。E-mail: 13350907018@163.com
基金资助:
国家现代农业产业技术体系(肉牛牦牛,CARS-37(肉牛牦牛);国家现代农业产业技术体系(肉牛牦牛,CARS-37(CARS-37);国家现代农业产业技术体系四川肉牛创新团队(SCCXTD-2020-13)

Effect of glucose concentrations on proinflammatory cytokine released from bovine alveolar macrophages

TAN Tianyu¹(), CAI Dongjie¹, WANG Zhisheng², ZUO Zhicai¹^,^*()

1. Sichuan Key Laboratory of Animal Diseases and Human Health,College of Veterinary Medicine, Sichuan Agricultural University, Chengdu 611130, China
2. Animal Nutrition Institute,Sichuan Agricultural University,Chengdu 611130,China

Received:2020-11-06 Online:2022-03-25 Published:2022-03-30
Contact: ZUO Zhicai

摘要/Abstract

摘要：

探讨葡萄糖浓度变化能否上调牛肺泡巨噬细胞(BAMs)RAGE-TLR4-MyD88-NF-κB信号通路,引起BAMs释放促炎细胞因子IL-1β、IL-6及TNF-α。Ⅰ、Ⅱ、Ⅲ、Ⅳ组分别用5.5、15.5、25.5 mmol·L^-1葡萄糖以及葡萄糖(5.5 mmol·L^-1)+甘露醇(20 mmol·L^-1)作用BAMs 12 h,收集培养细胞和上清液;另采用25.5 mmol·L^-1葡萄糖与BAMs分别作用 0、3、6、12、24 h,收集各时间点培养细胞和上清液。检测RAGE、TLR4、MyD88及NF-κB p65 mRNA水平以及上清液中IL-1β、IL-6及TNF-α浓度。结果显示,Ⅲ组RAGE、TLR4、MyD88和NF-κB p65 mRNA水平以及上清液中IL-1β、IL-6、TNF-α浓度极显著(P<0.01)高于Ⅰ组,Ⅱ组IL-1β及TNF-α浓度极显著(P<0.01)高于Ⅰ组;MyD88、TLR4、NF-κB p65 mRNA水平在3 h时显著(P<0.05)高于0 h,RAGE在6 h时显著(P<0.05)高于0 h。4个基因指标均于12 h达到峰值,极显著(P<0.01)高于其他时间点。只有RAGE、NF-κB p65在24 h仍极显著(P<0.01)高于0 h;IL-1β、IL-6及TNF-α浓度均在3 h后相比0 h显著(P<0.05)升高,并呈现时间依赖性,于24 h达到峰值。综上,25.5 mmol·L^-1葡萄糖能上调RAGE-TLR4-MyD88-NF-κB信号通路的基因表达,促进促炎细胞因子IL-1β、IL-6及TNF-α的释放。

关键词: 高糖, 牛肺泡巨噬细胞, 炎症信号通路

Abstract:

In order to investigate whether changes in glucose concentration can up-regulate the RAGE-TLR4-MyD88-NF-κB signaling pathway of bovine alveolar macrophages (BAMs) and cause BAMs to release proinflammatory cytokines IL-1β, IL-6 and TNF-α.Groups Ⅰ, Ⅱ, Ⅲ and Ⅳ were treated with 5.5, 15.5, 25.5 mmol·L^-1 glucose and glucose (5.5 mmol·L^-1)+mannitol (20 mmol·L^-1) for 12 h to collect the cultured cells and supernatants; and 25.5 mmol·L^-1 glucose and BAMs were used for 0, 3, 6, 12, 24 h to collect the cultured cells and supernatants at each time point.RAGE, TLR4, MyD88 and NF-κB p65 mRNA levels and IL-1β, IL-6 and TNF-α concentrations in the supernatant were detected.The results showed that the levels of RAGE, TLR4, MyD88 and NF-κB p65 mRNA and the concentrations of IL-1β, IL-6 and TNF-α in group Ⅲ were significantly (P<0.01)higher than those in group Ⅰ, the concentrations of IL-1β and TNF-α in group Ⅱ were significantly(P<0.01) higher than those in group Ⅰ, the levels of MyD88, TLR4 and NF-κB p65 mRNA were significantly (P<0.05)higher than those in group Ⅰ at 3 h, and RAGE was significantly (P<0.05)higher than that at 6 h.The four gene indices all reached the peak at 12 h, which was significantly (P<0.01)higher than other time points.Only RAGE and NF-κB p65 at 24 h were significantly(P<0.01) higher than 0 h. The concentrations of IL-1β, IL-6 and TNF-α were significantly (P<0.05)higher after 3 h than 0 h, and showed a time-dependent peak at 24 h. In conclusion, 25.5 mmol·L^-1 glucose can up-regulate the gene expression in the BAMs RAGE-TLR4-MyD88-NF-κB signaling pathway and promote the release of proinflammatory cytokines IL-1β, IL-6 and TNF-α.

Key words: high glucose, bovine alveolar macrophages, inflammatory signaling pathways

中图分类号:

S852

谭天宇, 才冬杰, 王之盛, 左之才. 糖浓度变化对牛肺泡巨噬细胞促炎细胞因子释放的影响[J]. 浙江农业学报, 2022, 34(3): 464-470.

TAN Tianyu, CAI Dongjie, WANG Zhisheng, ZUO Zhicai. Effect of glucose concentrations on proinflammatory cytokine released from bovine alveolar macrophages[J]. Acta Agriculturae Zhejiangensis, 2022, 34(3): 464-470.

图/表 5

表1 各基因实时荧光定量引物序列

Table 1 Primer sequence of genes for real-time PCR

基因 Gene	引物序列 Primer sequence(5'-3')	登录号 GenBank ID
β-actin	F:GCCCATCTATGAGGGGTACG	AF-191490.1
	R:TCACGGACGATTTCCGCT
RAGE	F:GACAGTCGCCCTGCTCATT	XM-010818313.1
	R:CCTCTGGCTGGTTCAGTTCC
TLR4	F:TGCCTTCACTACAGGGACTTT	NM-174198.6
	R:TGGGACACCACGACAATAAC
NF-κB p65	F:GAGATCATCGAGCAGCCCAA	XM-002695167.5
	R:ATAGTGGGGTGGGTCTTGGT
MyD88	F:AGAAGAGGTGCCGTCGGATGG	XM-005222379.3
	R:TTGGTGTAGTCACAGACAGTGATGAAG

图1 不同浓度葡萄糖处理BAMs 12 h对RAGE、TLR4、MyD88、NF-κBp65基因表达的影响 *表示与对照组差异显著(P<0.05),**表示与对照组差异极显著(P<0.01)。下同。

Fig.1 Effects of different concentrations of glucose on gene expression of RAGE, TLR4, MyD88 and NF-κB p65 in bovine alveolar macrophages induced for 12 hours * meaned there was a significant difference from the control group (P<0.05), ** meaned there was a very significant difference from the control group (P<0.01). The same as below.

图2 不同浓度葡萄糖诱导BAMs 12 h对IL-1β、IL-6及TNF-α释放的影响

Fig.2 Effects of different concentrations of glucose on releasing of IL-1β, IL-6, TNF-α in bovine alveolar macrophages induced for 12 hours

图3 高糖诱导时间对牛肺泡巨噬细胞RAGE、TLR4、MyD88、NF-κB p65基因表达的影响

Fig.3 Effects of different high glucose induction time on gene expression of RAGE, TLR4, MyD88 and NF-κB p65 in bovine alveolar macrophages

图4 高糖诱导时间对BAMs IL-1β、IL-6及TNF-α释放的影响

Fig.4 Effects of different high glucose induction time on releasing of IL-1β, IL-6, TNF-α in bovine alveolar macrophages

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糖浓度变化对牛肺泡巨噬细胞促炎细胞因子释放的影响

Effect of glucose concentrations on proinflammatory cytokine released from bovine alveolar macrophages

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