高硼暴露通过诱导细胞焦亡加剧鸡肝细胞损伤

doi:10.11843/j.issn.0366-6964.2024.10.044

摘要/Abstract

摘要：

旨在探索硼暴露诱导鸡肝细胞损伤的作用机制。试验选取30只AA肉鸡随机分为对照组(control group, CG)、低硼组(low boron group, LBG)和高硼组(high boron group, HBG)，在基础日粮中分别添加不同剂量硼进行饲喂：基础日粮(硼0 mg·kg^-1), 低硼日粮(硼120 mg·kg^-1)、高硼日粮(硼240 mg·kg^-1)，处理7及14 d后采集肝组织。使用ICP-MS测定肝组织中的硼含量，全自动血生化分析仪检测ASL及ALT水平，HE染色观察肝组织病理变化，透射电镜观察肝细胞超微结构变化，免疫组化和免疫荧光检测肝细胞焦亡相关蛋白NLRP3及IL-1β定位表达情况，RT-qPCR和Western blot检测NLRP3、Casapse-1、IL-1β、IL-18和GSDMD的mRNA和蛋白表达水平。结果显示，蓄积于肝组织中的硼含量随着硼暴露时间和剂量的增加而增加。此外，14 d-低硼组(14 d-LBG)和高硼组(14 d-HBG)的肝组织结构明显破坏，并且细胞焦亡水平随硼剂量的增加而加剧；与同时间的CG相比，14 d-LBG中NLRP3、Caspase-1、IL-18的mRNA表达水平显著上升(P < 0.05)，14 d-HBG的NLRP3、Caspase-1、IL-1β、IL-18及GSDMD的mRNA和蛋白表达水平显著上升(P < 0.05)。高硼暴露可通过诱导细胞焦亡进而造成鸡肝细胞损伤。

关键词: 硼, 细胞焦亡, 鸡, 肝细胞

Abstract:

We aim to explore the mechanism of boron-induced hepatocytes damage in broilers. Thirty AA broiler chickens were randomly divided into control group (CG), low boron group (LBG) and high boron group (HBG). They were fed with different additional doses of boron in the basal diet: the basal diet (boron 0 mg·kg^-1), low boron diet (boron 120 mg·kg^-1) and high boron diet (boron 240 mg·kg^-1), respectively. Liver tissues were collected 7 and 14 days after treatment. The content of boron in liver tissue was determined by ICP-MS, the ASL and ALT levels were tested by automatic blood biochemistry analyzer, the pathological changes in liver tissue were observed by H & E staining, ultrastructural changes in liver tissue were observed by transmission electron microscopy, and the localized expressions of NLRP3 and IL-1β in liver tissue were detected by immunohistochemistry and immunofluorescence. The mRNA and protein expression levels of NLRP3, Casapse-1, IL-1β, IL-18, and GSDMD were detected by RT-qPCR and Western blot. The results showed that the amount of boron stored in the liver tissue as along dietary boron levels increased, with the duration and dosage of exposure. In addition, the structural integrity of the liver tissue of 14-day-low boron group (14 d-LBG) and 14-day-high boron group (14 d-HBG) was significantly compromised, with an increase in pyroptosis levels with an increase in boron dosage. Compared with CG at the same time, a significant upregulation of mRNA expression levels of NLRP3, Caspase-1 and IL-18 was observed in the 14 d-LBG(P < 0.05). Furthermore, a significant upregulation of both mRNA and protein expression levels of NLRP3, Caspase-1, IL-1β, IL-18 and GSDMD was observed in the 14 d-HBG (P < 0.05). High levels of boron exposure could induce hepatocytes damage in broilers through the pyroptosis pathway.

Key words: boron, pyroptosis, broiler, hepatocyte

中图分类号:

S856.9

何婷, 刘思颖, 全锦雯, 苏伟谦, 黄江利, 李玙萌, 刘忠华, 余文兰. 高硼暴露通过诱导细胞焦亡加剧鸡肝细胞损伤[J]. 畜牧兽医学报, 2024, 55(10): 4747-4759.

Ting HE, Siying LIU, Jinwen QUAN, Weiqian SU, Jiangli HUANG, Yumeng LI, Zhonghua LIU, Wenlan YU. High Levels of Boron Exposure Aggravated Hepatocytes Damage in Broilers via Pyroptosis[J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(10): 4747-4759.

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日粮组成Diet composition		营养水平Nutrients levels
配料Ingredients	含量Content	指标Indices	水平Levels
玉米Conrn	54.82	干物质Dry matter	88.60
豆粕Soybean	37.77	粗蛋白Crude protein	23.00
豆油Soybean oil	2.50	代谢能/(MJ·kg^-1)	12.70
石粉Limestone	1.01	钙Calcium	0.95
磷酸氢钙Dicalcium phosphate	1.75	有效磷Avail-P	0.46
食盐Salt	0.40
添加剂^* Additive^*	0.50
DL-蛋氨酸DL-Methionine	0.25
玉米淀粉+铜Conrn starch+Cu	1.00

基因 Genes	引物序列(5′→3′) Primer sequence	产物长度/bp Product length	序列号 Sequence number
NLRP3	AGCTACCACACATCTAGGAT	207	XM_046918112.1
	GGTGTCCAAATCCTCAATCT
Caspase-1	TTCCTTCAACACCATCTACG	209	XM_040687588.2
	GGTGAGCTTCTCTGGTTTTA
IL-1β	CAGCCTCAGCGAAGAGACCTT	165	XM_046931582.1
	ACTGTGGTGTGCTCAGAATCC
IL-18	GCAGTACGGCTTAGAGAAA	198	XM_046932263.1
	GTACATTCCACTGCCAGATT
GADPH	GCCATCACAGCCACACAGAAGA	206	NM_204305.2
	CGGCAGGTCAGGTCAACAACAG
GSDMD	CTGCTCTGATCTACAGGTTGTTCCG	79	XM_046911140
	GCTCAGTGCCAAGGTGCCATC

组别Group	AST	ALT
7 d-对照组7 d-CG	1.600±0.100^a	57.625±8.922^a
7 d-低硼组7 d-LBG	2.250±0.208^a	139.925±15.897^b
7 d-高硼组7 d-HBG	3.100±0.200^b	206.825±1.053^b
14 d-对照组14 d-CG	1.650±0.250^a	52.633±14.836^a
14 d-低硼组14 d-LBG	2.575±0.171^b	230.725±17.814^b
14 d-高硼组14 d-HBG	3.967±1.159^b	282.575±23.517^b

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