植物乳植杆菌ZG03通过其代谢物短链脂肪酸缓解斑马鱼的氧化应激

doi:10.12122/j.issn.1673-4254.2025.10.18

摘要/Abstract

摘要：

目的分析植物乳植杆菌ZG03的菌株特性并通过代谢组学结合斑马鱼模式生物探究ZG03缓解氧化应激作用机制。方法测定并绘制ZG03生长曲线，通过场发射扫描电镜观察菌体形态，进一步通过全基因组学测序技术从基因层面评估其安全性和潜在功效。通过异硫氰酸荧光素（FITC）标记植物乳植杆菌ZG03，在荧光显微镜下观察其在斑马鱼肠道中的定植情况。运用2%葡萄糖溶液诱导斑马鱼建立氧化应激模型，通过荧光显微镜观察斑马鱼尾部造血组织（CHT）处中性粒细胞的生成情况，和检测斑马鱼体内SOD活力、ROS水平以及MDA含量，评估ZG03缓解氧化应激作用。运用液相色谱-质谱技术进行靶向代谢组学检测ZG03作用斑马鱼后体内的代谢产物短链脂肪酸（SCFAs）含量，明确抗氧化应激关键代谢物。运用氧化应激模型评价关键代谢物乙酸、丙酸和己酸的缓解氧化应激作用。结果植物乳植杆菌ZG03在MRS固体培养基上呈圆形、表面光滑、湿润、乳白色单菌落，菌体呈杆状，ZG03具有丰富的糖代谢通路基因簇。与空白组相比，斑马鱼肠球、中肠和后肠部位均清晰可见FITC标记植物乳植杆菌ZG03的绿色荧光。与模型组相比，ZG03显著降低葡萄糖诱导氧化应激模型斑马鱼体内的ROS水平（P<0.05）、斑马鱼CHT处中性粒细胞的数量（P<0.001）、升高斑马鱼体内SOD活性（P<0.05），同时植物乳植杆菌ZG03在斑马鱼体内的代谢物SCFAs乙酸、丙酸、己酸含量增加（P<0.05）。SCFAs乙酸钠、丙酸钠、己酸钠能够提升氧化应激模型斑马鱼体内的SOD活性（P<0.001）。结论植物乳植杆菌ZG03通过其代谢物SCFAs乙酸、丙酸、己酸改善葡萄糖诱导模型斑马鱼的氧化应激。

关键词: 植物乳植杆菌ZG03, 短链脂肪酸, 葡萄糖, 氧化应激

Abstract:

Objective To investigate the efficacy of Lactobacillus plantarum ZG03 (L. plantarum ZG03) for ameliorating oxidative stress in zebrafish. Methods We evaluated the growth pattern of L. plantarum ZG03, observed its morphology using field emission scanning electron microscopy, and assessed its safety and potential efficacy with whole-genome sequencing for genetic analysis. FITC-labeled ZG03 was used to observe its intestinal colonization in zebrafish. In a zebrafish model of 2% glucose-induced oxidative stress, the effect of ZG03 was evaluated by assessing the changes in neutrophils in the caudal hematopoietic tissue (CHT), superoxide dismutase (SOD) activity, reactive oxygen species (ROS) levels, and malondialdehyde (MDA) content. Liquid chromatography-mass spectrometry-based targeted metabolomics was used for analyzing short-chain fatty acids (SCFAs) in the zebrafish, and the antioxidant effects of the key metabolites (acetate, propionate, and caproate) were tested. Results On MRS agar, L. plantarum ZG03 formed circular, smooth, moist, and milky-white colonies with a rod-shaped cell morphology. Genomic analysis revealed abundant sugar metabolism gene clusters. After inoculation of FITC-labeled L. plantarum ZG03 in zebrafish, green fluorescence was clearly observed in the intestinal bulb, mid-intestine, and hind intestine. In zebrafish with glucose-induced oxidative stress, L. plantarum ZG03 significantly reduced ROS levels and the number of neutrophils in the CHT with increased SOD activity. L.plantarum ZG03 significantly increased the content of SCFAs including acetic acid, propionic acid, and caproic acid in zebrafish metabolites. In addition, sodium acetate, sodium propionate, and sodium caproate in the SCFAs significantly increased SOD activity in the zebrafish models. Conclusion L. plantarum ZG03 ameliorates oxidative stress in a glucose-induced zebrafish model through its metabolites, particularly the SCFAs including acetic acid, propionic acid and caproic acid.

Key words: Lactobacillus plantarum ZG03, short-chain fatty acids, glucose, oxidative stress

林淑娴, 郭丽娜, 马燕, 熊尧, 何盈犀, 许欣筑, 盛雯, 许素哗, 邱峰. 植物乳植杆菌ZG03通过其代谢物短链脂肪酸缓解斑马鱼的氧化应激[J]. 南方医科大学学报, 2025, 45(10): 2223-2230.

Shuxian LIN, Lina GUO, Yan MA, Yao XIONG, Yingxi HE, Xinzhu XU, Wen SHENG, Suhua XU, Feng QIU. Lactobacillus plantarum ZG03 alleviates oxidative stress via its metabolites short-chain fatty acids[J]. Journal of Southern Medical University, 2025, 45(10): 2223-2230.

图/表 8

图1 植物乳植杆菌ZG03菌株生长特性

Fig.1 Growth characteristics of L. plantarum ZG03. A: Growth curve of ZG03. B: Colony morphology of ZG03. C: Field emission scanning electron microscopy (FESEM) images of ZG03.

图2 植物乳植杆菌ZG03基因组特征

Fig.2 Genomic characteristics of L. plantarum ZG03. A: Genomic circular map (from outer to inner circles): The first circle represents genomic position information; the second circle shows GC content; the third circle indicates protein-coding genes on the positive strand (marked in red); the fourth circle denotes protein-coding genes on the negative strand (marked in green); the fifth circle displays non-coding elements on the positive strand (marked in blue); the sixth circle represents non-coding elements on the negative strand (marked in purple); and the seventh circle shows long fragment repeat sequences within the genome (marked in orange). B: KEGG enrichment analysis. C: COG enrichment analysis.

表1 植物乳植杆菌ZG03基因组信息

Tab.1 Genomic information of L. plantarum ZG03

Feature	Value
Size (bp)	3 613 757
G+C content (%)	46.42
Coding region (%)	80.70
Total genes	3 354
RNA genes	123
rRNA genes	16
tRNA genes	70
Protein-coding genes	3 231
Protein coding genes with enzymes	1 099
Genes with signal petides	176
Genes with transmembrane helices	911

图3 植物乳植杆菌ZG03在斑马鱼肠道定植情况

Fig.3 Colonization by L. plantarum ZG03 in the intestinal tract of zebrafish. ***P<0.001 vs control group.

图4 斑马鱼尾部造血组织中性粒细胞的生成情况

Fig.4 Number of neutrophils in the caudal hematopoietic tissue of zebrafish (n=10). A: Representative images of neutrophils (dashed box) in zebrafish. B: Quantitative analysis of neutrophil numbers in zebrafish. ###P<0.001 vs control group; ***P<0.001 vs model group.

图5 植物乳植杆菌ZG03缓解葡萄糖诱导斑马鱼体内的氧化应激

Fig.5 Effect of L. plantarum ZG03 on glucose-induced oxidative stress in zebrafish. A: Representative images of ROS staining in zebrafish. B: Statistical analysis of relative ROS levels (n=6). C: SOD activity in zebrafish (n=3). D: MDA levels in zebrafish (n=3). ##P<0.01 vs control group; *P<0.05, **P<0.01, ***P<0.001 vs model group.

图6 斑马鱼体内代谢物短链脂肪酸含量

Fig.6 Content of short-chain fatty acid metabolites in zebrafish (n=3). A: Acetic acid content in zebrafish. B: Propionic acid content in zebrafish. C: Hexanoic acid content in zebrafish. ##P<0.01, ###P<0.001vs control group; *P<0.05 vs model group.

图7 斑马鱼体内SOD活性统计图

Fig.7 Statistical chart of superoxide dismutase (SOD) activity in zebrafish (n=3). ##P<0.001 vs control group,***P<0.001 vs model group.

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