Arnica montana stimulates extracellular matrix gene expression in human macrophages differentiated to wound-healing phenotype.. Homo sapiens

Arnica m. effects were associated with a purported anti-inflammatory and tissue healing actions after trauma, bruises, or tissue injuries, but its cellular and molecular mechanisms are largely unknown. Here Arnica m. dilutions were tested using an in vitro model of macrophages polarized towards a “wound-healing” phenotype. The monocyte-macrophage human THP-1 cell line was cultured and differentiated with phorbol-myristate acetate and Interleukin-4, then exposed for 24 h to Arnica m. centesimal (c) dilutions 2c, 3c, 5c,9c, 15c or Control. None of these treatments affected cell viability. A total of 20 genes were differentially expressed comparing cells treated with Arnica m. 2c with those treated with Control only. Of these, 7 genes were up-regulated and 13 were down-regulated. Functional gene enrichment analysis showed that the most significantly upregulated function concerned 4 genes with a conserved site of EGF-like region (p<0.001) and three genes of proteinaceous extracellular matrix, including heparin sulphate proteoglycan 2 (HSPG2), fibrillin 2 (FBN2), and fibronectin (FN1) (p <0.01). Protein assay in supernatants confirmed a statistically significant increase of fibronectin production in Arnica m. 2c treated cells (p<0.05). Pooled extracts of cells treated with increasing dilutions of Arnica m. (3c, 5c, 15c) showed up-regulation of the same group of genes although with lower effect size. The down-regulated transcripts derive from mitochondrial genes coding for some components of electron transport chain. These findings provide new insights into the action of Arnica m. in tissue healing and repair, identifying increased fibronectin production by macrophages as a major therapeutic target. Overall design: Expression analysis of differentiated THP-1 cell line exposed at Arnica m. centesimal (c) dilution 2c, plus control non-exposed line both in 5 replicates.

山金车属（Arnica m.）的相关效应被认为与创伤、挫伤或组织损伤后据称具有的抗炎及组织修复作用存在关联，但其细胞与分子机制尚未明确。本研究采用极化至“创伤愈合”表型的巨噬细胞体外模型，对山金车母液的百分度（centesimal, c）稀释液进行了实验检测。将人单核细胞-巨噬细胞THP-1细胞系采用佛波醇-肉豆蔻酸酯-乙酸酯（phorbol-myristate acetate）与白细胞介素-4（Interleukin-4）进行培养并诱导分化，随后分别以山金车母液的2c、3c、5c、9c、15c百分度稀释液或对照液处理24小时。上述所有处理均未对细胞活力产生影响。仅对比山金车2c处理组与对照组细胞时，共有20个基因呈现差异表达，其中7个基因上调，13个基因下调。功能基因富集分析结果显示，上调最为显著的功能模块涉及4个携带EGF样保守结构域的基因（p<0.001），以及3个编码细胞外基质蛋白的基因：硫酸乙酰肝素蛋白聚糖2（heparin sulphate proteoglycan 2, HSPG2）、原纤维蛋白2（fibrillin 2, FBN2）与纤连蛋白（fibronectin, FN1）（p<0.01）。细胞上清液的蛋白检测证实，经山金车2c处理的细胞中纤连蛋白的分泌量存在统计学意义上的显著升高（p<0.05）。对经梯度稀释的山金车母液（3c、5c、15c）处理的细胞混合提取物进行检测，结果显示同一组基因同样呈现上调，但效应幅度相对较低。下调的转录本均源自编码电子传递链（electron transport chain）部分组分的线粒体基因。本研究结果为山金车属在组织愈合与修复中的作用机制提供了全新见解，明确巨噬细胞分泌纤连蛋白的增强效应可作为其核心治疗靶点。整体实验设计：对经诱导分化的THP-1细胞系分别施以山金车2c百分度稀释液处理与未处理对照，每组均设置5个生物学重复。