亚急性反刍酸中毒（SARA）将导致内毒素增加，对牛反刍胃上皮细胞（BREC）产生负面影响。黄酮类物质对内毒素引起的炎症具有很好的治疗效果。槲皮素是一种在水果和蔬菜中广泛存在的重要黄酮类物质，作为一种潜在的抗炎抗氧化剂引起了广泛关注。然而，槲皮素在抵抗脂多糖（LPS）引起的BREC损伤过程中的保护机制尚不清楚。本研究采用槲皮素和LPS诱导的BREC炎症模型，探讨槲皮素对LPS引起的BREC损伤的保护作用。将BREC分别用不同剂量的LPS（1、5和10 μg/mL）处理6小时或24小时，检测炎症因子的mRNA表达。实验结果表明，在1 μg/mL LPS处理6小时后，BREC中的前炎症细胞因子的mRNA高表达建立了BREC炎症模型。通过细胞计数试剂盒-8（CCK8）实验观察到80 μg/mL槲皮素对BREC生长的促进作用。槲皮素补充显著降低了炎症因子和趋化因子（尤其是肿瘤坏死因子α（TNF-α）、白细胞介素1β（IL-1β）、IL-6、CC趋化因子配体2（CCL2）、CCL20、CCL28和CXC趋化因子9（CXCL9））的表达水平。我们还通过qRT-PCR分析了相关通路的mRNA表达情况。验证实验结果表明槲皮素显著抑制了LPS处理的BREC中类胶质细胞柱状体联蛋白4（TLR4）和髓样分化原反应蛋白（MyD88）以及核因子κ-B（NF-κB）的mRNA表达。此外，西方印迹结果证实，LPS显著激活了p44/42外源性调节蛋白激酶（ERK1/2）和NF-κB的磷酸化，而添加槲皮素后此效应被逆转。为了辅助西方印迹结果，我们还通过免疫荧光法评估了p-ERK1/2和p-p65蛋白的表达情况，结果一致。因此，槲皮素抑制TLR4介导的NF-κB和MAPK信号通路的能力可能是其对LPS引起的BREC炎症反应产生抗炎作用的原因。根据这些结果，槲皮素可作为一种抗炎药物用于缓解高粒饲料引起的炎症，并为后期槲皮素的研发和利用奠定了概念基础。

Subacute rumen acidosis (SARA) will cause an increase in endotoxin, which will have a negative effect on the bovine rumen epithelial cells (BREC). Flavonoids are effective in treating inflammation caused by endotoxin. Quercetin is a vital flavonoid widely occurring in fruits and vegetables and has received significant interest as a prospective anti-inflammatory antioxidant. Nonetheless, quercetin's protective machinery against such damage to BREC induced by lipopolysaccharide (LPS) remains unclear. A combined quercetin and LPS-induced BREC inflammation model was utilized to elucidate the effect of quercetin protecting BREC from LPS-induced injury. After treating BREC with different doses of LPS (1, 5, and 10 μg/mL) for 6 h or 24 h, the mRNA expression of inflammatory factors was detected. Our experimental results show the establishment of the BREC inflammation model via mRNA high expression of pro-inflammatory cytokines in BREC following 6 h treatment with 1 µg/mL LPS. The promotive effect of 80 μg/mL quercetin on BREC growth via the cell counting kit-8 (CCK8) assay was observed. The expression of pro-inflammatory cytokines and chemokines, notably tumor necrosis factor α (TNF-α), Interleukin 1β (IL-1β), IL-6, CC-motif chemokine ligand 2 (CCL2), CCL20, CCL28, and CXC motif chemokine 9 (CXCL9), etc., was significantly reduced by quercetin supplementation. We also analyzed the mRNA detection of related pathways by qRT-PCR. Our validation studies demonstrated that quercetin markedly curbed the mRNA expression of the toll-like receptor 4 (TLR4) and myeloid differentiation primary response protein (MyD88) and the nuclear factor-κB (NF-κB) in LPS-treated BREC. In addition, western blot result outcomes confirmed, as expected, that LPS significantly activated phosphorylation of p44/42 extracellular regulated protein kinases (ERK1/2) and NF-κB. Unexpectedly, this effect was reversed by adding quercetin. To complement western blot results, we assessed p-ERK1/2 and p-p65 protein expression using immunofluorescence, which gave consistent results. Therefore, quercetin's capacity to bar the TLR4-mediated NF-κB and MAPK signaling pathways may be the cause of its anti-inflammatory effects on LPS-induced inflammatory reactions in BREC. According to these results, quercetin may be utilized as an anti-inflammatory medication to alleviate inflammation brought on by high-grain feed, and it also lays out a conceptual foundation regarding the development and utilization of quercetin in the later stage.