Yanthi Nova Dilla, Anggraeni Anneke, Said Syahruddin, Saputra Sugiyono, Soejoedono Retno Damayanti, Muladno Muladno, Herlina Nina, Fauziah Ima, Nugroho Herjuno Ari, Nasrulloh Mukh Fajar, Tiffarent Rida
Research Center for Applied Microbiology, Research Organization for Life Sciences and Environment, National Research and Innovation Agency, Cibinong Science Center, Jl. Raya Jakarta - Bogor KM. 46, Cibinong, Bogor 16911, West Java, Indonesia.
Research Center for Animal Husbandry, Research Organization for Agriculture and Food, National Research and Innovation Agency, Cibinong Science Center, Jl. Raya Jakarta - Bogor KM. 46, Cibinong, Bogor 16911, West Java, Indonesia.
Vet World. 2025 Apr;18(4):1014-1024. doi: 10.14202/vetworld.2025.1014-1024. Epub 2025 Apr 25.
Mastitis remains a major health challenge in dairy cattle, often caused by Gram-positive pathogens. Toll-like receptors (TLRs) and chemokine receptors (CXCRs) play essential roles in the innate immune response of mammary epithelial cells (MECs). However, the differential expression of these genes in response to specific mastitis-causing spp. has not been comprehensively evaluated. This study aimed to characterize the temporal gene expression patterns of TLR and CXCR family members in murine mammary epithelial HC11 cells exposed to and , thereby providing insights into their immunological roles in mastitis pathogenesis.
HC11 cells were cultured and infected with and (5 × 10 colony-forming units/mL) and incubated at 37°C with 95% O and 5% CO for 48 h in RPMI 1640 medium supplemented with serum and antibiotics. Gene expression of interleukin (IL)-6, IL-8, TLR2, TLR4, IL-1 alpha (IL-1α), and CXCR1 was evaluated by quantitative real-time polymerase chain reaction at 0, 6, 12, 24, 48, and 72 h post-infection. Expression levels were normalized to glyceraldehyde-3-phosphate dehydrogenase and analyzed using ΔCt methods and Spearman correlation.
TLR2 exhibited a biphasic expression pattern, with early upregulation followed by suppression, while TLR4 showed higher expression in response to than . IL-6 displayed prolonged expression under challenge but was transient under exposure. IL-1α showed consistent expression across both bacterial challenges, suggesting its potential as a stable biomarker for mastitis susceptibility. CXCR1 exhibited delayed but sustained expression, indicative of its role in secondary neutrophil recruitment. IL-8 expression correlated with early immune activation and chemotactic signaling.
The immune response of HC11 MECs to Gram-positive bacterial infection is gene- and pathogen-specific. and genes show distinct temporal profiles, underscoring their utility in understanding epithelial-driven immune defense. These findings provide molecular insights into mastitis pathogenesis and identify IL-1α, IL-6, and CXCR1 as promising targets for genetic selection and therapeutic intervention.
乳腺炎仍是奶牛养殖中的一项重大健康挑战，通常由革兰氏阳性病原体引起。Toll样受体（TLRs）和趋化因子受体（CXCRs）在乳腺上皮细胞（MECs）的固有免疫反应中发挥着重要作用。然而，这些基因在应对特定致乳腺炎病原体时的差异表达尚未得到全面评估。本研究旨在描述暴露于[具体病原体1]和[具体病原体2]的小鼠乳腺上皮HC11细胞中TLR和CXCR家族成员的基因表达时间模式，从而深入了解它们在乳腺炎发病机制中的免疫作用。
培养HC11细胞，并用[具体病原体1]和[具体病原体2]（5×10菌落形成单位/毫升）进行感染，在补充有血清和抗生素的RPMI 1640培养基中于37°C、95% O₂和5% CO₂条件下孵育48小时。在感染后0、6、12、24、48和72小时，通过定量实时聚合酶链反应评估白细胞介素（IL）-6、IL-8、TLR2、TLR4、IL-1α和CXCR1的基因表达。将表达水平标准化为甘油醛-3-磷酸脱氢酶，并使用ΔCt方法和Spearman相关性进行分析。
TLR2呈现双相表达模式，早期上调随后下调，而TLR4对[具体病原体1]的反应比[具体病原体2]表现出更高的表达。IL-6在[具体病原体1]刺激下呈现延长表达，但在[具体病原体2]暴露下是短暂的。IL-1α在两种细菌刺激下均表现出一致的表达，表明其作为乳腺炎易感性稳定生物标志物的潜力。CXCR1表现出延迟但持续的表达，表明其在次级中性粒细胞募集中的作用。IL-8表达与早期免疫激活和趋化信号相关。
HC11 MECs对革兰氏阳性细菌感染的免疫反应具有基因和病原体特异性。[具体病原体1]和[具体病原体2]基因显示出不同的时间特征，突出了它们在理解上皮驱动的免疫防御中的作用。这些发现为乳腺炎发病机制提供了分子见解，并确定IL-1α、IL-6和CXCR1为遗传选择和治疗干预的有前景靶点。
