Laboratory of Clinical Pharmaceutics, Yokohama University of Pharmacy, Yokohama 245-0066, Kanagawa, Japan.
General Health Medical Research Center, Yokohama University of Pharmacy, Yokohama 245-0066, Kanagawa, Japan.
Int J Mol Sci. 2024 Oct 19;25(20):11249. doi: 10.3390/ijms252011249.
Lactoferrin (LF) is a multifunctional protein abundant in breast milk that modulates the functions of neural stem cells. Recent studies have demonstrated the efficacy of bovine LF (bLF) in mitigating behavioral changes; however, the molecular mechanisms on the nervous system have not yet been elucidated. The presented study aimed to characterize the molecular mechanisms of bLF on nerve extension in PC12 cells. PC12 cells were treated with 0.01-1000 µg/mL of bLF, and cell viability was determined using the cell counting kit-8 assay after treatment for 24 h. Morphometric evaluation was performed after 24 or 72 h of treatment with 50 ng/mL nerve growth factor (NGF) or 100-500 µg/mL bLF. The molecular mechanisms were investigated using Western blotting and real-time quantitative PCR. Cell viability was significantly decreased after treatment with 600-1000 µg/mL bLF for 24 h compared with the control group. Morphometric evaluation revealed neurite outgrowth after 72 h of NGF treatment, with a significant increase in neurite outgrowth after treatment with 250 µg/mL bLF. The phosphorylated p44/42 expression ratio peaked at 5 min and persisted for up to 10 min. Quantitative real-time PCR revealed a significant decrease in MAP2 expression. Our findings suggested that bLF enhanced PC12 cell neurite outgrowth to a similar extent as NGF. These effects are thought to be mediated via the TrkA receptor and activated by the phosphorylated ERK signaling pathway. Therefore, this study demonstrates that bLF promotes neurite outgrowth via a pathway similar to that of NGF.
乳铁蛋白 (LF) 是一种丰富存在于母乳中的多功能蛋白质，可调节神经干细胞的功能。最近的研究表明牛乳铁蛋白 (bLF) 可有效缓解行为变化；然而，其在神经系统上的分子机制尚未阐明。本研究旨在探究 bLF 对 PC12 细胞神经突起延伸的分子机制。将 PC12 细胞用 0.01-1000μg/mL 的 bLF 处理 24 小时，用细胞计数试剂盒-8 法测定细胞活力。用 50ng/mL 神经生长因子 (NGF) 或 100-500μg/mL bLF 处理 24 或 72 小时后进行形态计量评估。用 Western blot 和实时定量 PCR 法研究分子机制。与对照组相比，600-1000μg/mL bLF 处理 24 小时后细胞活力显著下降。形态计量评估显示 NGF 处理 72 小时后神经突起生长，250μg/mL bLF 处理后神经突起生长显著增加。磷酸化 p44/42 表达比值在 5 分钟时达到峰值，并持续至 10 分钟。定量实时 PCR 显示 MAP2 表达显著下降。我们的研究结果表明，bLF 增强 PC12 细胞的神经突起生长，与 NGF 效果相当。这些作用被认为是通过 TrkA 受体介导的，通过磷酸化 ERK 信号通路激活。因此，本研究表明 bLF 通过类似于 NGF 的途径促进神经突起生长。
