蜂胶调控紧密连接蛋白对奶牛乳腺上皮细胞炎性损伤的保护机制

Bovine mastitis is one of the most serious diseases caused by a variety of factors, which is a costly disease for dairy producers. Prevention and treatment for bovine mastitis is a systematic project. Propolis is widely used in clinical veterinary medicine in controlling diseases of livestock and poultry. Nevertheless, the material basis and the mechanisms for the efficacy of propolis against bovine mastitis remains elusive, which restrict its application. Our previous studies found propolis has inhibitive effects against inflammation and oxidative stress in the bovine mammary epithelial cells. We also found propolis regulate tight junction proteins in the gut. Therefore, we postulated that the beneficial effects by propolis against mastitis might attribute to its regulation on the milk-blood barrier and tight junction proteins. To verify this hypothesis, this project try to establish the in vitro barrier system using the bovine epithelial cells and to induce inflammatory injuries with loss of tight junctions. Then we plan to apply the propolis and its isolated bioactive constituents to protect bovine epithelial cells against mastitis pathogen challenges. Based on evaluating the lactation activities and intercellular barrier structures, we try to explore the bioactive responsible constituents of propolis against mastitis in the bovine epithelial cells. Based on potential therapeutic targets, like tight junction proteins, we plan to use molecular biology techniques to clarify the underlying mechanisms of these protective effects. Based on animal studies using dairy cattle, we further verify the therapeutic mechanisms of propolis against bovine mastitis. This project provides us new insights and novel targets for the treatment/prevention of bovine mastitis using traditional Chinese medicine.

奶牛乳腺炎是一种多种原因引起的极为复杂的严重疾病，造成奶牛业重大损失，其防控是一个系统工程。蜂胶广泛用于兽医临床，但其防治奶牛乳腺炎药效物质基础及作用机制尚未阐明。我们前期研究发现蜂胶可抑制奶牛乳腺上皮细胞炎症反应和氧化应激，并能提升肠上皮紧密连接屏障功能。由此我们推测蜂胶可能通过调控乳腺上皮细胞紧密连接蛋白保护奶牛乳血屏障，进而发挥抗炎功效。本项目拟构建奶牛乳腺上皮细胞屏障系统并建立炎性损伤紧密连接模型，采用蜂胶乙醇提取物及其主要活性成分进行保护性干预和系统筛查；以细胞泌乳活性和胞间屏障结构功能为评价指标，系统揭示和挖掘蜂胶及其活性成分在乳腺炎发生发展过程中对乳腺上皮细胞紧密连接的保护作用；以紧密连接蛋白为靶标结合分子生物学干预阐明蜂胶对奶牛乳腺炎性损伤的保护作用机制；采用奶牛乳腺炎动物实验验证蜂胶疗效机理，为中药防治奶牛乳腺炎的研究提供新思路和新靶点。

奶牛乳腺炎是一种多种原因引起的极为复杂的严重疾病，造成奶牛业重大损失，其防控是一个系统工程。蜂胶广泛用于兽医临床，但其防治奶牛乳腺炎药效物质基础及作用机制尚未阐明。我们前期研究发现蜂胶可抑制奶牛乳腺上皮细胞炎症反应和氧化应激，并能提升肠上皮紧密连接屏障功能。由此我们推测蜂胶可能通过调控乳腺上皮细胞紧密连接蛋白保护奶牛乳血屏障，进而发挥抗炎功效。本项目基于蜂胶活性物质化学分析结果，从细胞水平探明蜂胶对细菌毒素（LPS）侵染下奶牛乳腺上皮细胞紧密连接的保护作用；从分子水平揭示紧密连接蛋白（包括Occludin, ZOs等）在抵御病原物诱导的乳腺上皮细胞损伤中的作用以及蜂胶对紧密连接蛋白的干预模式，解析乳腺上皮细胞在炎症情况下细胞上皮屏障功能所发挥的关键性调节作用；从动物水平验证了蜂胶对细菌毒素诱导的小鼠急性乳腺炎性损伤的保护效果及作用机理。本研究也进一步明确了中国蜂胶对抗细菌脂多糖诱导乳腺炎的作用效果，为中药防治奶牛乳腺炎的研究提供新思路和新靶点。