State Key Laboratory of Animal Nutrition, College of Animal Science and Technology, China Agricultural University, Yuanmingyuan West Road, Haidian District, Beijing, 100193, China.
Division of Nutritional Sciences, University of Illinois at Urbana-Champaign, 1207 W. Gregory Ave., Urbana, IL, 61801, USA.
J Nutr Biochem. 2017 May;43:18-26. doi: 10.1016/j.jnutbio.2017.01.013. Epub 2017 Jan 31.
Zinc plays an important role in maintaining intestinal barrier function as well as modulating cellular signaling recognition and protein kinase activities. The phosphatidylinositol 3-kinase (PI3K) cascade has been demonstrated to affect intercellular integrity and tight junction (TJ) proteins. The current study investigated the hypothesis that zinc regulates intestinal intercellular junction integrity through the PI3K/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) pathway. A transwell model of Caco-2 cell was incubated with 0, 50 and 100 μM of zinc at various time points. Transepithelial electrical resistance (TEER), paracellular permeability, TJ proteins, cell proliferation, differentiation and cell damage were measured. Compared with controls, 50 and 100 μM of zinc increased cell growth at 6, 12 and 24 h and the expression of proliferating cell nuclear antigen at 24 h. Zinc (100 μM) significantly elevated TEER at 6-24 h and reduced TJ permeability at 24 h, accompanied by the up-regulation of alkaline phosphatase (AP) activity and zonula occludens (ZO)-1 expression. In addition, zinc (100 μM) affected the PI3K/AKT/mTOR pathway by stimulating phosphorylation of AKT and the downstream target mTOR. Inhibition of PI3K signaling by LY294002 counteracted zinc promotion, as shown by a decrease in AP activity, TEER, the abundance of ZO-1 and phosphorylation of AKT and mTOR. Additionally, TJ permeability and the expression of caspase-3 and LC3II (markers of cell damage) were increased by addition of PI3K inhibitor. In conclusion, the activation of PI3K/AKT/mTOR signaling by zinc is involved in improving intestinal barrier function by enhancing cell differentiation and expression of TJ protein ZO-1.
锌在维持肠道屏障功能以及调节细胞信号识别和蛋白激酶活性方面发挥着重要作用。已证实磷脂酰肌醇 3-激酶 (PI3K) 级联反应影响细胞间的完整性和紧密连接 (TJ) 蛋白。本研究提出假设，即锌通过 PI3K/蛋白激酶 B (AKT)/哺乳动物雷帕霉素靶蛋白 (mTOR) 通路调节肠道细胞间连接的完整性。用不同浓度的锌（0、50 和 100μM）孵育 Caco-2 细胞的 Transwell 模型，在不同时间点测量跨上皮电阻 (TEER)、旁细胞通透性、TJ 蛋白、细胞增殖、分化和细胞损伤。与对照组相比，50 和 100μM 的锌在 6、12 和 24 小时增加了细胞生长，在 24 小时增加了增殖细胞核抗原的表达。锌（100μM）在 6-24 小时显著提高 TEER，在 24 小时降低 TJ 通透性，同时增加碱性磷酸酶 (AP) 活性和紧密连接蛋白-1 (ZO-1) 的表达。此外，锌（100μM）通过刺激 AKT 和下游靶标 mTOR 的磷酸化影响 PI3K/AKT/mTOR 通路。LY294002 抑制 PI3K 信号通路可减弱锌的促进作用，表现为 AP 活性、TEER、ZO-1 的丰度以及 AKT 和 mTOR 的磷酸化降低。此外，TJ 通透性和 caspase-3 和 LC3II（细胞损伤标志物）的表达增加是由 PI3K 抑制剂的加入引起的。总之，锌激活 PI3K/AKT/mTOR 信号通路通过增强细胞分化和 TJ 蛋白 ZO-1 的表达来改善肠道屏障功能。
