Institute of Animal Nutrition and Nutrition Physiology, Justus-Liebig-University Giessen, Giessen, Germany.
PLoS One. 2020 Feb 10;15(2):e0228945. doi: 10.1371/journal.pone.0228945. eCollection 2020.
Recent studies indicated that intramammary administration of active vitamin D3 hormone (1,25D3) inhibits the inflammatory process associated with mastitis. We hypothesized that attenuation of endoplasmic reticulum (ER) stress by 1,25D3 in mammary epithelial cells (MECs) is an important cellular mechanism contributing to this beneficial effect of intramammary treatment with 1,25D3. To test this hypothesis, the effect of 1,25D3 was studied on induction of ER stress in a transformed human MEC line, MCF-7 cells. Treatment with two different ER stress inducers, thapsigargin (TG) and tunicamycin (TM), caused a dose-dependent induction of ER stress as evident from up-regulation of protein kinase RNA-like ER kinase (PERK), heat shock protein family A (Hsp70) member 5 (HSPA5), activating transcription factor (ATF4), ATF6, DNA damage inducible transcript 3 (DDIT3) and spliced X-box binding protein 1 (sXBP1) and impaired cell viability and decreased expression of vitamin D receptor (VDR) in MCF-7 cells (P < 0.05). Treatment with 1,25D3 (100 nM) inhibited TG (10 nM)- and TM (1 μg/mL)-induced mRNA and/or protein levels of ATF4, ATF6, DDIT3 and HSPA5 in MCF-7 cells (P < 0.05). In addition, 1,25D3 (100 nM) antagonized the effect of TG (10 nM) and TM (1 μg/mL) on mRNA and protein levels of VDR and mRNA levels of genes involved in production and degradation of 1,25D3 in MCF-7 cells (P < 0.05). Moreover, 1,25D3 (100 nM) inhibited nuclear factor-κB (NF-κB) activation in response to TM (10 nM) and TG (1 μg/mL) in MCF-7 cells. In conclusion, the present findings show that 1,25D3 is effective in attenuating ER stress and the NF-κB-driven inflammatory response in MCF-7 cells. This indicates that attenuation of ER stress by 1,25D3 in MECs may contribute to the recently observed inhibitory effect of intramammary treatment of dairy cows with 1,25D3 on the inflammatory process associated with mastitis.
最近的研究表明，向乳腺内施用活性维生素 D3 激素（1,25D3）可抑制与乳腺炎相关的炎症过程。我们假设，1,25D3 可减轻乳腺上皮细胞（MEC）中的内质网（ER）应激，这是 1,25D3 对乳腺内治疗有益效果的重要细胞机制。为了验证这一假设，我们研究了 1,25D3 对转化的人乳腺上皮细胞系 MCF-7 细胞中 ER 应激诱导的作用。用两种不同的 ER 应激诱导剂，他莫昔芬（TG）和衣霉素（TM）处理，从蛋白激酶 RNA 样内质网激酶（PERK）、热休克蛋白家族 A（Hsp70）成员 5（HSPA5）、激活转录因子（ATF4）、ATF6、DNA 损伤诱导转录物 3（DDIT3）和剪接 X 盒结合蛋白 1（sXBP1）的上调，以及细胞活力的降低和维生素 D 受体（VDR）表达的下调，明显看出 ER 应激的剂量依赖性诱导（P<0.05）。用 1,25D3（100 nM）处理可抑制 MCF-7 细胞中 TG（10 nM）和 TM（1μg/mL）诱导的 ATF4、ATF6、DDIT3 和 HSPA5 的 mRNA 和/或蛋白水平（P<0.05）。此外，1,25D3（100 nM）拮抗了 TG（10 nM）和 TM（1μg/mL）对 MCF-7 细胞中 VDR 的 mRNA 和蛋白水平以及参与 1,25D3 产生和降解的基因的 mRNA 水平的影响（P<0.05）。此外，1,25D3（100 nM）抑制了 MCF-7 细胞中 TM（10 nM）和 TG（1μg/mL）对核因子-κB（NF-κB）激活的作用。总之，本研究结果表明，1,25D3 可有效减轻 MCF-7 细胞中的 ER 应激和 NF-κB 驱动的炎症反应。这表明，1,25D3 可减轻 MEC 中的 ER 应激，这可能有助于最近观察到的用 1,25D3 对奶牛乳腺内治疗对与乳腺炎相关的炎症过程的抑制作用。
