Department of Biomaterials Science, College of Natural Resources and Life Science/Life and Industry Convergence Research Institute, Pusan National University, Miryang 50463, Republic of Korea.
Department of Horticultural Life Sciences, College of Natural Resources and Life Science/Life and Industry Convergence Research Institute, Pusan National University, Miryang 50463, Republic of Korea.
Mol Med Rep. 2020 Dec;22(6):4685-4695. doi: 10.3892/mmr.2020.11563. Epub 2020 Oct 6.
Mulberry leaves have antioxidant activity and anti‑inflammatory effects in several types of cells. However, the efficacy of mulberry leaves fermented with Cordyceps militaris remains unknown. Therefore, the present study aimed to investigate whether the ethanol extracts of mulberry leaves fermented with C. militaris (EMfC) can prevent lipopolysaccharide (LPS)‑induced inflammation and autophagy in macrophages. To achieve this, RAW264.7 cells pretreated with three different dose of EMfCs were subsequently stimulated with LPS, and examined for alterations in the regulatory factors of inflammatory responses and key parameters of the autophagy signaling pathway. EMfC treatment inhibited the generation of reactive oxidative species; however, significant activity was observed for 2,2‑diphenyl‑1‑picrylhydrazyl (DPPH) radical scavenging (IC50=579.6703 mg/ml). Most regulatory factors in inflammatory responses were significantly inhibited following treatment with EMfC, without any significant cellular toxicity. EMfC‑treated groups exhibited marked suppression of nitrogen oxide (NO) levels, mRNA expression levels of iNOS/COX‑2, levels of all inflammatory cytokines (TNF‑α, IL‑1β and IL‑6) and phosphorylation of MAPK members, as well as recovery of cell cycle progression. Furthermore, similar effects were observed in the LPS‑induced autophagy signaling pathway of RAW264.7 cells. The expression levels of microtubule‑associated protein 1A/1B‑light chain 3 (LC3) and Beclin exhibited a dose‑dependent decrease in the EMfC+LPS‑treated groups compared with in the Vehicle+LPS‑treated group, whereas the phosphorylation of PI3K and mTOR were enhanced in a dose‑dependent manner in the same groups. Overall, the results of the present study provide evidence that exposure to EMfC protects against LPS‑induced inflammation and autophagy in RAW264.7 cells. These results indicated that EMfC is a potential candidate for treatment of inflammatory diseases.
桑叶具有抗氧化活性和多种细胞的抗炎作用。然而，桑椹与蛹虫草发酵后的功效尚不清楚。因此，本研究旨在探讨桑椹与蛹虫草发酵的乙醇提取物（EMfC）是否可以预防巨噬细胞中脂多糖（LPS）诱导的炎症和自噬。为了实现这一目标，用三种不同剂量的 EMfC 预处理 RAW264.7 细胞，然后用 LPS 刺激细胞，并检测炎症反应调节因子和自噬信号通路关键参数的变化。EMfC 处理抑制活性氧的产生；然而，对 2,2-二苯基-1-苦基肼（DPPH）自由基清除（IC50=579.6703mg/ml）具有显著活性。用 EMfC 处理后，大多数炎症反应调节因子显著受到抑制，同时没有明显的细胞毒性。EMfC 处理组表现出对一氧化氮（NO）水平、iNOS/COX-2 基因表达水平、所有炎症细胞因子（TNF-α、IL-1β 和 IL-6）和丝裂原活化蛋白激酶（MAPK）成员磷酸化水平的显著抑制，以及细胞周期进程的恢复。此外，在 LPS 诱导的 RAW264.7 细胞自噬信号通路中也观察到类似的作用。与 Vehicle+LPS 处理组相比，EMfC+LPS 处理组微管相关蛋白 1A/1B-轻链 3（LC3）和 Beclin 的表达水平呈剂量依赖性降低，而同一组中 PI3K 和 mTOR 的磷酸化呈剂量依赖性增强。总之，本研究结果表明，暴露于 EMfC 可防止 LPS 诱导的 RAW264.7 细胞炎症和自噬。这些结果表明，EMfC 是治疗炎症性疾病的潜在候选药物。
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