Arctigenin Confers Neuroprotection Against Mechanical Trauma Injury in Human Neuroblastoma SH-SY5Y Cells by Regulating miRNA-16 and miRNA-199a Expression to Alleviate Inflammation

Abstract

Mechanical trauma injury is a severe insult to neural cells. Subsequent secondary injury involves the release of inflammatory factors that have dramatic consequences for undamaged cells, leading to normal cell death after the initial injury. The present study investigated the capacity for arctigenin (ARC) to prevent secondary effects and evaluated the mechanism underlying the action of microRNA (miRNA)-199a and miRNA-16 in a mechanical trauma injury (MTI) model using SH-SY5Y cells in vitro. SH-SY5Y cells are often applied to in vitro models of neuronal function and differentiation. Recently, miRNAs have been demonstrated to play a crucial role in NF-κB and cholinergic signaling, which can regulate inflammation. The cell model was established by scratch-induced injury of human SH-SY5Y cells, which mimics the characteristics of MTI. A cell counting kit-8 (CCK-8), terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), and immunocytochemistry were used to measure cell viability. Enzyme-linked immunosorbent assay (ELISA) was used to evaluate the inflammatory cytokine and cholinesterase (CHE) content. The lactate dehydrogenase (LDH) content was measured to assess the degree of cell injury. The mRNA levels were measured by RT-PCR to analyze ARC's mechanism of action. miRNA inhibitors and mimics were used to inhibit and strengthen the expression of miRNAs. Protein expression was detected by western blotting analysis. ARC treatment reduced the TNF-α and IL-6 levels as well as the number of TUNEL+ apoptotic SH-SY5Y cells surrounding the scratch and increased the IL-10 level compared to the controls. ARC attenuated the increase of the cell damage degree and LDH content induced by scratching, indicating increased cell survival. Mechanistic studies showed that ARC upregulated the miRNA-16 and miRNA-199a levels to reduce upstream protein (IKKα and IKKβ) expression and inhibit NF-κB signaling pathway activity; moreover, the increased miRNA-199a suppresses cholinesterases to increase cholinergic signaling, resulting in decreased expression of proinflammatory cytokines. ARC treatment confers protection for SH-SY5Y cells through positive regulation of miRNA expression, thereby reducing the inflammatory response. In turn, these effects accelerate injury repair in the scratch-induced injury model. These results might provide insights into the pharmacological role of ARC in anti-inflammation and neuroprotection in neural cells.

Arctigenin protects against neuronal hearing loss by promoting neural stem cell survival and differentiation

“…Next, we found that proliferation of the cultured NSCs in the BrdU incorporation assay almost exactly recapitulated the trend as MTT assay, in which 0.5 μM ARC displayed the highest promotional effect on proliferation of the cultured NSCs. These results demonstrated that ARC increased viability as well as proliferation ability of NSCs derived from mouse cochlear in a dose‐dependent manner, with the optimal dosage at 0.5 μM in our study (Figure d), consistent with reported effective concentration of ARC on other cell lines in previous studies (Song et al, ; Zhang et al, ).…”

“…Moreover, ARC demonstrates neuroprotective effect on both primary neuron neurotoxicity, as well as memory and learning defects in mice induced by scopolamine with neural degenerative disease (Jang et al, ; Lee et al, ). In addition, ARC has also been reported to alleviate inflammation and confer neuroprotection against mechanical trauma injury in human neuroblastoma cells (Song et al, ), as well as in mouse primary neurons (Zhang et al, ). However, the effect of ARC in protecting against neuronal hearing loss has not been investigated.…”

Exosomes Derived From Low‐Intensity Pulsed Ultrasound‐Treated Dendritic Cells Suppress Tumor Necrosis Factor–Induced Endothelial Inflammation

“…miR‐16 targets IκB kinase (IKK)‐α and IKKβ to inhibit expression of these 2 proteins and consequently blocks the activity of NF‐κB signaling pathway . miR‐21 targets proinflammatory genes’ mRNA to limit the activation of NF‐κB .…”

“…Then NF‐κB proteins are translocated from the cytoplasm into the nucleus to activate the target gene expression . Low‐intensity pulsed ultrasonography–stimulated BMDC exosomes were reported to contain high levels of miR‐16, which targeted IKKα and IKKβ, then enhanced their degradation . Thus, IκBα levels increased by miR‐16 sequestered more NF‐κB proteins in the cytoplasm and alleviated the activity of the signaling pathway.…”

3′‐Desmethylarctigenin induces G2/M cell cycle arrest and apoptosis through reactive oxygen species generation in hepatocarcinoma cells

“…Arctigenin is a major biologically active component of Arctium lappa L. ( A. lappa ), which is a popularly used vegetable as well as herbal medicine (Liu et al, ). Arctigenin exhibits distinct biological activities, including antiviral (Dias et al, ; Hayashi, Narutaki, Nagaoka, Hayashi, & Uesato, ), neuroprotection (Song et al, ; Zhu et al, ), antidiabetic (Huang et al, ; Zeng et al, ), and anticancer (Chen et al, ; Huang et al, ; Hsieh et al, ; Jeong, Hong, Jeong, & Koo, ; Lei, Gan, Zhao, Yu, & Hu, ; Li, Liang, Tian, & Hu, ; Maimaitili et al, ; Wang, Solorzano, et al, ; Susanti et al, ). A growing body of literature documents that arctigenin can induce cell cycle arrests, apoptosis, autophagy, and antimetastasis in multiple types of cancer cells, including prostate (Wang, Solorzano, et al, ), breast (Feng et al, ; Hsieh et al, ; Lou, Zhu, Zhao, Zhu, & Zhao, ; Maxwell, Lee, Kim, & Nam, ), lung (Lei et al, ), liver (Susanti et al, ), gastric (Jeong et al, ), ovarian (Huang et al, ), colon cancer (Li et al, ), and glioma (Maimaitili et al, ).…”