Ascorbic acid 6-palmitate modulates microglia M1/M2 polarization in lipopolysaccharide-stimulated BV-2 cells via PERK/elF2α mediated endoplasmic reticulum stress.

BACKGROUND: Neuroinflammation-mediated microglia polarization is a major process in various central nervous system (CNS) diseases. Endoplasmic reticulum (ER) stress contributes to the inflammatory signals as well as to microglia polarization in lipopolysaccharide (LPS) induced neuroinflammation. Ascorbic acid 6-palmitate (L-AP) has been broadly used as a dietary antioxidant in foods and demonstrated a strong inhibitory effect on 5-LOX; however, the specific anti-inflammation mechanisms remain unclear. In this study, we investigated the effects and possible mechanisms of L-AP on LPS-induced neuroinflammation in BV-2 cells. METHODS: Immortalized murine microglia cell line BV-2 cells were employed to assess the effect of L-AP to modulate microglia M1/M2 polarization in vivo, and the molecular mechanism was evaluated by qRT-PCR and Western blotting analysis. Molecular docking was used to predict the binding activity of L-AP with protein kinase R-like ER kinase (PERK). RESULTS: L-AP at 62.5 µM significantly modulated LPS-induced microglia M1/M2 polarization (increases of interleukin (IL)-10 and arginase-1 (Arg-1) transcriptions) independent of cell growth. Besides, L-AP at 62.5 µM significantly down-regulated glucose-regulated protein 78 (GRP78) and CCAAT/enhancer-binding homologous protein (CHOP) mRNA levels. Similar data were shown in the tunicamycin (TM) induced ER stress cells model. Moreover, the protective effect of L-AP on TM-induced microglia M1/M2 polarization was similar to that of 4-phenyl butyric acid (4-PBA), the ER stress inhibitor. Molecular docking results indicated L-AP might directly bind with PERK, with a binding affinity of -7.7 kcal/mol. A further study unveiled that L-AP notably inhibited LPS-induced PERK/ eukaryotic initiation factor 2α (elf2α) activation. CONCLUSION: Together, this study revealed that L-AP possessed its effect on the reconstruction of microglia M1/M2 polarization balance in LPS-stimulated BV-2 cells via modulating PERK/elF2α mediated ER stress.

MANF: A Novel Endoplasmic Reticulum Stress Response Protein-The Role in Neurological and Metabolic Disorders.

The mesencephalic astrocyte-derived neurotrophic factor (MANF), also named as arginine-rich protein (ARP) or arginine-rich mutated in early-stage tumors (ARMET), is a novel evolutionary conserved protein related to unfolded protein response. Growing evidence suggests that MANF critically involves in many ER stress-related diseases with a protective effect. Here, we review the function of MANF based on its structure in neurological and metabolic disorders and summarize its potential applications in disease diagnosis and therapies.

Morphine-induced RACK1-dependent autophagy in immortalized neuronal cell lines.

BACKGROUND AND PURPOSE: Autophagy is a critical cellular catabolic process in cell homoeostasis and brain function. Recent studies indicate that receptor for activated C kinase 1 (RACK1) is involved in autophagosome formation in Drosophila and mice, and that it plays an essential role in morphine-associated memory. However, the exact mechanism of the role of RACK1 in morphine-induced autophagy is not fully understood. EXPERIMENTAL APPROACH: SH-SY5Y cells were cultured and morphine, rapamycin, 3-methyladenine and RACK1 siRNA were used to evaluate the regulation of RACK1 protein in autophagy. Western blotting and immunofluorescence were used to assess protein expression. KEY RESULTS: Activation of autophagy (i.e. autophagosome accumulation and an increase in the LC3-II/LC3-I ratio) induced by morphine contributes to the maintenance of conditioned place preference (CPP) memory in mice. Moreover, morphine treatment significantly increased Beclin-1 expression and decreased the p-mTOR/mTOR and SQSTM1/p62 levels, whereas knockdown of RACK1 prevented morphine-induced autophagy in vitro. Furthermore, we found that in the mouse hippocampus, knockdown of RACK1 also markedly suppressed morphine-induced autophagy (decreased LC3-II/LC3-I ratio and increased p-mTOR/mTOR ratio). Importantly, morphine-induced autophagy in a RACK1-dependent manner. Conversely, morphine-induced RACK1 upregulation in vitro is partially inhibited by autophagy feedback. CONCLUSIONS AND IMPLICATIONS: Our findings revealed a critical role for RACK1-dependent autophagy in morphine-promoted maintenance of CPP memory in mice and supported the notion that control of RACK1-dependent autophagic pathways may become an important target for novel therapeutics for morphine-associated memory.