Rubi Fructus Water Extract Alleviates LPS-Stimulated Macrophage Activation via an ER Stress-Induced Calcium/CHOP Signaling Pathway

Abstract

Despite the availability of antibiotics and vaccines, many intractable infectious diseases still threaten human health across the globe. Uncontrolled infections can lead to systemic inflammatory response syndrome and the excessive production of inflammatory cytokines, known as a cytokine storm. As cytokines also play necessary and positive roles in fighting infections, it is important to identify nontoxic and anti-inflammatory natural products that can modulate cytokine production caused by infections. Rubi Fructus, the unripe fruits of Rubus coreanus Miquel, are known to possess antioxidative properties. In this study, the effect of the water extract of Rubi Fructus (RF) on the lipopolysaccharide (LPS)-induced inflammatory response in RAW 264.7 macrophages was investigated using biochemical and cell biology techniques. Our data indicated that RF inhibits p38 phosphorylation, intracellular calcium release, and the production of nitric oxide (NO), interleukin (IL)-6, monocyte chemotactic activating factor (MCP)-1, tumor necrosis factor (TNF)-α, leukemia inhibitory factor (LIF), lipopolysaccharide-induced CXC chemokine (LIX), granulocyte-colony stimulating factor (G-CSF), granulocyte macrophage colony-stimulating factor (GM-CSF), vascular endothelial growth factor (VEGF), macrophage colony-stimulating factor (M-CSF), macrophage inflammatory protein (MIP)-1α, MIP-1β, MIP-2, and regulated on activation, normal T cell expressed and secreted (RANTES) in LPS-treated macrophages. In addition, we observed decreasing mRNA expression of Chop, Camk2a, Stat1, Stat3, Jak2, Fas, c-Jun, c-Fos, Nos2, and Ptgs2 without cytotoxic effects. We concluded that RF demonstrated immunoregulatory activity on LPS-stimulated macrophages via an endoplasmic reticulum (ER) stress-induced calcium/CCAAT-enhancer-binding protein homologous protein (CHOP) pathway and the Janus kinase (JAK)/signal transducers and activators of transcription (STAT) pathway.

A Combination Extract of Gardeniae Fructus and Perillae Folium Exerts Anti-Inflammatory Effects on LPS-Activated RAW 264.7 Mouse Macrophages via an ER Stress-Induced CHOP Pathway

“…Gardeniae Fructus and Perillae Folium were extracted together with boiling water for 2 h, filtered, and then lyophilized (yield: 20.75%). The powdered extract was dissolved in saline and then filtered through a 0.22 µM syringe filter [11,12].…”

“…The total flavonoid content of GP was determined using the diethylene glycol colorimetric method [11,12]. Total flavonoid content was expressed as milligrams of rutin equivalents (mg RE/g extract).…”

“…The RAW 264.7 cell lines (second passage) were purchased from Korea Cell Line Bank (Seoul, Korea). Cells were cultured in DMEM, 10% heat-inactivated FBS, 1% of penicillinstreptomycin [11,12]. The cells were incubated in atmosphere of 5% CO 2 and 95% humidity at 37 • C [11,12].…”

“…Cells were cultured in DMEM, 10% heat-inactivated FBS, 1% of penicillinstreptomycin [11,12]. The cells were incubated in atmosphere of 5% CO 2 and 95% humidity at 37 • C [11,12]. Cell viability was accessed by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5diphenyltetrazolium bromide) assay [11,12].…”

“…The cells were incubated in atmosphere of 5% CO 2 and 95% humidity at 37 • C [11,12]. Cell viability was accessed by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5diphenyltetrazolium bromide) assay [11,12]. Briefly, cell viabilities in RAW 264.7 cells which were incubated with GP at concentrations of 25, 50, 100, and 200 µg/mL for 24 h were 100.49 ± 7.32%, 100.76 ± 4.39%, 116.6 ± 12.66%, and 239.54 ± 11.09% of the control group treated with media only, respectively.…”