Effect of mucilage from yam on activation of lymphocytic immune cells

The immunostimulating activities of mucilage fraction from yam were investigated. The proliferation of BSA-primed lymph node cells was enhanced between 4.1- to 10.9-fold compare to control, when cultured with 1 to 25 µg/mL of yam-mucilage fraction. It showed strong immunopotentiating activity than ginseng extract and as remarkable as Bifidobacterium adolescentis M101-4 known as a positive immunostimulator. Mitogenicity to lymph node cells was fully induced by concanavalin A and lipopolysaccharide. The proliferation of splenocytes and Peyer's patch cells was enhanced between 5.0- to 14.1-fold and 2.4- to 6.4-fold, respectively, when cultured with 1 to 25 µg/mL of yam-mucilage fraction. It enhanced the production of cytokines such as tumor necrosis factor-α and IL-6 in the culture of RAW 264.7 macrophage cells. In the culture of lipopolysaccharide-stimulated RAW 264.7 cells, production of cytokines was as similar as compared to controls. In unstimulated RAW 264.7 cells, both tumor necrosis factor-α and IL-6 production were enhanced between 15.6- to 60.1-fold and 2.3- to 9.1-fold, respectively. Mucilage fraction from yam is expected to be a safe immunopotentiator to maintain the host immunity and develop a physiologically functional food

Soluble N-Ethylmaleimide-Sensitive Factor Attachment Protein Receptor-Derived Peptides for Regulation of Mast Cell Degranulation

Vesicle-associated V-soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins and target membrane-associated T-SNAREs (syntaxin 4 and SNAP-23) assemble into a core trans-SNARE complex that mediates membrane fusion during mast cell degranulation. This complex plays pivotal roles at various stages of exocytosis from the initial priming step to fusion pore opening and expansion, finally resulting in the release of the vesicle contents. In this study, peptides with the sequences of various SNARE motifs were investigated for their potential inhibitory effects against SNARE complex formation and mast cell degranulation. The peptides with the sequences of the N-terminal regions of vesicle-associated membrane protein 2 (VAMP2) and VAMP8 were found to reduce mast cell degranulation by inhibiting SNARE complex formation. The fusion of protein transduction domains to the N-terminal of each peptide enabled the internalization of the fusion peptides into the cells equally as efficiently as cell permeabilization by streptolysin-O without any loss of their inhibitory activities. Distinct subsets of mast cell granules could be selectively regulated by the N-terminal-mimicking peptides derived from VAMP2 and VAMP8, and they effectively decreased the symptoms of atopic dermatitis in mouse models. These results suggest that the cell membrane fusion machinery may represent a therapeutic target for atopic dermatitis

Chlorogenic Acid Isomers Isolated from <i>Artemisia lavandulaefolia</i> Exhibit Anti-Rosacea Effects In Vitro

Rosacea is a chronic inflammatory disease affecting facial skin. It is associated with immune and vascular dysfunction mediated via increased expression and activity of cathelicidin and kallikrein 5 (KLK5), a serine protease of stratum corneum. Therefore, KLK5 inhibitors are considered as therapeutic agents for improving the underlying pathophysiology and clinical manifestation of rosacea. Here, we isolated the active constituents of Artemisia lavandulaefolia (A. lavandulaefolia) and investigated their inhibitory effect on KLK5 protease activity. Using bioassay-guided isolation, two bioactive compounds including chlorogenic acid isomers, 3,5-dicaffeoylquinic acid (isochlorogenic acid A) (1), and 4,5-dicaffeoylquinic acid (isochlorogenic acid C) (2) were isolated from A. lavandulaefolia. In this study, we evaluated the effects of isochlorogenic acids A and C on dysregulation of vascular and immune responses to rosacea, and elucidated their molecular mechanisms of action. The two chlorogenic acid isomers inhibit KLK5 protease activity, leading to reduced conversion of inactive cathelicidin into active LL-37. This inhibition of LL-37 production by isochlorogenic acids A and C reveals the efficacy of suppressing the expression of inflammatory mediators induced by LL-37 in immune cells such as macrophages and mast cells. In addition, both isomers of chlorogenic acid directly inhibited the proliferation and migration of vascular endothelial cells induced by LL-37

TNF−α Secreted from Macrophages Increases the Expression of Prometastatic Integrin αV in Gastric Cancer

The tumor microenvironment comprising blood vessels, fibroblasts, immune cells, and the extracellular matrix surrounding cancer cells, has recently been targeted for research in cancer therapy. We aimed to investigate the effect of macrophages on the invasive ability of gastric cancer cells, and studied their potential mechanism. In transcriptome analysis, integrin αV was identified as a gene increased in AGS cells cocultured with RAW264.7 cells. AGS cells cocultured with RAW264.7 cells displayed increased adhesion to the extracellular matrix and greater invasiveness compared with AGS cells cultured alone. This increased invasion of AGS cells cocultured with RAW264.7 cells was inhibited by integrin αV knockdown. In addition, the increase in integrin αV expression induced by tumor necrosis factor-α (TNF-α) or by coculture with RAW264.7 cells was inhibited by TNF receptor 1 (TNFR1) knockdown. The increase in integrin αV expression induced by TNF-α was inhibited by both Mitogen-activated protein kinase (MEK) inhibitor and VGLL1 S84 peptide treatment. Finally, transcription of integrin αV was shown to be regulated through the binding of VGLL1 and TEAD4 to the promoter of integrin αV. In conclusion, our study demonstrated that TNFR1–ERK–VGLL1 signaling activated by TNF-α secreted from RAW264.7 cells increased integrin αV expression, thereby increasing the adhesion and invasive ability of gastric cancer cells

Munc18-1 induces conformational changes of syntaxin-1 in multiple intermediates for SNARE assembly

Abstract In neuronal exocytosis, SNARE assembly into a stable four-helix bundle drives membrane fusion. Previous studies have revealed that the SM protein Munc18-1 plays a critical role for precise SNARE assembly with the help of Munc13-1, but the underlying mechanism remains unclear. Here, we used single-molecule FRET assays with a nanodisc membrane reconstitution system to investigate the conformational dynamics of SNARE/Munc18-1 complexes in multiple intermediate steps towards the SNARE complex. We found that single Munc18-1 proteins induce the closed conformation of syntaxin-1 not only in the free syntaxin-1 but also in the t-SNARE (syntaxin-1/SNAP-25) complex. These results implicate that Munc18-1 may act as a gatekeeper for both binary and ternary SNARE complex formation by locking the syntaxin-1 in a cleft of Munc18-1. Furthermore, the kinetic analysis of the opening/closing transition reveals that the closed syntaxin-1 in the syntaxin-1/SNAP-25/Munc18-1 complex is less stable than that in the closed syntaxin-1/Munc18-1 complex, which is manifested by the infrequent closing transition, indicating that the conformational equilibrium of the ternary complex is biased toward the open conformation of syntaxin-1 compared with the binary complex

Synthesis and physicochemical characterization of acyl myricetins as potential anti-neuroexocytotic agents

Abstract Acyl myricetins (monopropionyl-, dipropionyl-, and monooctanoyl-myricetin, termed as MP1, MP2, and MO1, respectively) were synthesized through enzymatic or non-enzymatic esterification reaction of myricetin aglycone. Structure study indicated the hydroxyl group at C4′ in B-ring was highly susceptible to acylation. Over its parental myricetin, acylated compounds showed enhanced lipophilicity (from 7.4- to 26.3-fold) and oxidative stability (from 1.9- to 3.1-fold) on the basis of logP and decay rate, respectively. MO1, presenting the physicochemical superiority compared to the others, provided lowest EC50 value of 2.51 μM on inhibition of neutrotransmitter release and CC50 value of 59.0 μM, leading to widest therapeutic window. All myricetin esters did not show any irritation toxicity when assessed with a chicken embryo assay. This study describes information on acylation of myricetin that has not yet been explored, and suggests that MO1 has membrane fusion-arresting and anti-neuroexocytotic potential for industrial application due to its enhanced biological properties