Anti-asthma and antitussive effects of a fermented extract of a mixture of Ramulus mori, Anthriscus sylvestris, and Salvia plebeian.

Respiratory immunity is getting more important recently due to outbreak of respiratory diseases and increasing the concentration of fine dust. The aim of this study was to investigate respiratory protection effect of a fermented extract of medicinal plants (FEMP) containing Ramulus mori, Salvia plebeia, and Anthriscus sylvestris. The expression levels of IL-8 and IL-17 in LPS/poly-L-arginine (PLA) and FEMP-cotreated A549 cells were lower than those in LPS/PLA only-treated cells. The levels of IgE, IL-17, and IL-4 in the bronchoalveolar lavage fluid (BALF) and serum of FEMP-treated mice with ovalbumin/LPS-induced asthma were lower than the control levels. The lung inflammation score and the number of inflammatory cells in the BALF decreased by FEMP treatment. In the citric acid-induced coughing guinea pig, the FEMP treatment decreased the number of coughs. Therefore, FEMP shows anti-asthmatic and antitussive activities without hepatotoxicity and can be used as a compound aiming to improve respiratory health. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10068-021-00955-3.

Oxyresveratrol Ameliorates Dextran Sulfate Sodium-Induced Colitis in Rats by Suppressing Inflammation.

Colitis causes destruction of the intestinal mucus layer and increases intestinal inflammation. The use of antioxidants and anti-inflammatory agents derived from natural sources has been recently highlighted as a new approach for the treatment of colitis. Oxyresveratrol (OXY) is an antioxidant known to have various beneficial effects on human health, such as anti-inflammatory, antibacterial activity, and antiviral activity. The aim of this study was to investigate the therapeutic effect of OXY in rats with dextran sulfate sodium (DSS)-induced acute colitis. OXY ameliorated DSS-induced colitis and repaired damaged intestinal mucosa. OXY downregulated the expression of pro-inflammatory cytokine genes (TNF-α, IL-6, and IL-1ß) and chemokine gene MCP-1, while promoting the production of anti-inflammatory cytokine IL-10. OXY treatment also suppressed inflammation via inhibiting cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) expression in the colon, as well as the activity of myeloperoxidase (MPO). OXY exhibited anti-apoptotic effects, shifting the Bax/Bcl-2 balance. In conclusion, OXY might improve DSS-induced colitis by restoring the intestinal mucus layer and reducing inflammation within the intestine.

Oxyresveratrol improves tight junction integrity through the PKC and MAPK signaling pathways in Caco-2 cells.

Strengthening intestinal tight junctions (TJ) provides an effective barrier from the external environment and is important for recovery from inflammatory bowel disease. Oxyresveratrol (OXY), an isomer of hydroxylated resveratrol, is isolated from many plants. The aim of this study was to investigate the effect of OXY on intestinal TJ and to elucidate the mechanism underlying the OXY-mediated increase in TJ integrity in human intestinal Caco-2 cells. OXY-treated Caco-2 cell monolayers showed decreased monolayer permeability as evaluated by paracellular transport assay. The results showed that OXY significantly increased the levels of TJ-related genes and proteins (Claudin-1, Occludin and ZO-1) compared with those of the negative control. OXY activated protein kinase C (PKC) and increased expression levels of mitogen-activated protein kinase (MAPK) genes. OXY also increased gene and protein levels of the transcription factor Cdx-2. Expression levels of TJ, PKC and Cdx-2 proteins and transepithelial electrical resistance (TEER) value decreased in OXY-treated Caco-2 cells following treatment with a pan-PKC inhibitor compared with those of the untreated control. In conclusion, OXY strengthens the integrity of the intestinal TJ barrier via activation of the PKC and MAPK pathways.

Conditioned medium from LS 174T goblet cells treated with oxyresveratrol strengthens tight junctions in Caco-2 cells.

BACKGROUND: Strengthening of intestinal tight junctions provides an effective barrier from the external environment. Goblet cell-derived trefoil factor 3 (TFF3) increases transepithelial resistance by upregulating the expression of tight junction proteins. Oxyresveratrol (OXY) is a hydroxyl-substituted stilbene found in the roots, leaves, stems, and fruit of many plants and known to have various biological activities. In this study, we investigated the strengthening effect of OXY on intestinal tight junctions through stimulation of TFF production in goblet cells. METHODS: We prepared conditioned medium from LS 174T goblet cells treated with OXY (GCO-CM) and investigated the effect of GCO-CM on strengthening tight junctions of Caco-2 cells. The mRNA and protein expression levels of major tight junction components (claudin-1, occludin, and ZO-1) were measured by quantitative real-time PCR and western blotting, respectively. Transepithelial electric resistance (TEER) was measured using an ohm/V meter. Monolayer permeability was evaluated by paracellular transport of fluorescein isothiocyanate-dextran. RESULTS: OXY showed a strong antioxidant activity. It significantly increased the expression level of TFF3 in LS 174T goblet cells. GCO-CM prepared by treatment with 2.5, 5, and 10µg/ml OXY did not show cytotoxicity in Caco-2 cells. GCO-CM increased the mRNA and protein expression levels of claudin-1, occludin, and ZO-1. It also significantly increased tight junction integrity and reduced permeability in a dose-dependent manner. CONCLUSION: OXY stimulates the expression of TFF3 in goblet cells, which might increase the integrity of the intestinal tight junction barrier.

Brief Communication: SIR-2.1-dependent lifespan extension of Caenorhabditis elegans by oxyresveratrol and resveratrol.

Resveratrol (RES) has been studied for its effects on the lifespan extension of Caenorhabditis elegans, but controversy still remains on its mechanism related with SIR-2. In this study, longevity assay was performed to confirm SIR-2-dependent lifespan extension of C. elgeans with RES and oxyresveratrol (OXY), an isomer of hydroxylated RES using loss-of-function mutants of C. elegans including sir-2.1 mutant. The results showed that OXY and RES significantly (P < 0.05) extended the lifespan of C. elegans compared with the control. OXY and RES also significantly (P < 0.05) increased the mRNA expression levels of sir-2.1 and aak-2 in a dose-dependent manner and increased the protein expression levels of SIR-2.1. OXY and RES treatment extended the lifespan in daf-16 loss-of-function mutants, which suggested that lifespan extension was not occurring via the activation of DAF-16. However, OXY and RES failed to extend the lifespan in loss-of-function mutants of sir-2.1 and aak-2 Therefore, OXY and RES extend the lifespan of C. elegans by overexpression of SIR-2.1, which is related to lifespan extension through calorie restriction and the AMP-activated protein kinase (AMPK) pathway, although this process is independent of the FOXO/DAF-16 pathway.

An ethanol extract of Ramulus mori improves blood circulation by inhibiting platelet aggregation.

Inappropriate platelet aggregation can cause blood coagulation and thrombosis. In this study, the effect of an ethanol extract of Ramulus mori (ERM) on blood circulation was investigated. The antithrombotic activity of ERM on rat carotid arterial thrombosis was evaluated in vivo, and the effect of ERM on platelet aggregation and blood coagulation time was evaluated ex vivo. To evaluate the safety of ERM, its cytotoxicity to platelets and its effect on tail bleeding time were assessed; ERM was not toxic to rat platelets and did not prolong bleeding time. Moreover, administering ERM to rats had a significant preventive effect on carotid arterial thrombosis in vivo, and significantly inhibited adenosine diphosphate- and collagen-induced platelet aggregation ex vivo, whereas it did not prolong coagulation periods, such as prothrombin time and activated partial thromboplastin time. The results suggest that ERM is effective in improving blood circulation via antiplatelet activity rather than anticoagulation activity.

Anti-inflammatory action of ethanolic extract of Ramulus mori on the BLT2-linked cascade.

Mulberry tree twigs (Ramulus mori) contain large amounts of oxyresveratrols and have traditionally been used as herbal medicines because of their anti-inflammatory properties. However, the signaling mechanism by which R. mori exerts its anti-inflammatory action remains to be elucidated. In this study, we observed that R. mori ethanol extracts (RME) exerted an inhibitory effect on the lipopolysaccharide (LPS)-induced production of the pro-inflammatory cytokine interleukin-6 (IL-6) in Raw264.7 macrophage cells. Additionally, RME inhibited IL-6 production by blocking the leukotriene B4 receptor- 2 (BLT2)-dependent-NADPH oxidase 1 (NOX1)-reactive oxygen species (ROS) cascade, leading to anti-inflammatory activity. Finally, RME suppressed the production of the BLT2 ligands LTB4 and 12(S)-HETE by inhibiting the p38 kinase- cytosolic phospholipase A2-5-/12-lipoxygenase cascade in LPS-stimulated Raw264.7 cells. Overall, our results suggest that RME inhibits the 'BLT2 ligand-BLT2'-linked autocrine inflammatory axis, and that this BLT2-linked cascade is one of the targets of the anti-inflammatory action of R. mori. [BMB Reports 2016; 49(4): 232-237].

Antihyperlipidemic effects of rhapontin and rhapontigenin from rheum undulatum in rats fed a high-cholesterol diet.

Rhapontin was purified from a methanol extract from the roots of Rheum undulatum, and rhapontigenin was produced by an enzymatic transformation of rhapontin. Rats were fed a high-cholesterol diet to induce hyperlipidemia, followed by oral treatment with rhapontin or rhapontigenin (1-5 mg/kg/day). Rhapontin and rhapontigenin treatment resulted in a significant (p<0.05) dose-dependent decrease in the serum lipid level, while the high-density lipoprotein cholesterol level increased slightly compared with the experimental control. Furthermore, rhapontin and rhapontigenin treatment improved the pathological characteristics of the degenerating fatty liver in high-cholesterol diet-induced hyperlipidemic rats dose-dependently. Aspartate aminotransferase and alanine aminotransferase levels in rhapontin- and rhapontigenin-treated hyperlipidemic rats were not significantly different from those in the control. These results indicate that rhapontin and rhapontigenin can be used as potent antihyperlipidemic agents.

Antihyperlipidemic effects of stilbenoids isolated from Morus alba in rats fed a high-cholesterol diet.

Mulberroside A (MUL) was purified from an ethanol extract of Morus alba root, and oxyresveratrol (OXY) was produced by enzymatic conversion of MUL. Normal rats, Triton WR-1339-induced hyperlipidemic rats, and high-cholesterol diet (HCD)-induced hyperlipidemic rats were orally treated with MUL or OXY (1-5mg/kg/day). MUL and OXY were administered 1h prior to concomitant treatment with Triton WR-1339 for a further 24h, whereas the drugs were administered concurrently with HCD for 4weeks. Oral MUL and OXY pre-treatment vs. water pre-treatment of Triton WR-1339-induced hyperlipidemic rats significantly (p<0.05) reduced the levels of serum lipids in a dose-dependent manner, while high-density lipoprotein cholesterol (HDL-C, or "good" cholesterol) levels were increased. Oral MUL and OXY treatment of HCD-fed rats also showed a significant (p<0.05) dose-dependent decrease in serum lipids, coronary artery risk index (CRI), and atherogenic index (AI), but not HDL-C. Furthermore, MUL and OXY treatment of HCD-induced hyperlipidemic rats demonstrated a significant dose-dependent improvement in the histological features of hepatic fatty degeneration. Aspartate aminotransferase and alanine aminotransferase values in OXY-treated normal rats were not significantly different from those in water-treated control rats. These results indicate that MUL and OXY might be developed as novel antihyperlipidemic agents.

Evaluation of the inhibition of mushroom tyrosinase and cellular tyrosinase activities of oxyresveratrol: comparison with mulberroside A.

The inhibitory effects of oxyresveratrol, the aglycone of mulberroside A, on mushroom and cellular tyrosinase activities and melanin synthesis were evaluated. Mulberroside A and oxyresveratrol showed inhibitory activity against mushroom tyrosinase, with oxyresveratrol demonstrating a greater inhibitory effect than that of mulberroside A. Oxyresveratrol and mulberroside A strongly inhibited melanin production in Streptomyces bikiniensis and exhibited dose-dependent inhibition of tyrosinase activity and inhibition of melanin synthesis in B16F10 melanoma cells. However, the compounds exhibited nearly similar inhibitory effects on the activity of cellular tyrosinase and melanin synthesis in murine melanocytes. The inhibition of melanin synthesis by mulberroside A and oxyresveratrol was involved in suppressing the expression level of melanogenic enzymes, tyrosinase, tyrosinase-related protein-1 (TRP-1), and tyrosinase-related protein-2 (TRP-2). These results indicate that the inhibition rate of mushroom tyrosinase might not provide an accurate estimate of the inhibition rate of melanin synthesis in melanocytes.

Inhibitory effect of mulberroside A and its derivatives on melanogenesis induced by ultraviolet B irradiation.

Mulberroside A was isolated from the ethanol extract of Morus alba roots. The enzymatic hydrolysis of mulberroside A with Pectinex produced oxyresveratrol and oxyresveratrol-3-O-glucoside. We tested oxyresveratrol, oxyresveratrol-3-O-glucoside, and mulberroside A to determine whether they could inhibit ultraviolet B (UVB) irradiation-induced melanogenesis in brown guinea pig skin. Topical application of mulberroside A, oxyresveratrol, and oxyresveratrol-3-O-glucoside reduced the pigmentation in guinea pig skin. These compounds suppressed the expression of melanogenic enzymes tyrosinase, tyrosinase-related protein-1, and microphthalmia transcription factor. The anti-melanogenesis effect was highest with oxyresveratrol, intermediate with oxyresveratrol-3-O-glucoside, and lowest with mulberroside A. Mulberroside A is a glycosylated stilbene of oxyresveratrol; thus, the deglycosylation of mulberroside A resulted in enhanced inhibition of melanogenesis. Histological analysis with Fontana-Masson staining confirmed that these compounds significantly reduced the melanin content in the epidermis of UVB-irradiated guinea pig skin compared to the vehicle control. Thus, these compounds effectively reduced pigmentation and may be suitable cosmetic agents for skin whitening.

Biotransformation of mulberroside A from Morus alba results in enhancement of tyrosinase inhibition.

Mulberroside A, a glycosylated stilbene, was isolated and identified from the ethanol extract of the roots of Morus alba. Oxyresveratrol, the aglycone of mulberroside A, was produced by enzymatic hydrolysis of mulberroside A using the commercial enzyme Pectinex. Mulberroside A and oxyresveratrol showed inhibitory activity against mushroom tyrosinase with an IC(50) of 53.6 and 0.49 microM, respectively. The tyrosinase inhibitory activity of oxyresveratrol was thus approximately 110-fold higher than that of mulberroside A. Inhibition kinetics showed mulberroside A to be a competitive inhibitor of mushroom tyrosinase with L-tyrosine and L-DOPA as substrate. Oxyresveratrol showed mixed inhibition and noncompetitive inhibition against L-tyrosine and L-DOPA, respectively, as substrate. The results indicate that the tyrosinase inhibitory activity of mulberroside A was greatly enhanced by the bioconversion process.

Evaluation of the antibacterial activity of rhapontigenin produced from rhapontin by biotransformation against Propionibacterium acnes.

Biotransformation is often used to improve chemical activity. We evaluated the antimicrobial activity of rhapontigenin, converted from rhapontin after treatment with Pectinex. Rhapontigenin showed 4-16 times higher antimicrobial activity than rhapontin. Activity was higher against Gram positive strains than Gram negative strains. Minimum inhibitory concentrations (MICs) of rhapontigenin, retinol, and five antibiotics were determined by microbroth dilution method for antibiotic-sensitive and -resistant Propionibacterium acnes. We also investigated the in vitro antibacterial activity of rhapontigenin in combination with antibiotic against antibiotic-resistant P. acnes. The antibiotic combination effect against resistant P. acnes was studied by checkerboard method. The combination formulations (rhapontigenin and clindamycin, retinol and clindamycin) showed synergic effects on the inhibition of the growth of clindamycin-resistant P. acnes. It is predictable that the combination of antibiotics with rhapontigenin is helpful to treat acne caused by antibiotic resistant P. acnes. The antibacterial activity of rhapontigenin was enhanced by biotransformation.