Iridoid Glycosides and Coumarin Glycoside Derivatives from the Roots of Nymphoides peltata and Their In Vitro Wound Healing Properties.

Nymphoides peltata has been used as a medicinal herb in traditional medicines to treat strangury, polyuria, and swelling. The phytochemical investigation of the MeOH extract of N. peltata roots led to the isolation of three iridoid glycosides and three coumarin glycoside derivatives, which were characterized as menthiafolin (1), threoninosecologanin (2), callicoside C (3), and scopolin (4), as well as two undescribed peltatamarins A (5) and B (6). The chemical structures of the undescribed compounds were determined by analyzing their 1 dimensional (D) and 2D nuclear magnetic resonance (NMR) spectra and using high-resolution (HR)-electrospray ionization mass spectroscopy (ESI-MS), along with the chemical reaction of acid hydrolysis. The wound healing activities of the isolated compounds 1-6 were evaluated using a HaCaT cell scratch test. Among the isolates, scopolin (4) and peltatamarin A (5) promoted HaCaT cell migration over scratch wounds, and compound 5 was the most effective. Furthermore, compound 5 significantly promoted cell migration without adversely affecting cell proliferation, even when treated at a high dose (100 µM). Our results demonstrate that peltatamarin A (5), isolated from N. peltata roots, has the potential for wound healing effects.

3‴-O-Foliamenthoyl-Rutin, a New Flavonoid Glycoside from the Roots of Nymphoides peltata.

Nymphoides peltata (Menyanthaceae) has been used as a medicinal herb in traditional medicines to treat conditions such as strangury, polyuria, swelling, and as a diuretic and antipyretic. In our ongoing research to discover novel structural and/or biological natural products in natural resources, five flavonoids, quercetin (1), quercitrin (2), isoquercetin (3), quercetin-3-O-vicianoside (4), and rutin (5), as well as a new flavonoid glycoside, 3‴-O-foliamenthoyl-rutin (6), were isolated from the MeOH extract of N. peltata roots. The chemical structure of the new compound (6) was determined by analyzing 1D and 2D NMR spectra and high-resolution (HR) electrospray ionization mass spectroscopy (ESIMS), along with a chemical reaction. The wound-healing activities of the isolated compounds (1-6) were evaluated using a HaCaT cell scratch test. Among the isolates, isoquercetin (3), quercetin-3-O-vicianoside (4), and 3‴-O-foliamenthoyl-rutin (6) promoted HaCaT cell migration over scratch wounds, with compound 4 being the most effective. Our findings provide experimental data supporting the potential of quercetin-3-O-vicianoside (4) as a wound-healing agent.

Anti-Wrinkling Effect of 3,4,5-tri-O-caffeoylquinic Acid from the Roots of Nymphoides peltata through MAPK/AP-1, NF-κB, and Nrf2 Signaling in UVB-Irradiated HaCaT Cells.

Nymphoides peltata has been widely used pharmacologically in traditional Chinese medicine to treat heat strangury and polyuria. The aim of this study was to isolate the bioactive components from N. peltata and evaluate their potential use as antioxidant and anti-wrinkle agents. Phytochemical investigation of the methanolic extract of N. peltata roots led to the isolation of 15 compounds (1-15), which were structurally determined as α-spinasterol (1), 3-O-ß-D-glucopyranosyl-oleanolic acid 28-O-ß-D-glucuronopyranoside (2), 4-hydroxybenzoic acid (3), protocatechuic acid (4), vanillic acid (5), p-coumaric acid (6), caffeic acid (7), ferulic acid (8), neochlorogenic acid (neo-CQA) (9), chlorogenic acid (CQA) (10), cryptochlorogenic acid (crypto-CQA) (11), isochlorogenic acid B (3,4-DCQA) (12), isochlorogenic acid A (3,5-DCQA) (13), isochlorogenic acid C (4,5-DCQA) (14), and 3,4,5-tri-O-caffeoylquinic acid (TCQA) (15). Of these 15 compounds, compound 2 was a new oleanane saponin, the chemical structure of which was characterized by 1D and 2D nuclear magnetic resonance (NMR) spectroscopic data and high-resolution electrospray ionization mass spectrometry (HRESIMS), as well as chemical reaction. Biological evaluation of the isolated compounds revealed that 3,4,5-tri-O-caffeoylquinic acid (TCQA) significantly improved Nrf2 levels in an Nrf2-ARE reporter HaCaT cell screening assay. TCQA was found to potently inhibit the Nrf2/HO-1 pathway and to possess strong anti-wrinkle activity by modulating the MAPK/NF-κB/AP-1 signaling pathway and thus inhibiting MMP-1 synthesis in HaCaT cells exposed to UVB. Our results suggest that TCQA isolated from N. peltata might be useful for developing effective antioxidant and anti-wrinkle agents.

A High-Performance Liquid Chromatography with Photodiode Array Detection Method for Simultaneous Determination of Three Compounds Isolated from Wikstroemia ganpi: Assessment of the Effects on Cytochrome P450-Mediated Metabolism In Vitro and In Vivo.

In natural products, the content and quality of the marker components differ depending on the part, production area, collection period, and extraction method; therefore, a standardized analysis method is required to obtain consistent results. This study developed a simultaneous analysis method for three marker components (7-methoxylutolin-5-O-glucoseide, pilloin 5-O-ß-d-glucopyranoside, rutarensin) isolated and purified from Wikstroemia ganpi (W. ganpi). Simultaneous analysis was performed using high-performance liquid chromatography with photodiode array detection (HPLC-PDA) method that was validated according to the International Council for Harmonisation (ICH) guidelines. The developed analytical method exhibited linearity (r2 > 0.999), detection limits (0.72-3.34 µg/mL), and quantification limits (2.19-10.22 µg/mL). The relative standard deviation (RSD) value of intra- and inter-day precisions was less than 1.68%, and analyte recoveries (93.42-117.55%; RSD < 1.86%) were validated according to the analytical procedures, and all parameters were within the allowable range. Quantitative analysis of the three marker components from W. ganpi MeOH extract (WGM) showed 7-methoxylutolin-5-O-glucoseide with the highest content (51.81 mg/g). The inhibitory effects of WGM on cytochrome P450 (CYP) substrate drugs were further investigated. The in vitro study revealed that WGM inhibited the CYP3A-mediated metabolism of buspirone and that 7-methoxylutolin-5-O-glucoseide and pilloin 5-O-ß-d-glucopyranoside inhibited the metabolism of buspirone with IC50 values of 2.73 and 18.7 µM, respectively. However, a single oral dose of WGM did not have significant effects on the pharmacokinetics of buspirone in rats, suggesting that WGM cannot function as an inhibitor of CYP3A-mediated metabolism in vivo.

Lobelia chinensis Extract and Its Active Compound, Diosmetin, Improve Atopic Dermatitis by Reinforcing Skin Barrier Function through SPINK5/LEKTI Regulation.

The skin acts as a mechanical barrier that protects the body from the exterior environment, and skin barrier function is attributed to the stratum corneum (SC), which is composed of keratinocytes and skin lipids. Skin barrier homeostasis is maintained by a delicate balance between the differentiation and exfoliation of keratinocytes, and keratinocyte desquamation is regulated by members of the serine protease kalikrein (KLK) family and their endogenous inhibitor SPINK5/LEKTI (serine protease inhibitor Kazal type 5/lympho-epithelial Kazal-type-related inhibitor). Furthermore, SPINK5/LEKTI deficiency is involved in impaired skin barrier function caused by KLK over-activation. We sought to determine whether increased SPINK5/LEKTI expression ameliorates atopic dermatitis (AD) by strengthening skin barrier function using the ethanol extract of Lobelia chinensis (LCE) and its active compound, diosmetin, by treating human keratinocytes with UVB and using a DNCB-induced murine model of atopic dermatitis. LCE or diosmetin dose-dependently increased the transcriptional activation of SPINK5 promoter and prevented DNCB-induced skin barrier damage by modulating events downstream of SPINK5, that is, KLK, PAR2 (protease activated receptor 2), and TSLP (thymic stromal lymphopoietin). LCE or diosmetin normalized immune response in DNCB treated SKH-1 hairless mice as determined by reductions in serum immunoglobulin E and interleukin-4 levels and numbers of lesion-infiltrating mast cells. Our results suggest that LCE and diosmetin are good candidates for the treatment of skin barrier-disrupting diseases such as Netherton syndrome or AD, and that they do so by regulating SPINK5/LEKTI.

7-O-Methylluteolin Suppresses the 2,4-Dinitrochlorobenzene-Induced Nrf2/HO-1 Pathway and Atopic Dermatitis-like Lesions.

7-O-methylluteolin (7-ML) is a flavonoid isolated from the aerial parts of Wikstroemia ganpi (W. ganpi). We describe the anti-atopic dermatitis (AD) effects of 7-ML in tert-butyl hydroperoxide (tBHP)-induced HepG2 cells and 2,4-dinitrochlorobenzene (DNCB)-induced SKH-1 hairless mice. Results demonstrated that 7-ML dose-dependently inhibited the activation of Nrf2 (nuclear factor-erythroid 2-related factor 2) in tBHP-induced HepG2 cells. 7-ML applied topically to our DNCB-induced mouse model upregulated the antioxidant protein expression (phosphorylated Nrf2 (pNrf2), Nrf2, and heme oxygenase-1 (HO-1)) in skin tissues, improved epidermal thickness, and reduced mast cell infiltration into the skin. In addition, 7-ML reduced the serum levels of immunoglobulin E (IgE) and interleukin-4 (IL-4) and improved skin barrier functions. These results suggest that 7-ML should be considered a novel antioxidant and anti-AD agent.

Inhibitory Effects of Luteolin 7-Methyl Ether Isolated from Wikstroemia ganpi on Tnf-A/Ifn-Γ Mixture-Induced Inflammation in Human Keratinocyte.

Plants of the genus Wikstroemia are traditionally used in China to treat various inflammatory diseases. The purpose of this study was to isolate the components of Wikstroemia ganpi (Siebold & Zucc.) Maxim., to evaluate their anti-atopic activities and to identify candidates with anti-atopic therapeutics. A total of 24 compounds were isolated by bioassay-guided separation, including one novel compound, which was tilianin 5-methyl ether. The anti-atopic activities of the isolated compounds were determined using TNF-α-treated RBL-2H3 cells and HaCaT cells. The mRNA expressions of IL-4, IL-6, GM-CSF, G-CSF and TRPV1 were reduced by luteolin 7-methyl ether. The study shows that the luteolin 7-methyl ether isolated from W. ganpi is a potential therapeutic agent for the treatment of atopic dermatitis.

Haplopine Ameliorates 2,4-Dinitrochlorobenzene-Induced Atopic Dermatitis-Like Skin Lesions in Mice and TNF-α/IFN-γ-Induced Inflammation in Human Keratinocyte.

This study aimed to investigate the anti-inflammatory, antioxidant, and anti-atopic dermatitis (AD) effects of haplopine, which is one of the active components in D. dasycarpus. Haplopine (12.5 and 25 µM) inhibited the mRNA expressions of inflammatory cytokines IL-6, TSLP, GM-CSF, and G-CSF and the protein expressions of IL-6 and GM-CSF in TNF-α/INF-γ-stimulated HaCaT cells. In H2O2-induced Jukat T cells, haplopine (25 and 50 µM) suppressed the productions of proinflammatory cytokines (IL-4, IL-13, and COX-2) and increased the mRNA and protein expressions of oxidative stress defense enzymes (SOD, CAT, and HO-1) in a concentration-dependent manner. In vivo, haplopine significantly attenuated the development of AD symptoms in 2,4-dinitrochlorobenzene (DNCB)-stimulated Balb/c mice, as evidenced by reduced clinical dermatitis scores, skin thickness measurements, mast cell infiltration, and serum IgE concentrations. These findings demonstrate that haplopine should be considered a novel anti-atopic agent with the potential to treat AD.

Wikstroemiaganpi Extract Improved Atopic Dermatitis-Like Skin Lesions via Suppression of Interleukin-4 in 2,4-Dinitrochlorobenzene-Induced SKH-1 Hairless Mice.

Plants of the genus Wikstroemia are used in Chinese traditional medicine to treat inflammatory diseases, such as arthritis, bronchitis, and pneumonia. The present study was designed to determine whether Wikstroemia ganpi (Siebold and Zucc.) Maxim. offers a potential means of treating 2,4-dinitrochlorobenzene (DNCB)-induced atopic dermatitis (AD) in mice. Symptoms such as redness, edema, and keratinization in AD mice induced by DNCB were alleviated by the co-application of an ethanolic extract of W. ganpi for 2 weeks. The severity of skin barrier function damage was evaluated by measuring TEWL (transepidermal water loss). TEWLs of DNCB sensitized mouse dorsal skin were reduced by the application of a W. ganpi ethanolic extract, and skin hydration was increased. In addition, the infiltration of inflammatory cells into the dermis was significantly reduced, as were blood levels of IgE and IL-4, which play an important role in the expression of AD. The results of this experiment suggest that W. ganpi is a potential therapeutic agent for AD.

Quercitrin, the Main Compound in Wikstroemia indica, Mitigates Skin Lesions in a Mouse Model of 2,4-Dinitrochlorobenzene-Induced Contact Hypersensitivity.

Hapten-induced contact hypersensitivity (CHS) is widely utilized to induce immune activation in animal models of allergic contact dermatitis. Our previous findings suggested that the 95% EtOH extract of Wikstroemia indica (L.) C. A. Mey. has antiallergic and anti-inflammatory effects in DNCB-treated CHS SKH-1 hairless mice. The aim of this study was to evaluate the protective effects of compounds isolated from the EtOAc fraction of W. indica in RBL-2H3 cells and 2,4-dinitrochlorobenzene- (DNCB-) induced CHS mice. Of eight compounds in W. indica, that is, umbelliferone, daphnoretin, wikstrocoumarin, (+)-syringaresinol, tricin, (+)-lariciresinol, erythro-guaiacylglycerol-ß-coniferyl ether, and quercitrin, quercitrin exhibited the most antiallergic activity against antigen-induced ß-hexosaminidase release and IL-4 mRNA expression, which are markers of degranulation in RBL-2H3 cells. After a 7-sensitizing period, 14 days of DNCB treatment with or without topical pimecrolimus (1%) or quercitrin (0.5%) treatment, quercitrin was found to suppress DNCB-induced increases in serum IL-4 and IgE concentrations and transepidermal water loss. These results indicate that quercitrin has therapeutic potential for treatment of allergies and allergy-related contact dermatitis.

Suppression of DNCB-Induced Atopic Skin Lesions in Mice by Wikstroemia indica Extract.

Wikstroemia indica (L.) C.A. Mey. is used in traditional Chinese medicine to treat inflammatory diseases such as arthritis and bronchitis. In this study, we aimed to investigate the effects of an ethanolic extract of W. indica on cutaneous inflammation in mice with 2,4-dinitrochlorobenzene (DNCB)-induced atopic dermatitis (AD). Dermal administration of W. indica ethanolic extract to DNCB-sensitized hairless mice with dermatitis, for two weeks, reduced erythema, scaling, and edema. Skin hydration was improved and transepidermal water loss was reduced at a W. indica concentration of 1%. Furthermore, W. indica also significantly reduced serum IgE and IL-4 concentrations in our mouse model. These results suggest that W. indica has potential as a topical treatment for AD and as an adjunctive agent to control AD.

Stellera chamaejasme and Its Main Compound Luteolin 7-O-Glucoside Alleviates Skin Lesions in Oxazolone- and 2,4-Dinitrochlorobenzene-Stimulated Murine Models of Atopic Dermatitis.

Stellera chamaejasme, also known as "Langdu", has been traditionally used for the management of skin-related diseases such as psoriasis and skin ulcers. The aim of this study was to determine whether S. chamaejasme and its major component, luteolin 7-O-glucoside, have a preventive effect on the development of atopic dermatitis in oxazolone-treated BALB/c mice and 2,4-dinitrochlorobenzene-treated hairless mice. The epicutaneous applications of oxazolone and 2,4-dinitrochlorobenzene evoke an experimental murine atopic dermatitis-like reaction in BALB/c mouse ears and SKH-1 hairless mice. Atopic skin symptoms, including erythema (redness), pruritus (itching), exudation (weeping), excoriation (peeling), and lichenification (skin thickening), responded to treatment with S. chamaejasme aerial parts EtOH extract for 2 or 3 weeks. Histopathological examination revealed S. chamaejasme aerial parts EtOH extract significantly reduced inflammatory cell infiltration when applied to atopic dermatitis mice. In addition, luteolin 7-O-glucoside, the major active compound of the S. chamaejasme aerial parts EtOH extract, decreased serum IgE and IL-4 levels and transepidermal water loss and increased skin hydration, therefore exhibiting strong anti-atopic dermatitis activity in 2,4-dinitrochlorobenzene-induced atopic dermatitis mice. In this study, we confirmed antipruritic and antidermatitic effects of S. chamaejasme extract and its main component luteolin 7-O-glucoside in atopic dermatitis murine models. The study shows S. chamaejasme aerial parts EtOH extract and luteolin 7-O-glucoside are most likely to be potential drug candidates for atopic dermatitis treatment.

Isolation of maltol derivatives from Stellera chamaejasme and the anti-atopic properties of maltol on skin lesions in DNCB-stimulated mice.

The aim of this study was to isolate maltol derivatives from S. chamaejasme and to investigate the anti-atopic dermatitis (anti-AD) effect of maltol in a 2,4-dinitrochlorobenzene (DNCB)-sensitized mouse model of AD. A novel compound, maltol 3-O-(4'-O-cis-p-coumaroyl)-ß-d-glucoside (named isosoyamaloside I), and two known maltol derivatives (maltol and soyamaloside I) were isolated from S. chamaejasme using chromatographic methods. Dermal application of maltol to DNCB-sensitized AD mice reduced erythema, pruritus, and lichenification scores. Histopathological examinations revealed significant decline in mast cell infiltration in maltol-treated AD mice. In addition, maltol accelerated skin barrier recovery by reducing TEWL and skin pH and increasing skin hydration. Maltol was also found to suppress atopy-induced IL-4 and IgE elevations in serum, which are known to be essential for the development of atopy. The results of this study show that maltol is a potential therapeutic candidate for the treatment of AD-related skin diseases.

Anti-Atopic Properties of Gracillin Isolated from Dioscorea quinqueloba on 2,4-Dinitrochlorobenzene-Induced Skin Lesions in Mice.

Naturally occurring saponins have been reported to have anti-inflammatory and immunomodulatory effects. However, the effects of gracillin, a main saponin component of Dioscorea quinqueloba (D. quinqueloba), on atopic dermatitis (AD), have not been previously studied. The aim of this study was to determine whether gracillin isolated from D. quinqueloba has an anti-AD effect on 2,4-dinitrochlorobenzene (DNCB)-induced AD-like skin lesions in SKH-1 hairless mice. Topical co-treatment of gracillin and DNCB for two weeks markedly reduced symptoms typical of AD (redness, itching, swelling and skin lichenification), decreased transepidermal water loss (TEWL) and increased skin hydration. In addition, gracillin strongly inhibited PI-induced IL-4 expression in RBL-2H3 cells and in the skins of AD mice. Our results suggest gracillin is a potential candidate for the prevention and treatment of AD and other inflammatory skin disorders.

A new flavonoid from Stellera chamaejasme L., stechamone, alleviated 2,4-dinitrochlorobenzene-induced atopic dermatitis-like skin lesions in a murine model.

Stellera chamaejasme L. (family Thymelaeaceae), also known as 'Langdu', has been traditionally used to treat of skin-related diseases, such as, psoriasis and skin ulcers. The aim of this study was to identify the biologically active component of S. chamaejasme and evaluate its preventive effects on IL-4 and mast cell degranulation in RBL-2H3 cells and on the development of atopic dermatitis (AD) in 2,4-dinitrochlorobenzene (DNCB)-treated SKH-1 hairless mice. A novel flavonoid, genkwanin 5-O-xylosyl(1 → 2)glucoside (named stechamone), and three known compounds (umbelliferone, luteolin, and luteolin-7-O-glucoside) were isolated from the aerial parts of S. chamaejasme using chromatographic methods. Of these four compounds, stechamone most potently inhibited IL-4 production and mast cell degranulation in RBL-2H3 cells. Topical application of 0.5% stechamone improved atopic skin symptoms, including, erythema (redness), pruritus (itching), exudation (weeping), excoriation (peeling), and lichenification (skin thickening) in DNCB-treated AD mice by accelerating skin barrier recovery function and suppressing inflammatory cell infiltration. In addition, stechamone attenuated DNCB-induced increases in IL-4 (an inflammatory TH2 cytokine) expression and in serum IgE levels in our murine model of AD. DNCB induced AD-like skin lesions, but treatment with stechamone exhibited strong anti-atopic activity by regulating skin barrier function and reducing inflammatory responses. The results obtained suggest stechamone is a potential anti-atopic agent and treatment for skin inflammatory diseases.