Total flavone of flowers of Abelmoschus manihot (L.) Medic inhibits the expression of adhesion molecules in primary mesenteric arterial endothelial cells and ameliorates dextran sodium sulphate-induced ulcerative colitis in mice.

BACKGROUND: Flowers of Abelmoschus manihot (L.) medic (AM) is a traditional Chinese medicine used to treat chronic nephritis, nephrotic syndrome, diabetic nephropathy, and colonic inflammation. PURPOSE: This study aimed to explore the influence of the total flavone of AM flowers (TFA) on acute ulcerative colitis (UC) and the potential underlying mechanism. METHODS: Efficacy of TFA (30, 60, 120 mg/kg) on UC was evaluated in a dextran sodium sulphate (DSS)-induced colonic inflammatory mouse model by analyzing disease activity index (DAI), histopathological score, colon length, and cytokine expression. Expression levels of critical adhesion molecules and nuclear factor kappa B (NF-κB) were examined by qRT-PCR, Western blotting, or immunofluorescence labeling. Myeloperoxidase activity was examined using ELISA. In vitro THP-1 adhesion assay was used to evaluate monocyte adhesion. RESULTS: TFA significantly reduced DAI score, prevented colon shortening, and ameliorated histological injuries of colons in DSS-treated mice. TFA inhibited the expression of cytokines (IL-1ß and TNF-α) and adhesion molecules (ICAM-1, VCAM-1, and MAdCAM-1) in colon tissues of DSS mice. In vitro studies on mesenteric arterial endothelial cells (MAECs) showed that TFA attenuated TNF-α-induced upregulation of ICAM-1, VCAM-1, and MAdCAM-1, as well as THP-1 cell adhesion to MAECs. TFA also suppressed the phosphorylation and nuclear translocation of NF-κB in MAECs. CONCLUSION: TFA efficaciously ameliorates UC possibly by inhibiting monocyte adhesion through blocking TNF-α-induced NF-κB activation, which in turn suppresses the upregulation of adhesive molecules in colon endothelial cells. Inhibiting the expression of adhesion molecule in MAECs may represent a useful strategy for therapeutic development to treat UC, with TFA being a safe and efficacious therapeutic agent.

Scutellarein attenuates atopic dermatitis by selectively inhibiting transient receptor potential vanilloid 3 channels.

BACKGROUND AND PURPOSE: Atopic dermatitis (AD) is one of the most common chronic inflammatory cutaneous diseases with unmet clinical needs. As a common ingredient found in several medicinal herbs with efficacy on cutaneous inflammatory diseases, Scutellarein (Scu) has been shown to possess anti-inflammatory and anti-proliferative activities. We aimed to evaluate the therapeutic efficacy of Scu against AD and its underlying molecular mechanism. EXPERIMENTAL APPROACH: Efficacy of Scu on AD was evaluated in 2,4-dinitrofluorobenzene (DNFB) and carvacrol-induced dermatitis mouse models. Cytokine mRNA and serum IgE levels were examined using qPCR and ELISA, respectively. Voltage clamp recordings were used to measure currents mediated by transient receptor potential (TRP) channels. In silico docking, site-direct mutagenesis, and covalent modification were used to explore the binding pocket of Scu on TRPV3. KEY RESULTS: Subcutaneous administration of Scu efficaciously suppresses DNFB and carvacrol-induced pruritus, epidermal hyperplasia and skin inflammation in wild type mice but has no additional benefit in Trpv3 knockout mice in the carvacrol model. Scu is a potent and selective TRPV3 channel allosteric negative modulator with an apparent affinity of 1.18 µM. Molecular docking coupled with site-direct mutagenesis and covalent modification of incorporated cysteine residues demonstrate that Scu targets the cavity formed between the pore helix and transmembrane helix S6. Moreover, Scu attenuates endogenous TRPV3 activity in human keratinocytes and inhibits carvacrol-induced proliferative and proinflammatory responses. CONCLUSION AND IMPLICATIONS: Collectively, these data demonstrate that Scu ameliorates carvacrol-induced skin inflammation by directly inhibiting TRPV3, and TRPV3 represents a viable therapeutic target for AD treatment.

TRPV1 SUMOylation regulates nociceptive signaling in models of inflammatory pain.

Although TRPV1 channels represent a key player of noxious heat sensation, the precise mechanisms for thermal hyperalgesia remain unknown. We report here that conditional knockout of deSUMOylation enzyme, SENP1, in mouse dorsal root ganglion (DRG) neurons exacerbated thermal hyperalgesia in both carrageenan- and Complete Freund's adjuvant-induced inflammation models. TRPV1 is SUMOylated at a C-terminal Lys residue (K822), which specifically enhances the channel sensitivity to stimulation by heat, but not capsaicin, protons or voltage. TRPV1 SUMOylation is decreased by SENP1 but upregulated upon peripheral inflammation. More importantly, the reduced ability of TRPV1 knockout mice to develop inflammatory thermal hyperalgesia was rescued by viral infection of lumbar 3/4 DRG neurons of wild-type TRPV1, but not its SUMOylation-deficient mutant, K822R. These data suggest that TRPV1 SUMOylation is essential for the development of inflammatory thermal hyperalgesia, through a mechanism that involves sensitization of the channel response specifically to thermal stimulation.

Subunit-specific inhibition of glycine receptors by curcumol.

Emerging evidence has suggested that inhibitory glycine receptors (GlyRs) are an important molecular target in the treatment of numerous neurological disorders. Rhizoma curcumae is a medicinal plant with positive neurological effects. In this study, we showed that curcumol, a major bioactive component of R. curcumae, reversibly and concentration-dependently inhibited the glycine-activated current (IGly) in cultured rat hippocampal neurons. The inhibitory effect was neither voltage- nor agonist concentration-dependent. Moreover, curcumol selectively inhibited homomeric α2-containing, but not α1- or α3-containing, GlyRs. The addition of ß subunit conferred the curcumol sensitivity of α3-containing, but not α1-containing, GlyRs. Site-directed mutagenesis analysis revealed that a threonine at position 59 of the α2 subunit is critical for the susceptibility of GlyRs to curcumol-mediated inhibition. Furthermore, paralleling a decline of α2 subunit expression during spinal cord development, the degree of IGly inhibition by curcumol decreased with prolonged culture of rat spinal dorsal horn neurons. Taken together, our results suggest that the GlyRs are novel molecular targets of curcumol, which may underlie its pharmaceutical effects in the central nervous system.

Calmodulin regulates Ca(2+)-dependent feedback inhibition of store-operated Ca(2+) influx by interaction with a site in the C terminus of TrpC1.

The mechanism involved in [Ca(2+)](i)-dependent feedback inhibition of store-operated Ca(2+) entry (SOCE) is not yet known. Expression of Ca(2+)-insensitive calmodulin (Mut-CaM) but not wild-type CaM increased SOCE and decreased its Ca(2+)-dependent inactivation. Expression of TrpC1 lacking C terminus aa 664-793 (TrpC1DeltaC) also attenuated Ca(2+)-dependent inactivation of SOCE. CaM interacted with endogenous and expressed TrpC1 and with GST-TrpC1 C terminus but not with TrpC1DeltaC. Two CaM binding domains, aa 715-749 and aa 758-793, were identified. Expression of TrpC1Delta758-793 but not TrpC1Delta715-749 mimicked the effects of TrpC1DeltaC and Mut-CaM on SOCE. These data demonstrate that CaM mediates Ca(2+)-dependent feedback inhibition of SOCE via binding to a domain in the C terminus of TrpC1. These findings reveal an integral role for TrpC1 in the regulation of SOCE.