Jung A Seo

Sulforaphane Suppresses Oligomerization of TLR4 in a Thiol-Dependent Manner

TLRs are pattern recognition receptors that detect invading microorganisms and nonmicrobial endogenous molecules to trigger immune and inflammatory responses during host defense and tissue repair. TLR activity is closely linked to the risk of many inflammatory diseases and immune disorders. Therefore, TLR signaling pathways can provide efficient therapeutic targets for chronic diseases. Sulforaphane (SFN), an isothiocyanate, has been well known for its anti-inflammatory activities. In this study, we investigated the modulation of TLR activity by SFN and the underlying mechanism. SFN suppressed ligand-induced and ligand-independent TLR4 activation because it prevented IL-1R–associated kinase-1 degradation, activation of NF-κB and IFN regulatory factor 3, and cyclooxygenase-2 expression induced by LPS or overexpression of TLR4. Receptor oligomerization, which is one of the initial and critical events of TLR4 activation, was suppressed by SFN, resulting in the downregulation of NF-κB activation. SFN formed adducts with cysteine residues in the extracellular domain of TLR4 as confirmed by liquid chromatography-tandem mass spectrometry analysis and the inhibitory effects of SFN on oligomerization and NF-κB activation were reversed by thiol donors (DTT and N-acetyl-l-cysteine). These suggest that the reactivity of SFN to sulfhydryl moiety contributes to its inhibitory activities. Blockade of TLR4 signaling by SFN resulted in the reduced production of inflammatory cytokines and the decreased dermal inflammation and edema in vivo in experimental inflammatory animal models. Collectively, our results demonstrated that SFN downregulated TLR4 signaling through the suppression of oligomerization process in a thiol-dependent manner. These present a novel mechanism for beneficial effects of SFN and a novel anti-inflammatory target in TLR4 signaling.

TRPV1 antagonist can suppress the atopic dermatitis-like symptoms by accelerating skin barrier recovery

BACKGROUND Transient receptor potential vanilloid type 1 (TRPV1) is a cation channel activated by diverse obnoxious stimuli like capsaicin, low pH or heat. Recently, it was revealed that TRPV1 might be deeply associated with skin permeability barrier function, suggesting that modulation of TRPV1 might be beneficial for the skin disorders with barrier damages. OBJECTIVE We aimed to investigate whether the blockade of TRPV1 activation might accelerate skin barrier recovery and alleviate atopic dermatitis (AD)-like symptoms, employing a novel TRPV1 antagonist, PAC-14028. METHODS TRPV1 antagonistic effects of PAC-14028 in human keratinocytes and skin were confirmed through capsaicin-evoked calcium influx assay and capsaicin-induced blood perfusion increase. Effects of PAC-14028 on skin barrier recovery were examined in vivo tape-stripping-induced barrier disruption in hairless mice. To determine the effects of PAC-14028 on AD, Dermatophagoides farina (Df)- and oxazolone (OXZ)-induced AD models were employed. RESULTS PAC-14028 could inhibit capsaicin-evoked calcium influx in keratinocytes at sub-micromolar concentrations. This potent TRPV1 antagonistic activity in keratinocytes was manifested in vivo as the blockade of capsaicin-induced blood perfusion increase, and the accelerated barrier recovery from tape-stripping-induced barrier damages in hairless mice. PAC-14028 could also attenuate dermatitis-associated barrier damages in Df and OXZ models as determined by lower TEWL (trans-epidermal water loss), reformation of neutral lipid layer and reversion of changes in loricrin and filaggrin expression. Importantly, along with accelerated recovery of skin barrier function, PAC-14028 alleviated the general AD-like symptoms, including serum IgE increase, mast cell degranulation, scratching behavior and clinical severity of dermatitis. CONCLUSIONS These results reflect that the blockade of TRPV1 activation can suppress the atopic dermatitis-like symptoms by accelerating skin barrier recovery.

Immunohistological comparison of cutaneous pathology of three representative murine atopic dermatitis models

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