Topical Application Of Oxazolone Research Articles

Inhibition of gasdermin D (GSDMD) as a promising therapeutic approach for atopic dermatitis

Atopic dermatitis (AD) is a chronic inflammatory skin disease characterized by pruritus, erythema, and skin barrier dysfunction. Gasdermin D (GSDMD) is the key executioner of an inflammatory cell death mechanism known as pyroptosis. However, the role of GSDMD in the pathogenesis of AD remains unclear. Through the analysis of publicly available Gene Expression Omnibus (GEO) datasets, we observed an upregulation of Gsdmd mRNA in the skin tissue of AD patients. Moreover, we delved into the impact of GSDMD deletion and inhibition on AD-like skin lesions using a mouse model induced by the topical application of oxazolone (Oxa). We found that mice lacking GSDMD exhibited relieved AD signs and symptoms in terms of reduced skin thickness, scarring and scratching behavior compared to wild-type mice after induction of AD-like skin lesions. This was associated with decreased infiltration of inflammatory cells, reduced epidermal thickness, and decreased serum levels of IgE and IL-4. Western blot analysis further revealed decreased GSDMD cleavage in the skin of GSDMD knockout mice, and reduced expression of IL-1β and IL-18. Inhibition of GSDMD using the pharmacological agent disulfiram or the herbal compound matrine significantly attenuated the symptoms of AD-like skin lesions in wild-type mice, GSDMD cleavage and pro-inflammatory cytokines were reduced as well. Our results suggest that GSDMD-mediated pyroptosis plays a critical role in the development of AD-like skin lesions, and targeting GSDMD may be a promising therapeutic strategy for AD.

Histopathologic Features in Animal Model of Atopic Dermatitis Induced by Topical Application of Oxazolone

Animal models of atopic dermatitis (AD) are widely used to investigate therapeutic effects of candidates for AD. However, the characteristics of each model are not fully understood. This study was designed to compare the animal models of dermatitis induced by dinitrofluorobenzene (DNFB) and oxazolone (Ox). We investigated the effects of DNFB and Ox on skin thicknesses and weights as well as skin lesions associated with AD such as scale, crust and erythematous eruption, and histopathological changes such as hyperkeratosis, dermal and epidermal hyperplasia and immune cell infiltration in inflamed tissues. Multiple application of 0.5% Ox onto the skin increased skin thickness and weight compared to those of DNFB treated mice, as well as those of normal mice. In addition, topical application of DNFB induced marked scale, crust and erythematous eruption, while Ox induced erythematous eruption and mild scale and crust. Histopathological examination revealed that 0.5% Ox induced marked hyperplasia in the dermis and epidermis, large vesicles, spongiotic changes, mild hyperkeratosis and immune cell infiltration in balb/c mice. These data suggest that multiple applications of Ox can induce chronic AD like dermatitis in balb/c mice.

380. Use of Somatotransgenic Bioimaging as a Platform for Studying NFkB Pathway in a Tissue-Specific Manner in Two Mouse Models of Inflammation: Rheumatoid Arthritis and Contact Dermatitis

Somatotransgenic bioimaging is a technology for in vivo imaging using lentiviral mediated delivery of a luciferase construct into neonatal mice. This luciferase marker, under the control of transcription factor binding elements or surrogate promoters, can be activated during physiological and pathological processes and can be quantified as photonic readout by whole-body bioluminescence imaging technology. The technique permits lifelong expression with immune tolerance to the transgenic proteins.In this study, we show the utility of somatotransgenic bioimaging using NFkB signaling as a model of inflammation in mouse models of rheumatoid arthritis and contact dermatitis.An NFkB transcription factor response element was inserted upstream of a mouse minimal promoter and the genes encoding firefly luciferase and green fluorescent protein. The cassette was inserted into a lentivirus vector.VSV-G-pseudotyped lentivirus vector was injected subcutaneously into CD1 outbread neonatal (P0) mice either in the footpad or in the abdominal skin to generate paw or skin specific light emitting transgenic mice.Canonical NFkB signaling (through TLR4) was first validated using ultrapure LPS in both the paw and skin of somatic transgenic mice. Following intraperitoneal LPS injection we observed upregulation of luciferase activity in the paw or the skin which returned to background levels within a few days.As a model of inflammatory mediated rheumatoid arthritis, we injected zymosan, which activates NFkB via TLR2 into the mouse paw fat pad. Induction of contact dermatitis was achieved by applying topically oxazolone, which plays an indirect activator role in the NFkB pathway. Luciferase expression was then monitored by whole body bioluminescence.Stimulation with Zymosan of the paw biosensor mice induced a prolonged upregulation of luciferase expression, specifically into the mouse paw which peaked at day 3 post injection and gradually returned to background level after twenty days.Similarly, after topical application of oxazolone onto the skin biosensor mice, we observed a specific, localised and increased luciferase activity which peaked at day 2 and returned to baseline by day 5.We have therefore shown that targeted gene expression can be achieved in two mouse models of inflammation. This also allowed us to quantify NFkB-mediated response to pro-inflammatory stimuli in a disease- and tissue-specific manner. Applied to collagen-induced arthritis, this technique could improve quantitation of disease severity compared to subjective scoring.

Inhibition of Neurite Outgrowth by a Neuropilin-1 Binding Peptide Derived from Semaphorin 3A

Semaphorin 3A (Sema3A), an axon guidance molecule, inhibits neurite outgrowth of sensory neurons. Recombinant Sema3A protein has also inhibited scratching behavior and improved skin inflammation in an atopic dermatitis model. In the present study, we investigated whether Sema3A-derived peptides could bind its receptor, neuropilin-1 (NRP1), to inhibit neurite outgrowth. Here, two candidate NRP1-binding (NPB) peptides, NPB7 and NPB15, were found to inhibit NGF-induced survival and neurite outgrowth of PC12 cells and rat primary neurons in serum-free medium. To investigate the preventive effect of the two NPB peptides in vivo, we assessed whether they could inhibit skin inflammation induced by repeated topical application of oxazolone in mice. NPB15 peptide, but not NPB7, inhibited ear swelling. The NPB15 peptide solution and Vaseline ointment groups showed slightly decreased epidermal nerve densities compared with controls. The combination of NPB15 peptide and Vaseline ointment increased the inhibitory effect of NPB15 peptide on epidermal nerve densities. These results suggest that Sema3A-derived peptides can bind to NRP1 and inhibit neurite outgrowth both in vitro and in vivo. Thus, these peptides may be potent candidates for the treatment of atopic dermatitis.

Topical Apigenin Alleviates Cutaneous Inflammation in Murine Models

Herbal medicines have been used in preventing and treating skin disorders for centuries. It has been demonstrated that systemic administration of chrysanthemum extract exhibits anti-inflammatory properties. However, whether topical applications of apigenin, a constituent of chrysanthemum extract, influence cutaneous inflammation is still unclear. In the present study, we first tested whether topical applications of apigenin alleviate cutaneous inflammation in murine models of acute dermatitis. The murine models of acute allergic contact dermatitis and acute irritant contact dermatitis were established by topical application of oxazolone and phorbol 12-myristate 13-acetate (TPA), respectively. Inflammation was assessed in both dermatitis models by measuring ear thickness. Additionally, the effect of apigenin on stratum corneum function in a murine subacute allergic contact dermatitis model was assessed with an MPA5 physiology monitor. Our results demonstrate that topical applications of apigenin exhibit therapeutic effects in both acute irritant contact dermatitis and allergic contact dermatitis models. Moreover, in comparison with the vehicle treatment, topical apigenin treatment significantly reduced transepidermal water loss, lowered skin surface pH, and increased stratum corneum hydration in a subacute murine allergic contact dermatitis model. Together, these results suggest that topical application of apigenin could provide an alternative regimen for the treatment of dermatitis.

The Signaling Adaptor Eps8 Is an Essential Actin Capping Protein for Dendritic Cell Migration

Dendritic cells (DCs) flexibly adapt to different microenvironments by using diverse migration strategies that are ultimately dependent on the dynamics and structural organization of the actin cytoskeleton. Here, we have shown that DCs require the actin capping activity of the signaling adaptor Eps8 to polarize and to form elongated migratory protrusions. DCs from Eps8-deficient mice are impaired in directional and chemotactic migration in 3D invitro and are delayed in reaching the draining lymph node (DLN) invivo after inflammatory challenge. Hence, Eps8-deficient mice are unable to mount a contact hypersensitivity response. We have also shown that the DC migratory defect is cell autonomous and that Eps8 is required for the proper architectural organization of the actin meshwork and dynamics of cell protrusions. Yet, Eps8 is not necessary for antigen uptake, processing, and presentation. Thus, we have identified Eps8 as a unique actin capping protein specifically required for DC migration.

Contact sensitivity in the murine oral mucosa. I. An experimental model of delayed-type hypersensitivity reactions at mucosal surfaces

We have examined in a murine model, the potential of the oral mucosa (OM) to serve as inductive and/or expression site(s) of delayed-type hypersensitivity (DTH) reactions. The expression of DTH reactions in the murine buccal mucosa was studied after topical application of oxazolone or picryl chloride onto the OM of animals previously sensitized with either hapten. Irrespective of the site of priming (skin or buccal mucosa), inflammatory cells appeared in the OM following buccal elicitation with the pertinent hapten. The density of infiltrating cells peaked at 24 h after hapten elicitation. Such inflammatory reactions, which comprised mainly mononuclear cells at 24 h, were preceded by an early inflammatory reaction that developed only in animals previously sensitized at skin sites. This early reaction, comprising mainly PMN neutrophils, peaked at 6-8 h, declined by 8-16 h, and was not observed in mice previously sensitized in the buccal mucosa. The 24 h reactions failed to develop in nude mice similarly treated, in intact unsensitized mice, as well as in animals sensitized with an irrelevant hapten. These reactions could be adoptively transferred to naive animals by LN cells but not by serum from sensitized syngeneic donors. Furthermore, LN cell suspensions depleted of T cells failed to transfer sensitization for subsequent OM DTH. Topical application of contact sensitizing haptens onto OM induced priming for subsequent DTH reactions elicited with recall antigen applied at a distant skin site or at a local buccal site. These results demonstrate that the OM has the capacity to serve both as an inductive and as an expression site for T cell-mediated inflammatory reactions, be these expressed or induced at local mucosal sites or at remote systemic (skin) sites. This animal model should be valuable for studying the regulation of T cell-mediated inflammatory responses at mucosal surfaces.

Serum Amyloid A-Luciferase Transgenic Mice: Response to Sepsis, Acute Arthritis, and Contact Hypersensitivity and the Effects of Proteasome Inhibition

Acute phase serum amyloid A proteins (A-SAAs) are multifunctional apolipoproteins produced in large amounts during the acute phase of an inflammation and also during the development of chronic inflammatory diseases. In this study we present a Saa1-luc transgenic mouse model in which SAA1 gene expression can be monitored by measuring luciferase activity using a noninvasive imaging system. When challenged with LPS, TNF-alpha, or IL-1beta, in vivo imaging of Saa1-luc mice showed a 1000- to 3000-fold induction of luciferase activity in the hepatic region that peaked 4-7 h after treatment. The induction of liver luciferase expression was consistent with an increase in SAA1 mRNA in the liver and a dramatic elevation of the serum SAA1 concentration. Ex vivo analyses revealed luciferase induction in many tissues, ranging from several-fold (brain) to >5000-fold (liver) after LPS or TNF-alpha treatment. Pretreatment of mice with the proteasome inhibitor bortezomib significantly suppressed LPS-induced SAA1 expression. These results suggested that proteasome inhibition, perhaps through the NF-kappaB signaling pathway, may regulate SAA1 expression. During the development of acute arthritis triggered by intra-articular administration of zymosan, SAA1 expression was induced both locally at the knee joint and systemically in the liver, and the induction was significantly suppressed by bortezomib. Induction of SAA1 expression was also demonstrated during contact hypersensitivity induced by topical application of oxazolone. These results suggest that both local and systemic induction of A-SAA occur during inflammation and may contribute to the pathogenesis of chronic inflammatory diseases associated with amyloid deposition.

Inhibition of contact sensitivity in the mouse by topical application of corticosteroids.

1. Topical application of oxazolone to the ears of mice previously sensitized to this hapten resulted in a marked increase in ear weight 24 h later.2. Topical application of hydrocortisone, betamethasone 17-valerate, fluocinolone acetonide or triamcinolone acetonide inhibited this response in a dose dependent manner.3. The free alcohols of the three fluorinated compounds were only weakly active.4. Activity in this model was truly local. Oral administration of hydrocortisone or fluocinolone acetonide at doses comparable to those used topically failed to inhibit the ear swelling.5. Some steroids with poor anti-inflammatory properties had no significant effect.