Sarsasapogenin and fluticasone combination improves DNFB induced atopic dermatitis lesions in BALB/c mice.

OBJECTIVE: Atopic dermatitis (AD) is a pruritic, chronic, relapsing inflammatory skin disease. The research aims to study the effects of Sarsasapogenin and its combination with Fluticasone in 2, 4-Dinitrofluorobenzene (DNFB) induced atopic dermatitis in BALB/c mice. MATERIAL AND METHODS: Thirty male Balb/c mice were divided into 5 groups: (i) Normal control (NC), (ii) Disease control (DNFB), (iii) Sarsasapogenin (SG) (50 µg/mice), (iv) Fluticasone (FC) (50 µg/mice), (v) Sarsasapogenin + Fluticasone (SG + FC) combination (25 µg/mice). Dermatitis was induced by repeated application of DNFB in Balb/c mice. On topical application of SG, FC, and SG + FC combination on the ear and skin lesions, body weight, ear weight, ear thickness, erythema score, spleen weight, cytokines, immunoglobulin E (IgE) levels, nitric oxide (NO) level, hematological parameters, and oxidative stress markers were evaluated. Histological analysis of the ear tissue was also done. RESULTS: The results stated that SG and SG + FC treatment to mice considerably decrease the ear weight, ear thickness, spleen weight, serum IgE, cytokines, NO levels, and restoration of antioxidant stress markers with elevation in the hematological parameters. The observations were further confirmed by histopathological analysis of ear tissue. CONCLUSION: These data specify that SG has been demonstrated as a probable therapy for the treatment of allergic skin diseases in combination with FC by decreasing its dose from 50 to 25 µg/mice to avoid the chronic side effects of FC. Hence, it can be concluded that SG and SG + FC combination significantly improved the AD-like symptoms in the DNFB sensitized mice through mitigating the production of proinflammatory mediators and restoration of oxidative stress markers.

Qingxue jiedu formulation ameliorated DNFB-induced atopic dermatitis by inhibiting STAT3/MAPK/NF-κB signaling pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Qingxue jiedu Formulation (QF) is composed of two classic prescriptions which have been clinically used for more than 5 centuries and appropriately modified through basic theory of traditional Chinese medicine for treating various skin inflammation such as atopic dermatitis (AD), acute dermatitis and rash. Although QF possesses a prominent clinical therapeutic effect, seldom pharmacological studies on its anti-AD activity are conducted. AIM OF THE STUDY: We used AD mice model to investigate the anti-AD activities of QF, as well as its underlying molecular mechanisms which involved signal transducer and activator of transcription 3 (STAT3), nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) signaling pathways. MATERIALS AND METHODS: 2,4-dinitrofluorobenzene (DNFB)-induced AD mice were used to collect serum and skin tissues for consequential determination. The levels of various inflammatory factors [interleukin (IL)-12, Interferon (IFN)-γ, tumor necrosis factor (TNF)-α, IL-4, IL-6 and immunoglobulin E (IgE)] were determined by enzyme-linked immunosorbent assay (ELISA). Real-time polymerase chain reaction (RT-PCR) was contributed to detect the effects of relevant inflammatory factors on mRNA. The roles of STAT3, NF-κB and MAPK signaling pathways in AD response were analyzed by Western blotting (WB), and the thickening of mice dorsal skin and inflammatory cell infiltration were observed by hematoxylin and eosin (H&E) staining. RESULTS: QF significantly reduced the skin thickening, inflammatory cell infiltration and other symptoms in AD mice. The levels of IL-12, TNF-α, IL-4, IL-6 and IgE were decreased, while IFN-γ was increased by QF in the ELISA analysis. QF lessened the levels of lL-6 and elevated IFN-γ on the mRNA level. In addition, WB analysis showed QF thoroughly inhibited the activation of NF-κB, STAT3 and phosphorylation of JAK1, JAK2, JAK3, while partially suppressed MAPK signaling pathways. CONCLUSIONS: QF inhibited the activations of STAT3, MAPK and NF-κB signaling pathways and possessed a significant therapeutic effect on AD. Therefore, QF deserves our continuous attention and research as a prominent medicine for AD.

Prediction of the therapeutic mechanism responsible for the effects of Sophora japonica flower buds on contact dermatitis by network-based pharmacological analysis.

ETHNOPHARMACOLOGICAL RELEVANCE: The flower buds of Sophora japonica L. are a major traditional medicine in China, Japan, and Korea and are used to stop bleeding and 'cool the blood'. Accordingly, they are used to treat bleeding haemorrhoids, hypertension, and pyoderma. In addition, it was recently found that the flower buds of S. japonica (SJ) have cosmetic whitening properties. MATERIALS AND METHODS: Compounds in SJ and their targets and related diseases were investigated using the Traditional Chinese Medicine Systems Pharmacology (TCMSP) database and analysis platform. Target gene information was obtained from the UniProt database. Network construction was carried out using Cytoscape 3.72. Contact dermatitis (CD)-related gene searching was performed using the Cytoscape string App. Docking analysis was conducted using AutoDock Vina. Six-week-old Balb/c male mice with DNFB (1-fluoro-2,4-dinitrofluorobenzene)-induced CD were treated with a methanol extract of the flower buds of S. japonica (MESJ), and its effects on skin colour, lesions, and immune cell infiltration, and on histopathological abnormalities such as epidermal hyperplasia were investigated. RESULTS: Eleven compounds targeted 13 CD-related genes, that is, serum albumin (ALB), prostaglandin G/H synthase (COX) 2, C-X-C motif chemokine (CXCL) 2, CXCL10, ICAM1, IFN-γ, IL-10, IL-1α, IL-1ß, IL-2, IL-6, E-selectin, and TNF. In the murine DNFB model, MESJ significantly suppressed scaling, erythema, and skin thickening as compared with DNFB controls and epithelial hyperplasia and immune cell infiltrations induced by repeated DNFB application. CONCLUSIONS: Our animal study showed that the mode of action of MESJ was closely related to the prevention of epithelial hyperplasia and immune cell infiltration. The results obtained demonstrated that the flower buds of S. japonica offer a potential means of treating CD, and suggest that the therapeutic mechanism of CD is explained by relations between 11 major components of SJ, including kaempferol and quercetin, and 13 CD-related genes.

Omega-3 fatty acid-derived mediator, Resolvin E1, ameliorates 2,4-dinitrofluorobenzene-induced atopic dermatitis in NC/Nga mice.

Atopic dermatitis (AD) is a common inflammatory skin disease for which few effective treatments are available. Resolvin E1 (RvE1; 5S,12R,18R-trihydroxy-6Z,8E,10E,14Z,16E-eicosapentaenoic acid) is an endogenous lipid mediator derived from omega-3 fatty eicosapentaenoic acid, which is a potent inhibitor of inflammation. AD-like skin lesion was induced by repetitive skin contact with DNFB in NC/Nga mice and the effects of RvE1 were evaluated on the basis of histopathological findings of skin, ear swelling and cytokine production of CD4(+) T cells. Intraperitoneal injection of RvE1 for one week after DNFB challenge significantly lowered ear swelling and improved back skin lesions. In addition, RvE1 significantly suppressed production of interferon-gamma (IFN-γ) and interleukin-4 (IL-4) by activated CD4(+) T cells and serum IgE level. Furthermore, RvE1 reduced DNFB-induced infiltration of eosinophils, mast cells, CD4(+) T cells, and CD8(+) T cells in skin lesions. Therefore, RvE1 may suppress the development of AD-like skin lesions in DNFB-treated NC/Nga mice by reducing IL-4 and IFN-γ of activated CD4(+) T cells and serum IgE levels and infiltration of immune cells to skin lesion.

Effect of Pyeongwee-San (KMP6) on 2,4-dinitrofluorobenzene-induced atopic dermatitis-like skin lesions in NC/Nga mice.

AIMS: Recently, some studies reported that digestive tract disease is closely associated with atopic dermatitis (AD). Pyeongwee-San (KMP6) is a Korean medicine, which has come onto the drugstore for the treatment of digestive tract disease. The aim of the present study was to examine whether KMP6 could suppress 2,4-dinitrofluorobenzene (DNFB)-induced AD-like skin lesions in NC/Nga mice. MAIN METHODS: Mice were sensitized with DNFB by applying to shaved dorsal skin. At that time, the drugs or saline were orally administrated to DNFB-applied mice. KEY FINDINGS: The administration of KMP6 or glycyrrhizic acid (GL), a major component of KMP6, inhibited the scratching number in DNFB-induced AD model. The mRNA expressions of interleukin (IL)-4, interferon (IFN)-γ, tumor necrosis factor (TNF)-α, and CCR3 were upregulated by DNFB sensitization, but the upregulated mRNA expressions were significantly reduced by the administration of KMP6 or GL. In addition, the levels of IgE, histamine, and IL-4 were significantly reduced by the administration of KMP6 or GL in serum of DNFB-induced AD model. However, the level of IFN-γ in serum was significantly increased by KMP6 or GL. KMP6 or GL also significantly inhibited the numbers of inflammatory cells, mast cells, and protein level of IL-4 in lesions of DNFB-induced AD model. Finally, KMP6 or GL significantly decreased the productions of IL-4, IFN-γ, and TNF-α in anti-CD3 plus anti-CD28 antibody-stimulated splenocytes. SIGNIFICANCE: KMP6 showed anti-atopic potential in this setting; hence we suggest it as a potential prospect for anti-atopic agent besides being just a medicine for the stomach and bowels.

Anti-inflammatory and immunomodulatory effects of bortezomib in various in vivo models.

Bortezomib (Velcade®) is a proteasome inhibitor that has been approved for the treatment of multiple myeloma and mantle cell lymphoma. It has been shown to inhibit the expression of cell adhesion molecules, co-stimulatory molecules, and NFκB activation, to deplete alloreactive T lymphocytes, and to decrease Th1 cytokine production. The anti-inflammatory effects of bortezomib were further investigated in this current set of studies. Systemic treatment with bortezomib was efficacious in the thioglycolate-induced MCP-1 production model, and the dinitrofluorobenzene-induced delayed-type hypersensitivity model. Psoriasis is an autoimmune disease that affects about 2% of the world population. Many treatments have been reported with varying degrees of efficacy. A topical bortezomib formulation was developed to minimize systemic exposure. Its tolerability was investigated in a topical imiquimod (IMQ)-induced psoriasis model. Daily application of IMQ on mouse skin induced inflamed scaly skin lesions resembling plaque-type psoriasis. Fatality was observed in the 1-mg/ml dose group. At 0.1 and 0.01 mg/ml, bortezomib potentiated IMQ-induced erythema, scaling, skin thickening, and caused necrotic lesions. Lower doses had no effect on the clinical observations. Histologically, bortezomib dose-dependently increased parakeratosis, hyperkeratosis, acanthosis, and inflammatory cell infiltration. This study demonstrated that topical bortezomib is not suitable for the treatment of psoriasis.

Alleviation of atopic dermatitis-related symptoms by Perilla frutescens Britton.

To ascertain whether an aqueous fraction of Perilla frutescens Britton (PfB/af) has advantageous anti-atopic dermatitis activity, we used a 2,4-dinitrofluorobenzene (DNFB)-induced animal model of atopic dermatitis symptoms to investigate the effects of the extract. We performed an ear swelling assay by comparing thickness of the DNFB-induced ear, and measured the numbers of eosinophils as well as total immune cells. We analyzed the expression levels of matrix metalloproteinase (MMP)-9, interleukin (IL)-31 and of the T-bet transcription factor. The results revealed that PfB/af (100 µg/ml) exhibited strong anti-atopic dermatitis activity, interceding drastic reduction (35%) of the immune response, as measured by the thickness of ear epidermis swelling, and resulting in decreased eosinophil levels (73.7%) in adjacent skin tissues. Collectively, the present results suggest that PfB/af has potential for mitigation of atopic dermatitis-like symptoms induced by DNFB in the mouse.

Inhibitory effect of Platycodon grandiflorum on T(H)1 and T(H)2 immune responses in a murine model of 2,4-dinitrofluorobenzene-induced atopic dermatitis-like skin lesions.

BACKGROUND: Platycodon grandiflorum is a traditional Asian medicine that is used to treat pulmonary and respiratory allergic disorders. OBJECTIVE: to investigate the effects of P grandiflorum in vivo in an animal model of atopic dermatitis (AD), with particular emphasis on its effects on T(H)1 and T(H)2 immune responses. METHODS: we established a model of AD-like skin lesions in NC/Nga mice. After oral administration of P grandiflorum, we measured cytokine and immunoglobulin profiles along with histologic examination of skin. RESULTS: P grandiflorum was nontoxic in a 2,4-dinitrofluorobenzene-induced model of AD-like skin lesions in NC/Nga mice. AD symptoms in skin lesions improved after oral administration of P grandiflorum. IgE secretion was significantly downregulated in P grandiflorum-treated animals, accompanied by decreased levels of interleukin (IL) 4 and IgG1 and increased serum levels of IL-12p40 and IgG2a. In isolated splenocytes, the production of the T(H)1 cytokines IL-12p40 and interferon-γ was upregulated by P grandiflorum, whereas the levels of the T(H)2 cytokines IL-4 and IL-5 were downregulated in a mouse model of AD-like skin lesions. CONCLUSIONS: these results suggest that P grandiflorum inhibits the development of AD-like skin lesions in NC/Nga mice by suppressing the T(H)2 cell response and increasing the T(H)1 cell responses. Our results indicate that P grandiflorum is safe and effective as a natural herbal medicine for the treatment of AD-like skin lesions.

Stable form of galectin-9, a Tim-3 ligand, inhibits contact hypersensitivity and psoriatic reactions: a potent therapeutic tool for Th1- and/or Th17-mediated skin inflammation.

Tim-3 is a cell surface molecule preferentially expressed in Th1 and Th17 cells. Galectin-9 is a ligand for Tim-3 and the binding of galectin-9 to Tim-3 induces apoptosis. We recently developed a stable form of galectin-9 (sGal-9) by partial deletion of the linker peptide. In this study, we characterized the therapeutic effects of sGal-9 on inflammatory reactions in contact hypersensitivity and IL-23-induced psoriatic mouse models. In contact hypersensitivity in mice, the ear swelling response was suppressed by sGal-9. In vitro treatment with sGal-9 resulted in cell apoptosis of CD4, CD8, and hepatic NK cells. sGal-9-treated mice had decreased IFN-gamma- and IL-17-producing T cells. Similarly, sGal-9 reduced epidermal thickness and dermal cellular infiltrate levels in IL-23-induced psoriasis-like skin inflammation. This was accompanied by decreased skin lesion levels of IL-17 and IL-22. sGal-9 may be a unique and useful therapeutic tool for the treatment of Th1- and/or Th17-mediated skin inflammation.

Oral administration of Astragalus membranaceus inhibits the development of DNFB-induced dermatitis in NC/Nga mice.

Epicutaneously administered chemical antigens like 2,4-dinitrofluorobenzene (DNFB), evoke an atopic dermatitis (AD)-like dermatitis reaction in NC/Nga mice under specific pathogen free (SPF) conditions. Astragalus membranaceus (AM), is a popular herbal medicine used to treat allergic diseases in East Asia. In the present study, we examined whether AM suppress AD-like skin lesions in NC/Nga mice treated with DNFB under SPF conditions. Oral administration of AM to DNFB-treated NC/Nga mice was found to inhibit ear thickness increases and the skin lesions induced by DNFB. Moreover, IFN-gamma production by CD4(+) T cells from the lymph nodes of DNFB-treated NC/Nga mice was significantly inhibited by AM treatment, although levels of IL-4 and total IgE in serum were not. Study findings suggest that AM may suppress the development of AD-like dermatitis in DNFB-treated NC/Nga mice by reducing IFN-gamma production.

Amelioration of experimental colitis by Astragalus membranaceus through anti-oxidation and inhibition of adhesion molecule synthesis.

AIM: To investigate the protective effects of Astragalus membranaceus (Am) against hapten-induced colitis in male Sprague-Dawley rats as well as its underlying mechanism. METHODS: Experimental colitis was induced in rats by enema administration of 2,4-dinitrobenzene sulfonic acid (DNBS). Rats were either pretreated with Am extract (2 or 4 g/kg, p.o. once daily) starting from 10 d before DNBS enema, or received Am post-treatment (2 or 4 g/kg, p.o. twice daily) on the three consecutive days following DNBS administration. Colonic lesion area and histological damage were determined, while the activities of myeloperoxidase (MPO) and xanthine oxidase, as well as reduced glutathione (GSH) content were measured in the excised colonic tissues. Besides, protein expression of inducible nitrite oxide synthase (iNOS), intercellular adhesion molecule-1 (ICAM-1) and P-selectin was also detected by Western blot analysis. RESULTS: Our findings had shown that both macroscopic lesion area and histological colonic damage induced by DNBS were significantly reduced by both Am pre- and post-treatments. These were accompanied by attenuation of the elevated colonic MPO activity and downregulation of the iNOS, P-selectin, and ICAM-1 protein expression. Besides, deprivation of colonic GSH level under colitis condition was also preserved. CONCLUSION: These results demonstrate that Am possesses both preventive and therapeutic potential in experimental colitis. The anti-inflammatory actions involve anti-oxidation along with inhibition of adhesion molecule synthesis in the colonic tissues.

[Il-10 expression is involved in the regulation of the immune response by omega 3 fatty acids]. / La expresión de IL-10 interviene en la regulación de la respuesta inflamatoria por los acidos grasos omega 3.

INTRODUCTION: Polyunsaturated fatty acids play a key role in a huge number of biological functions. Western diets are highly rich in w-6 fatty acids. However the content of w-3 fatty acids is not suitable in those diets, despite of their importance in normal development of the human body and regulation of immune response. The aim of this work is to examine the effect of w-3 fatty acids enriched diet in the regulation of inflammatory response. MATERIAL AND METHODS: Balb/c mice were fed either w-6 fatty acids rich diet (100% sunflower oil) or w-3 fatty acids fortified diet (12% fish oil plus 88% sunflower oil) during 28 days. Twelve hours prior to sacrifice, the mice were treated with 2,4-ninitro-1-fluorobezene on the left ear to induce the inflammatory reaction. Afterwards the mice were sacrificed and the different samples collected were analized. RESULTS: Ear inflammation of mice fed the w-3 diet was significantly lower. Leukocyte infiltration and oxidative stress were also lower in those mice. To explain these results, cytokine expression and plasma eicosanoid concentration were measured. An increase in IL-10 levels and a down regulation of Th1 and Th2 responses were observed in mice fed the w-3 diet. CONCLUSION: Not only n-3 fatty acids exerts an antiinflammatory and an antialergical role but also they enhance some of the organism defenses. Our data suggest that w-3 fatty acids downregulate the inflammatory response by enhancing IL10 expression.

Treatment of silymarin, a plant flavonoid, prevents ultraviolet light-induced immune suppression and oxidative stress in mouse skin.

It is well documented that ultraviolet (UV) light-induced immune suppression and oxidative stress play an important role in the induction of skin cancers. Earlier, we have shown that topical treatment of silymarin, a plant flavonoid from milk thistle (Silybum marianum L. Gaertn.), to mouse skin prevents photocarcinogenesis, but the preventive mechanism of photocarcinogenesis in vivo animal system by silymarin is not well defined and understood. To define the mechanism of prevention, we employed immunostaining, analytical assays and ELISA which revealed that topical treatment of silymarin (1 mg/cm2 skin area) to C3H/HeN mice inhibits UVB (90 mJ/cm2)-induced suppression of contact hypersensitivity (CHS) response to contact sensitizer dinitrofluorobenzene. Prevention of UVB-induced suppression of CHS by silymarin was found to be associated with the inhibition of infiltrating leukocytes, particularly CD11b+ cell type, and myeloperoxidase activity (50-71%). Silymarin treatment also resulted in significant reduction of UVB-induced immunosuppressive cytokine interleukin-10 producing cells and its production (58-72%, p<0.001). Topical treatment of silymarin also resulted in significant reduction of the number of UVB-induced H2O2 producing cells and inducible nitric oxide synthase expressing cells concomitant with decrease in H2O2 (58-65%, p<0.001) and nitric oxide (65-68%, p<0.001) production. Together, these data suggest that prevention of UVB-induced immuno-suppression and oxidative stress by silymarin may be associated with the prevention of photocarcinogenesis in mice. The data obtained from this study also suggest: i) phase-I clinical trial of silymarin in high skin cancer risk human population and ii) development of sunscreen containing silymarin as an antioxidant (chemopreventive agent) or silymarin can be supplemented in skin care products.

Anti-allergic effects of cnidii monnieri fructus (dried fruits of Cnidium monnieri) and its major component, osthol.

Anti-allergic effects (types I and IV) of the 70% ethanol extract (CM-ext) obtained from Cnidii Monnieri Fructus (dried fruits of Cnidium monnieri) were investigated on 48 h homologous passive cutaneous anaphylaxis (PCA), 2, 4-dinitrofluorobenzene (DNFB)-induced contact dermatitis and picryl chloride (PC)-induced contact dermatitis in experimental animals. CM-ext showed inhibitory effects on these allergic models. Osthol isolated from CM-ext also had the inhibitory effects. These results suggested that Cnidii Monnieri Fructus might be useful as an agent for allergic diseases and that its anti-allergic effect was partially attributable to a coumarin derivative, osthol.

Low-dose dithranol treatment and tape stripping induce tolerance to dithranol in a mouse ear oedema model.

BACKGROUND: It is well known from clinical practice that repeated treatment with dithranol leads to the development of tolerance. OBJECTIVES: To investigate the characteristics and mechanism of such dithranol tolerance. METHODS: The mouse ear was pretreated with a low dose of dithranol or croton oil or, in previously sensitized animals, with dinitrofluorobenzene (DNFB). Twenty-four hours later irritant dermatitis was elicited by painting the mouse ear with a high dose of dithranol, croton oil or DNFB, and the dermatitis was characterized by measurement of ear thickness. RESULTS: Low-dose dithranol significantly suppressed dithranol-induced oedema, whereas it had no effect on croton oil- or DNFB-induced dermatitis, suggesting that dithranol-induced tolerance is specific. Tolerance to dithranol could not be induced by pretreatment of the mouse ear with a low dose of croton oil or DNFB. Mild tape stripping of the mouse ear also inhibited the inflammatory effect of dithranol applied 24 h later. Superoxide dismutase treatment abolished the tolerance-inducing effect of low-dose dithranol or stripping. CONCLUSIONS: These results suggest that superoxide anion radicals are involved not only in the inflammatory effect of dithranol, but also in the induction of tolerance.

The effect of VAS972 on allergic contact hypersensitivity.

BACKGROUND: Contact hypersensitivity (CHS) is a Th1-mediated immune response that can be down-regulated by immunosuppressive agents such as cyclosporine and environmental stimuli such as ultraviolet light. Recently, an immunomodulation therapy, VAS972, has been developed which is believed to down-regulate the Th1 arm of the immune response. This VAS972 involves modifying autologous blood by controlled exposure to the oxidizing agent ozone and UVC light, at an elevated temperature ex vivo. The processed blood is then administered by intramuscular injection. OBJECTIVE: To further evaluate the immune modulating effect of VAS972. METHODS: We examined the effect of VAS972 treatment on CHS. Contact hypersensitivity was induced with dinitrofluorobenzene (DNFB) in animals receiving VAS972- processed blood, control blood, or saline. A preliminary study was also conducted to evaluate the effect of plasma and cellular fractions of processed blood. RESULTS: Mice injected with VAS972-processed blood demonstrated a significantly lower (46%) CHS response than controls. Histologic examination of challenged ear skin from control mice displayed edema with a significant lymphocytic infiltration, whereas animals administered processed blood demonstrated a reduction in lymphocytic infiltration. Mice injected with either plasma or the cellular fraction of the VAS972-treated blood also demonstrated a significant suppression (49% and 41%, respectively). CONCLUSION: The results of this study demonstrated that VAS972 suppresses CHS and cellular infiltration. Furthermore, the plasma and cellular components of the VAS972 treatment were also able to induce immunosuppression. This further supports the hypothesis that VAS972 down-regulates the Th1 arm of the immune response.

The noninvasive mouse ear swelling assay. I. Refinements for detecting weak contact sensitizers.

The noninvasive mouse ear swelling assay (MESA) is a model for delayed-type hypersensitivity that holds promise as a testing protocol for allergic contact dermatitis (ACD). The MESA employs only topical sensitization on the abdomen and does not use injections, adjuvants, anesthesia, occlusion, or disruption of the stratum corneum. Five days after induction, the ears are challenged topically and ear swelling measurements taken at 24, 48, and 72 hr indicate the extent of ACD. In this study, refinements of the assay were explored in BALB/cBy mice using dinitrofluorobenzene (DNFB) and dinitrochlorobenzene (DNCB). A complete dose-response curve was developed for DNFB and the dose which sensitized half the mice in a group (SD50, 0.001%, w/v) was used to test noninvasive enhancement protocols. Several triple-dose protocols tested produced no increase in responsiveness and daily dosing showed a trend toward tolerance induction yielding 20% positive responses. Dietary vitamin A supplementation produced a dramatic enhancement of the responses: ear thickness increase was doubled and the SD50 sensitized 94 to 100% of the mice in the vitamin A groups. We conclude that the MESA allowed identification of ACD potency for known sensitizers at very low concentrations which do not produce ACD with other techniques. The importance of dose-response studies for avoiding the high-dose reduced-response region was also shown. Based on the observation that the vitamin A-augmented MESA was considerably more sensitive than with regular feed, a companion study (P.S. Thorne. C. Hawk, S.D. Kaliszewski, P.D. Guiney, Fundam. Appl. Tox. 17, 807-820, 1991) presents tests of the enhancements to the MESA developed in this work, using weak sensitizers and complex mixtures.

Expression of Ia antigen on epidermal keratinocytes is a consequence of cellular immunity.

Ia antigen is seen in the normal epidermis only in Langerhans cells. However, when there is damage to the epidermis induced by cellular immunity Ia is expressed in the keratinocytes. This phenomenon is not seen in trauma or chemical inflammation. It is suggested that the expression of Ia in keratinocytes is due to cellular immunity possibly due to a lymphokine.