ABSTRACT
OBJECTIVES@#Ozone is widely applied to treat allergic skin diseases such as eczema, atopic dermatitis, and contact dermatitis. However, the specific mechanism remains unclear. This study aims to investigate the effects of ozonated oil on treating 2,4-dinitrochlorobenzene (DNCB)-induced allergic contact dermatitis (ACD) and the underling mechanisms.@*METHODS@#Besides the blank control (Ctrl) group, all other mice were treated with DNCB to establish an ACD-like mouse model and were randomized into following groups: a model group, a basal oil group, an ozonated oil group, a FcεRI-overexpressed plasmid (FcεRI-OE) group, and a FcεRI empty plasmid (FcεRI-NC) group. The basal oil group and the ozonated oil group were treated with basal oil and ozonated oil, respectively. The FcεRI-OE group and the FcεRI-NC group were intradermally injected 25 µg FcεRI overexpression plasmid and 25 µg FcεRI empty plasmid when treating with ozonated oil, respectively. We recorded skin lesions daily and used reflectance confocal microscope (RCM) to evaluate thickness and inflammatory changes of skin lesions. Hematoxylin-eosin (HE) staining, real-time PCR, RNA-sequencing (RNA-seq), and immunohistochemistry were performed to detct and analyze the skin lesions.@*RESULTS@#Ozonated oil significantly alleviated DNCB-induced ACD-like dermatitis and reduced the expressions of IFN-γ, IL-17A, IL-1β, TNF-α, and other related inflammatory factors (all P<0.05). RNA-seq analysis revealed that ozonated oil significantly inhibited the activation of the DNCB-induced FcεRI/Syk signaling pathway, confirmed by real-time PCR and immunohistochemistry (all P<0.05). Compared with the ozonated oil group and the FcεRI-NC group, the mRNA expression levels of IFN-γ, IL-17A, IL-1β, IL-6, TNF-α, and other inflammatory genes in the FcεRI-OE group were significantly increased (all P<0.05), and the mRNA and protein expression levels of FcεRI and Syk were significantly elevated in the FcεRI-OE group as well (all P<0.05).@*CONCLUSIONS@#Ozonated oil significantly improves ACD-like dermatitis and alleviated DNCB-induced ACD-like dermatitis via inhibiting the FcεRI/Syk signaling pathway.
Subject(s)
Animals , Mice , Dinitrochlorobenzene/metabolism , Skin/metabolism , Cytokines/metabolism , Interleukin-17/metabolism , Tumor Necrosis Factor-alpha/metabolism , Dermatitis, Allergic Contact/pathology , Dermatitis, Atopic/chemically induced , Signal Transduction , RNA, Messenger/metabolism , Mice, Inbred BALB CABSTRACT
Objective: To explore the anti-inflammatory and antioxidant effects of caraway on atopic dermatitis (AD) in mice. Methods: AD was induced in two stages, including sensitization and challenge with the application of 2,4 dinitrochlorobenzene 2% and 0.2%, respectively. Clinical symptoms and histological analysis of the skin were assessed. The effects of caraway on oxidant/ antioxidant parameters as well as Th1- and Th2-related cytokines were also evaluated. Results: Caraway reduced the severity of dermatitis in AD-induced mice, as evidenced by significant inhibition of Th2-related cytokines (IL-4 and IL-13) and increased Th1-related cytokine (IFN-γ). Additionally, treatment with caraway significantly increased superoxide dismutase and catalase activity and decreased the malondialdehyde level in the serum of AD mice. Furthermore, caraway inhibited the differentiation of Th2 cells while favoring Th1 cell differentiation in the spleen via regulating their master transcription factors GATA3 and T-bet. Conclusions: Caraway could improve AD autoimmune responses and could be considered a potential candidate to treat AD disease.
ABSTRACT
ObjectiveTo explore the pharmacodynamic effect of gramine on 2，4-dinitrochlorobenzene （DNCB）-induced atopic dermatitis （AD） in mice and its potential mechanism. MethodThe mice were divided into the normal control group， model group， dexamethasone （0.05 g·kg-1） group， and high- and low-dose （0.12，0.06 g·kg-1） gramine groups. Mice in all groups except for the normal control group were stimulated with DNCB， followed by medication 13 d later. The changes in skin lesions were then observed， and the skin thickness， moisture content， and transepidermal water loss （TWEL） in each group were measured. The pathological changes in skin lesions were observed by hematoxylin-eosin （HE） staining， and the effects of drugs on CD4+/CD8+T-cell ratio in the spleen were detected by flow cytometry. The levels of immunoglobulin E （IgE）， interleukin （IL）-4， and IL-6 in serum were detected by enzyme-linked immunosorbent assay （ELISA）， and the changes in serum alanine aminotransferase （ALT）， aspartate aminotransferase （AST）， blood urea nitrogen （BUN）， and creatinine （CRE） by microplate method. The mRNA expression levels of inflammatory cytokines γ-interferon（IFN-γ）， IL-13， IL-17， IL-1β， IL-6， and tumor necrosis factor-α （TNF-α） in skin lesions were assayed by real-time polymerase chain reaction （Real-time PCR）， and the protein expression levels of nuclear transcription factor -κB (NF-κB) and NF-κB inhibitory protein α (IκBα) in skin lesions by Western blot. ResultCompared with the normal control group， the model group showed skin edema， erythema， scab， scratch， and lymphocyte and neutrophil infiltration， decreased skin moisture content， as well as increased skin thickness， TWEL （P<0.01）， spleen index， CD4+/CD8+ T-cell ratio in the spleen （P<0.05）， mRNA expression of IFN-γ， IL-13， IL-17， IL-1β， IL-6， and TNF-α in the skin lesions （P<0.05）， serum contents of IgE， IL-4， and IL-6 （P<0.05）， and protein expression of IκBα and NF-κB in skin lesions （P<0.05）. Compared with the model group， dexamethasone and gramine at different doses alleviated skin erythema， scale， scab， and inflammatory cell infiltration， elevated skin moisture content， inhibited skin thickening and TWEL， and decreased spleen index， CD4+/CD8+T-cell ratio in the spleen， mRNA expression of inflammatory factors in the skin lesions， serum contents of IgE and inflammatory factors， and protein expression of IκBα and NF-κB in skin lesions， especially in the dexamethasone group and the high- dose gramine group（P<0.05，P<0.01）. ConclusionGramine can inhibit the expression of related inflammatory factors and regulate the immune function of AD mice via the IκBα/NF-κB pathway， enabling it become a potential drug for treating AD.
ABSTRACT
Objective@#To investigate the developmental toxicity of 2,4-dinitrochlorobenzene in zebrafish embryos.@*Methods@#AB wild-type male and female zebrafish were selected to mate and spawn, then the eggs were cultured with Holt buffer solution. Six dose groups ( 0.4, 0.8, 1.0, 1.2, 1.6, 2.0 μg/mL ), a solvent control group and a cosolvent control group, were set up with 20 embryos each. Malformations and death of embryos were observed at 48, 72 and 96 hpf ( hours post fertilization ), the mortality and 50% lethal concentration ( LC50 ) were also calculated. @*Results@#At 48, 72 and 96 hpf, the LC50 of 2,4-dinitrochlorobenzene on zebrafish embryos were 1.668, 1.043 and 0.895 μg/mL, respectively, with a downward trend. After 72 hpf, when the concentration reached 2.0 μg/mL, all the zebrafish died. In the range of 0.4-2.0 μg/mL, the mortality of zebrafish at 48, 72 and 96 hpf increased with the increase of 2,4-dinitrochlorobenzene concentration ( all P<0.05 ); the malformation rate of zebrafish embryos at 48 hpf increased with the increase of 2,4-dinitrochlorobenzene concentration ( P<0.05 ). Zebrafish embryos exposed to 2,4-dinitrochlorobenzene led to yolk sac edema, pericardial edema and spinal curvature. @*Conclusion@#2,4-dinitrochlorobenzene can affect the development of zebrafish embryos, which will lead to lethal and teratogenic effects.
ABSTRACT
Objective: To induce the atopic dermatitis (AD) by 2, 4-dinitrobenzene (DNCB) in the BALB/c mice, and to explore the possible mechanism of DNCB as hapten to induce AD. Methods: A total of 12 BALB/c mice were randomly divided into control group (n = 6) and AD model group (n = 6). The dorsal skin of the mice in AD model group was sensitized by 1.0% DNCB at days 1, 4, and 7 and the back skin of the left ears were challenged by 0. 5% DNCB at days 14, 17, 19, 22, 24, 27, and 29. The mice in control group were given substrate solution with the same volume at the same time points. The inflammation scores of the skin tissue of the mice in two groups were evaluated, the appearance changes of the ear skin of the mice in two groups were observed, the pathomorphology of the skin tissue in left ear of the mice in two groups were observed by HE staining and toluidine blue staining, the epidermal thickness of the skin tissue at lesion site of the mice in two groups was measured, and the number of mast cells in the skin tissue at lesion site of the mice in two groups was counted; the expression levels of interleukin-4 (IL-4) in the skin tissue at lesion site of the mice in two groups was detected by immunohistochemistry staining, and the levels of serum IgE was detected by ELISA method. Results: The inflammation score of the mice in AD model group was higher that in control group (P < 0. 01). The skin tissue in left ear of the mice in AD model presented as redness swelling and hardening accompaning with abnormal scratching marks; the epidermal thickness, the number of mast cells, the expression level of IL-4 in skin tissue at lesion site and the level of serum IgE of the mice in AD model group were all increased compared with control group (P < 0. 05 or P < 0. 01). Conclusion: The AD mouse can be successfully induced by 1. 0% DNCB for sensitizing and 0. 5% DNCB for chllenging in the BALB/c mice, and its mechanism may be related with the increasing of IL-4 expression level in skin tissue and IgE level in serum.
ABSTRACT
OBJECTIVE@#To observe the therapeutic effect of different doses of dihydroartemisinin (DHA) on atopic dermatitis (AD) in mice and explore the mechanism.@*METHODS@#Forty-two C57BL/6 mice were randomly divided into 7 groups (@*RESULTS@#Treatment with 25, 75, and 125 mg/kg DHA and dexamethasone all alleviated AD symptoms of mice, reduced the severity scores of skin lesions, and ameliorated pathological changes of the skin tissue. DHA at 125 mg/kg produced the most obvious therapeutic effect and significantly alleviated mast cell infiltration in the lesions as compared with the other treatment groups (@*CONCLUSIONS@#DHA is effective for the treatment of AD in mice with an optimal dose of 125 mg/kg. The therapeutic effect of DHA is achieved probably through regulation of local immunity by inhibiting mast cell infiltration in the lesions.
Subject(s)
Animals , Mice , Anti-Inflammatory Agents/therapeutic use , Artemisinins , Cytokines , Dermatitis, Atopic/drug therapy , Immunoglobulin E , Mast Cells , Mice, Inbred BALB C , Mice, Inbred C57BL , SkinABSTRACT
Objective Chronic dermatitis-eczema is a kind of inflammatory skin disease characterized by pruritic and eczematous skin lesions. In the present study, we investigated the anti-allergic effect of Saxifrage cream on eczema-like skin lesions induced by 2,4-dinitrochlorobenzene (DNCB) in mice. Methods KM mice were randomly divided into eight groups: normal group (Norm), model group (DNCB), positive group 1 (Dexamethasone cream, DEX), positive group 2 (Chinese herbal medicine Paeonol cream, PA), Saxifrage cream groups (equivalent to crude herb 1.25, 2.50, 5.00 g/g, S1, S2, S4), and negative group (Norm + S4). Except for the normal control group and negative control group, right ears of mice in other groups were repeatedly induced with 0.5% DNCB for four times for 3 d and previously sensitized with 7% DNCB to induce eczema. Then mice in different groups were treated by DEX, PA, S1, S2, and S4, respectively. After mice were sacrificed, weight difference of two ears, immune organ change, and pathological change of right ear were used to assess the effects of the drugs. Results Compared with model group, DEX, PA, S2, and S4 could significantly lower mice ear thickness and swelling degree, and improve eczema skin symptoms. Meanwhile, the index of thymus and spleen in treatment groups were significantly reduced (P < 0.05). What's more, the histological analysis demonstrated that thickening of the skin lesions were significantly reduced in DEX, S2, and S4-treated group, and the levels of IL-4 and IL-6 in serum could be reduced by DEX, S2, and S4. Conclusion Saxifrage cream has a protective function against DNCB-induced eczematous. The mechanism might be related to alleviate eczematous inflammatory reaction.
ABSTRACT
Gastrodia elata Blume is a well-known kind of natural products used as a folk medicine for thousands of years. However, anti-atopic dermatitis-like effects of G. elata Blume had not been evaluated until now. The aim of the present study is to investigate the protective effects of water extract from the roots of G. elata Blume (GE) on 2, 4-dinitrochlorobenzene-induced atopic dermatitis-like skin lesions using balb/c mice. Combination treatment of GE (at a dose of 12.5 mg/kg body weight by administrated per os + 0.5 mg/cm2 as ointment to apply on ear and dorsal skin) was significantly inhibited spleen weight, ear thickness, levels of serum immunoglobulin E and number of mast cells, compared with that of 2, 4-dinitrochlorobenzene-included groups without GE. Furthermore, combination application by oral administration plus by ointment of GE significantly inhibited the histamine release from rat peritoneal mast cells. These results suggest that combination treatment of oral administration plus ointment form of GE could be helpful as a potentially natural pharmaceutical treatment on atopic-like dermatitis.
Subject(s)
Animals , Mice , Rats , Administration, Oral , Biological Products , Body Weight , Dermatitis , Dermatitis, Atopic , Ear , Gastrodia , Histamine Release , Immunoglobulin E , Immunoglobulins , Mast Cells , Medicine, Traditional , Skin , Spleen , WaterABSTRACT
The various murine models have contributed to the study of human atopic dermatitis (AD). However limitations of the models involve low reproducibility and long time to develop AD. In an attempt to overcome these limitations and establish an atopic dermatitis murine model, we repeated the application of 2, 4-dinitrochlorobenzene (DNCB) patch in NC/Nga and BALB/c mice, which has advantages in reproduction and cost. For the sensitization, a 1 cm2 gauze-attached patch, where 1% or 0.2% DNCB was periodically attached on the back of NC/Nga and BALB/c mice. To estimate how homologous our model was with human atopic dermatitis, clinical, histological and immunological alterations were evaluated. Both strains showed severe atopic dermatitis, increase in subiliac lymph node weight, mast cells, epidermal hyperplasia and serum IgE levels. Though both exhibited a high IL-4/IFN-gamma and IL-4/TNF-beta ratio in the expression of mRNA, the shifting of DNCB-treated BALB/c mice was increased to more than double that of NC/Nga mice. These results suggest that our DNCB patched model using BALB/c mice were more suitable than NC/Nga mice in demonstrating the immune response. We anticipate that our novel model may be successfully used for pathogenesis of atopic dermatitis and assessment of therapeutic approaches.
Subject(s)
Animals , Humans , Mice , Dermatitis, Atopic , Dinitrochlorobenzene , Hyperplasia , Immunoglobulin E , Lymph Nodes , Mast Cells , Reproduction , RNA, MessengerABSTRACT
OBJECTIVE:To determine the content of hydroxyproline in liver tissue in rats with hepatic fibrosis.METHO_ DS:RP-HPLC was used with precolumn derivation by2,4-dinitrochlorobenzene(DNCB)in the Borax buffer solution for lh in boiling water.Acetonitrile-acetate buffer linear gradient elution was used.Detecting wavelength was360nm.RESULTS:Derivation of hydroxyproline could be separated from other amino acids and accurately determined.CONCLUSION:The method is rapid,accurate,and suitable for detection of biopsy samples obtained by liver puncture.