| Objective:To investigate the effectiveness of a herbal combination of Sophora flavescens and Angelica sinensis(SG)in treating eczema and the underlying mechanism involving the TLR4/My D88/NF-κ B signaling pathway.This study aims to provide theoretical and experimental evidence for the potential use of SG in the treatment of acute eczema.Methods: Sixty male BALB/c mice were divided into a blank group(n=10)and a model group(n=50)to manufacture the acute eczema model using2Magne4-dinitrochlorobenzene(DNCB).The day prior to the formal experiment,the abdominal hair of six groups of mice was trimmed short.The blank and model groups were shaved with a depilatory cream at the site.On the first day of the experiment,a 5% DNCB acetone solution was evenly coated on the abdomen of the model mice,and then strengthened once on the third day.On the sixth day,the mice in both groups were shaved with a 3.0cm × 3.0cm dimension.On the seventh day,the back skin of the model group was smeared with 1 DNCB acetone solution and 100 μL of normal saline,once every other day,for a total of six times.After the mice in the model group were challenged,swelling,telangiectasia,and exudate were observed with the naked eye to confirm whether the acute eczema animal model was successful.The model was made uniformly on the ipsilateral ear(left).After successful modeling,model mice were randomly assigned to five groups: SG low-dose,SG medium-dose,SG high-dose,model,and positive drug(prednisone)groups.IL-1β and TNF-α in serum were detected by ELISA.Skin histopathological changes were observed by HE staining,and TNF-α in skin tissue was semi-quantitatively detected by immunohistochemical technique.The content of TLR4 and NF-κB,protein expression of TLR4,NF-κB,and My D88 were detected by Western blot,and the gene expression of TLR4,NF-κB,and My D88 were detected by q RT-PCR to explore the mechanism of SG in treating acute eczema through the TLR4/My D88/NF-κB pathway.Results:1.Establishment of eczema model: Erythema,squamous epidermis,scabs,and papules were visibly present on the back skin of the model mice.The Eczema Area and Severity Index(EASI)score significantly increased,and the mice showed noticeable symptoms of itching.The back skin of the model mice exhibited erythema,squamous epidermis,papules,and pruritus.Post-treatment with SG led to a significant decrease in EASI score,alleviation of itching,and an improvement in the skin tissue condition of the mice.These results suggest that SG may offer promising therapeutic benefits for eczema.Post-treatment analysis revealed that SG is effective in significantly reducing the EASI score,relieving itching,and improving the skin tissue condition in mice,thus indicating that SG may present a superior therapeutic effect on eczema.2.The spleen and liver indexes indicated that the model group had higher indexes compared to the blank group(P < 0.01).Following the administration of SG and prednisone control group,each group showed a decrease in spleen and thymus indexes compared to the model group(P <0.01).This suggests that the immune balance improved and showed better therapeutic effects.3.The ELISA results revealed that the TNF-α and IL-1 βconcentrations were elevated in all groups compared to the blank group.Additionally,the TNF-α and IL-1β concentrations were higher in the model and SG groups compared to the blank group.Compared to the model group,there was a significant decrease(P<0.05 or P<0.01)in TNF-α and IL-1β concentrations in the SG and positive drug groups.4.The HE and immunohistochemistry outcomes revealed that the hyperkeratosis,edema and inflammatory cell infiltration in the positive drug group,high,middle and low dose SG groups were lower compared to that of the model group,while the most significant inhibitory effect on acute eczema was observed in the positive drug group and high dose SG group.The immunohistochemical technique indicated that the TNF-α,NF-κB-p65 and TLR4 levels in the skin tissue of the model group were significantly higher than those of the blank group(P < 0.01).The content of TNF-α,NF-κB-p65 and TLR4 in SG groups increased(P < 0.05 or P > 0.05).In comparison with the model group,the content of TNF-α and NF-κB-p65 in the low,middle,high dose and positive drug groups of SG decreased significantly,while that of TLR4 reduced significantly in the high dose and positive drug groups.5.The q RT-PCR results indicated significantly higher relative expression levels of My D88,NF-κB-p65,and TLR4 m RNA in the model group as compared to the blank group,with each SG group showing significantly higher TLR4 m RNA level compared to the blank group(P < 0.01 or P < 0.05).The levels of My D88,NF-κB-p65,and TLR4 m RNA were lower in each group than the model group.The My D88 m RNA levels were significantly lower in SG low dose group and positive drug group,compared to the model group.The NF-κ B-p65 m RNA level was decreased in different degrees in SG low dose group,high dose group,and positive drug group.The TLR4 m RNA level was significantly lower in the low dose SG group than that in the model group(P < 0.05).The level of TLR4 m RNA significantly decreased in middle dose group,high dose group,and positive drug group(P < 0.05).The Western blot results showed a significant increase in My D88,NF-κB-p65,and TLR4 expression in the model group as compared to the blank group.The expression of My D88 in each SG group,as well as the expression of NF-κB-p65 and TLR4 in the SG group also increased(P < 0.05).As compared to the model group,My D88 in SG high dose group and positive drug group showed a significant decrease,while NF-κ B-p65 and TLR4 showed a significant decrease in SG low,middle,high,and positive drug groups with statistically significant differences.Conclusion: SG has the ability to inhibit the production of the TLR4 inflammatory receptor,decrease the production of My D88 ligands in cells,lower the production of IL-1β and TNF-α in serum,and ultimately reduce the production of NF-κB.This suggests that SG’s therapeutic impact on eczema could be achieved through the inhibition of the TLR4/My D88/NF-κB signaling pathway. |
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