Study On The Anti-inflammatory And Antipruritic Effects And Molecular Mechanism Of Two Drugs

Itching was originally defined by the German doctor Samuel Hafenreffer as "an unpleasant sensation that triggers the idea of scratching in order to eliminate harmful irritants." The molecular mechanism of itching is complex,and the causes of itching are varied.It is generally believed that the cells related to pruritus mainly include keratinocytes and immune cells,which release various metabolites to produce allergic pruritus and cause various inflammatory skin diseases.In the cycle of "pruritus-scratching-itch",the pruritus and the inflammation are aggravated,and the normal life of the patient is seriously affected.At present,the drugs used to treat chronic itching are simply divided into topical and systemic treatments,including emollients and moisturizers,anesthetics,cooling agents,and glucocorticoids,as well as antihistamines,neuroactive drugs,antidepressants,and phototherapy and so on.However,long-term use of antibiotics,immunosuppressive agents and glucocorticoids is difficult to avoid the side effects of drugs,and the symptoms are prone to recurrence.There are a variety of traditional Chinese medicine methods in China,which can relieve symptoms,reduce side effects,and improve curative effects.Commonly used traditional Chinese medicines include liquorice,divaricate saposhniovia root,fineleaf schizonepeta root,angelica dahurica,coptis root,Sophora flavescens,common cnidium fruit,densefruit pittany root-bark,borneol,etc.In this study,we established IMQ-induced psoriasis model and SADBE-induced ACD model and took oxymatrine(OMT)and borneol as examples to explore the anti-inflammatory and antipruritic effects of traditional Chinese medicine and its molecular mechanism.Firstly,IMQ model was established to explore the antipruritic and anti-inflammatory effects and mechanisms of OMT.Animal behavior results showed that OMT has a significant antipruritic effect in the IMQ model.We detect the expression of four heat shock proteins(HSP)in skin tissues by Western blotting and RT-PCR technology,and visually detect the expression and localization of heat shock proteins by immunohistochemistry and immunofluorescence staining techniques.The results exhibited that OMT played an antipruritic effect on mice with psoriasis by reducing the expression of HSP90 and HSP60 in keratinocytes.The results of morphological experiments such as H&E staining and toluidine blue staining indicated that OMT could significantly improve the inflammatory phenotype of the skin of the IMQ model in mice.In addition,OMT could reduce the mRNA expression of psoriasis marker inflammatory factors in the skin tissues of psoriasis mice such as IL-17,IL-22,IL-23,TNF-α and IL-6.Moreover,it was found that OMT could reduce the protein expression of MAPK pathway signaling molecules in skin tissues.Furthermore,a cellular inflammation model was established through TNF-α and IFN-γ.After using OMT,the mRNA expression of inflammatory factors all showed a decline and the increasing protein expression of p-P38 and p-ERK were reversed.The MAPK inhibitor U0126 also showed the similar inhibitory effects.Moreover,silencing the expression of HSP90 and HSP60 in the cells by siRNA technology could reverse the anti-inflammatory effect of OMT in HaCaT cells.The above results indicated that OMT exerts anti-inflammatory effects by reducing the expression of HSP90 and HSP60 in keratinocytes,decreasing the production of inflammatory effects and inhibiting the activation of MAPK pathway.Then SADBE model was estabilished to explore the mechanism of borneol’s anti-inflammatory effect.Using RT-PCR and Elisa technology to detect the influence of borneol on inflammatory factors,the results showed that borneol could effectively inhibit the expression of inflammatory factors in ear tissues,and immunohistochemical results displayed that borneol could down-regulate the expression of neutrophils,dendritic cells and M1 macrophages.Compared to those of immunohistochemistry,the results of Flow cytometry detection showed the same trend as immunohistochemistry,which borneol can inhibit the expression of dendritic cells and macrophages in ear tissues.Then,using LPS established an inflammatory macrophages model in vitro.The results indicated that borneol could inhibit the activation of M1 macrophages by LPS,decrease the expression of IL-6 mRNA in primary macrophages,and down-regulate the expression of protein molecules in the MAPK pathway.As explained above,the anti-inflammatory effect of borneol may involve in down-regulating the expression of four immune cells and inflammatory factors,inhibiting the activation of M1 type macrophages,and suppressing the mRNA level of IL-6.In summary,this study proved that OMT and borneol have anti-inflammatory and antipruritic effects and explored their mechanism of action.By regulating the expression of HSP90 and HSP60 in keratinocytes,OMT inhibited the itching of psoriasis,suppressed the MAPK signaling pathway and down-regulated the expression of inflammatory factors to exert an anti-inflammatory effect.Borneol could decrease the expression of dendritic cells,neutrophils and macrophages,inhibit the activation of M1 macrophages and the polarization of IL-6 on M1 macrophages,and down-regulate the p-P38 and p-ERK protein in MAPK signal pathway to exert its anti-inflammatory effect.Through this study,the anti-inflammatory and antipruritic pharmacological effects of OMT and borneol and their potential molecular mechanisms have been further confirmed.The relevant research could provide a important reference for their clinical effects and has good clinical application prospects.