Effect And Mechanism Of Berberine On Mast Cell Mediated Allergic Diseases

Anaphylaxis is defined as a serious allergic hypersensitivity reaction that is rapid in onset and can be life-threatening and even fatal,and it is rapidly increasing worldwide.Mast cell is one kind of key effector cell in IgE-mediated allergic responses.Type I hypersensitivity is caused by many environmental factors including certain foods,dust,mites and pollen,which evoke the production of antigen-specific IgE antibodies that bind to high-affinity IgE receptors(Fc?RI)on the surface of mast cells.After IgE binds to the Fc?RI,mast cells can be activated by crosslinking of the Fc?RIs with multivalent antigens,resulting in the degranulation response and then happen of the allergic diseases.Objective:The anti-allergic effect of berberine was evaluated in vitro and in vivo.The effects of berberine in IgE stimulated RBL-2H3 cells and related mechanisms were investigated in vitro.The anti-allergic effect of berberine on histamine-induced vascular permeability in rats or mice and IgE medicated passive cutaneous anaphylaxis in mice were investigated in vivo.Methods:In vitro experiments:1.To confirm the effect of berberine,coptisine,palmatine,jatrorrhizine on the cell viability was checked by Thiazolyl blue tetrazolium bromide(MTT)in IgE stimulated RBL-2H3 cells.2.Effect of berberine,coptisine,palmatine,and jatrorrhizine on ?-HEX release in IgE stimulated RBL-2H3 cells.3.Effect of berberine in IgE stimulated RBL-2H3 cells degranulation.(1)Effect of berberine on the levels of ?-HEX,histamine,IL-4,and TNF-?release in IgE stimulated RBL-2H3 cells.(2)Using the toluidine blue dye to observe the effect of berberine on toluidine blue staining in RBL-2H3 cells.(3)Using the fluorescent dye iFluor488 phalloidin to stain the membrane structure F-actin and observe the membrane structure changes.(4)Effect of berberine on related proteins of the Src family,p-Lyn/Lyn,p-Syk/Syk,p-Gab2/Gab2,and related proteins of the MAPK family,p-JNK/JNK,p-ERK/ERK,p-P38/P38 expression levels.In vivo experiment:1.Effect of berberine on histamine-induced vascular permeability in rats or mice.After intraperitoneal injection of histamine phosphate for 30 min,the content of Evans blue dye in the skin of the back of the rats or mice was determined.2.Effect of berberine on IgE medicated passive cutaneous anaphylaxis in mice.After 30 min of intradermal injection of DNP-HSA,the content of Evans blue dye in the ear skin of the mice was determined.Results:In vitro experiments:1.The MTT results showed that the cell viability of RBL-2H3 cells was not affected by berberine,ketotifen,coptisine,palmatine,jatrorrhizine at 30 ?M.2.Berberine,coptisine,palmatine and jatrorrhizine can inhibit mast cell degranulation,and the levels of ?-HEX after pretreatment with berberine were significantly lower than pretreatment with coptisine,palmatine or jatrorrhizine(P <0.05,P < 0.01,P < 0.001).3.Effect of berberine in IgE stimulated RBL-2H3 cells degranulation.(1)Effects of berberine on ?-HEX,histamine,IL-4 and TNF-? levels.Berberine significantly inhibited the levels of ?-HEX,histamine,IL-4 and TNF-?in IgE stimulated RBL-2H3 cells(P < 0.05,P < 0.01,P < 0.001).(2)Effect of berberine on toluidine blue staining in RBL-2H3 cells.Pretreatment with berberine restored elongated shape and intracytoplasmic granule content increase in IgE sensitized RBL-2H3 cells.(3)Effect of berberine on F-actin rearrangement in RBL-2H3 cells.Berberine inhibits DNP-IgE/HSA-induced reorganization of F-actin in RBL-2H3 cells.(4)Effect of berberine on related proteins of the Src family,p-Lyn/Lyn,p-Syk/Syk,p-Gab2/Gab2,and related proteins of the MAPK family,p-JNK/JNK,p-ERK/ERK,p-P38/P38 expression levels.Pretreatment with berberine suppressed the phosphorylation of antigen-induced Lyn,Syk,and Gab2,thus suppressing the downstream MAPK pathways,p-JNK and p-ERK,p-P38(P < 0.05,P < 0.01)In vivo experiment:1.Effect of berberine on histamine-induced vascular permeability in rats or mice.Berberine significantly inhibits the content of Evans blue in the skin on histamine-induced vascular permeability in rats or mice(P < 0.001).2.Effect of berberine on IgE medicated passive cutaneous anaphylaxis in mice.Berberine significantly suppressed passive cutaneous anaphylaxis as observed by impaired extravasation of Evans blue dye in the ear skin(P < 0.05,P < 0.01).Conclusion:Berberine may be suppress mast cell degranulation via regulating Fc?RI-mediated and MAPK signaling in vitro.Orally administration with berberine alleviated increased vascular permeability caused by histamine in rats or mice and IgE medicated passive cutaneous anaphylaxis(PCA)in mice.