Regulation And Mechanism Of Combined Intervention Of L-theanine And EGCG On Intestinal Specific Immunity Of Ovalbumin Sensitized Mice

Specific immunity is the protective mechanism of the body against external antigens,and intestinal specific immune disorders caused by food antigens such as ovalbumin are closely related to body health.L-theanine and EGCG are the two main active components in green tea,both of which have many biological activities,such as anti-inflammation,antioxidation and regulating immunity,but there are few studies on the combination of L-theanine and L-theanine to regulate immunity,especially specific immune function.In this study,this study established a mouse model of ovalbumin sensitization,and intervened 6-week-old female BALB/c mice of SPF grade with L-theanine(100 mg·kg^-1·d^-1),EGCG(100 mg·kg^-1·d^-1),L-theanine+EGCG low,middle and high dose(50mg·kg^-1·d^-1,100mg·kg^-1·d^-1,150 mg·kg^-1·d^-1;the mass ratio of L-theanine to EGCG is 1:4)for 41days.The regulatory effect and mechanism of L-theanine and EGCG on intestinal specific immunity of ovalbumin-sensitized mice were studied and analyzed from the aspects of body weight,allergic reaction,intestinal morphology,serum specific antibody and allergy medium,intestinal cytokines,Peyer patch lymphocyte grouping,intestinal flora and expression of intestinal related proteins by enzyme-linked immunosorbent assay,flow cytometry,16Sr DNA and Westernblot.The main results and conclusions are as follows:（1）Establishment of ovalbumin sensitized mouse model.Female BALB/c mice were sensitized with 50μg of OVA and challenged for 4 times.The allergic score and OVA-IgE index of the mice could meet the requirements of the sensitization standard,and the ovalbumin sensitized mouse model was successfully established.（2）Regulation of intestinal specific immunity.Compared with OVA group,L-theanine,EGCG and L-theanine+EGCG could significantly inhibit the production of serum IgE and OVA-specific antibodies,significantly reduce the release of allergic mediators HIS and m MCP-1（P<0.05）,and alleviate the occurrence of allergic symptoms.The combination of middle and high doses of L-theanine and EGCG significantly inhibited the increase of immune organ index induced by OVA stimulation（P<0.05）.At the same time,L-theanine,EGCG and L-theanine+EGCG could combined intervention can also inhibit the differentiation of intestinal Th17 cells,DC cells（MHC-Ⅱ⁺CD11c⁺）and MC cells（FcεRⅠα⁺c-KIT⁺）,and reduce the contents of Th2 cytokines IL-4,IL-5 and IL-13.It promoted Th1 cells to secrete IFN-γand Treg cells to secrete IL-10 and TGF-β1（P<0.05）,thus maintain the balance of Th1/Th2 and inhibiting the occurrence of overimmunity.In addition,L-theanine,EGCG and L-theanine+EGCG could inhibit the decrease of intestinal microflora diversity caused by OVA sensitization,increase the abundance of Lactobacillaceae and Lactobacillus,and decrease the abundance of harmful Helicobacteraceae and Helicobacter（P<0.05）.In summary,the combined intervention of middle and high doses of L-theanine and EGCG in regulating intestinal specific immunity was better than EGCG or L-theanine alone.（3）Mechanism of intestinal specific immune regulation.Compared with OVA group,L-thenine,EGCG and L-theanine combined with EGCG could significantly down-regulated Lyn and Syk phosphorylation expression of key upstream protein of the allergy-related FcεRⅠsignal pathway,and prevent Lyn and Syk from initiating PI3K/AKT signaling pathway.Significantly down-regulated the phosphorylation expression of PI3K and AKT signaling molecules（P<0.05）,inhibited the activation of FcεRⅠsignal pathway,inhibited the activation of mast cells,enhanced the function of Treg immune cells,inhibited the development of Th2 immunity,and maintained Th1/Th2 balance and intestinal homeostasis.Regulate the intestinal specific immune function,so as to inhibit the occurrence of allergic reactions.