The Role Of Mouse Mrgprb2 Gene In Allergic Reactions Induced By Traditional Chinese Medicine Injection

Traditional Chinese medicine(TCM)was a national treasure of China.With the development of science and technology,the modernization of TCM combining the characteristics of traditional TCM and modern technology was the general trend of the development of TCM,among which TCM injection was one of the typical representatives.Reports of adverse reactions caused by Traditional Chinese medicine injections(TCMI)have increased in recent years,largely curbing their development.Studies have shown that Chinese medicine injections mainly induce anaphylaxis,which can occur at the first dose without IgE mediated reaction.In this study,Mrgprb2 gene knockout mouse model was established,and DNA,gene sequencing and protein phenotype identification were performed to evaluate the anaphylactic reaction induced by Yuxingcao injection and positive drug C48/80.In order to determine the role of Mrgprb2 gene in anaphylactoid reaction,we evaluated whether the inhibition of Mrgprb2 gene on Yuxingcao injection and C48/80 induced by Mrgprb2 gene knockout in mice was decreased by the release of serum hiamine,the release of ?-hexosaminidase in mouse peritoneal mast cells,pathological tissue sections and the degree of blue staining in the hind paw of mice.Finally,the pathway of anaphylaxis induced by TCMI was preliminarily studied by transcriptome sequencing technology to provide experimental basis for the study of the mechanism of anaphylaxis induced by TCMI.The experimental results are as follows:1.Construction and identification of Mrgprb2 gene knockout mouse model.The Mrgprb2 gene knockout mouse model was established in cooperation with Jiangsu Jicuiyaokang Biological Science and Technology Co.,Ltd.The genotypes were identified by AGE.The homozygous were screened out and the characteristic amplified fragments were sequenced and verified.Western blot result revealed that MRGPRB2 was not found in peritoneal mast cells of Mrgprb2-null mice.Mrgprb2 KO mice and wild-type C57BL/6 mice showed no significant differences in appearance,behavior,reproductive capacity and organ index,suggesting that the establishment of Mrgprb2 knockout mouse model was successful.2.The mediating role of Mrgprb2 gene in anaphylaxis induced by TCMI.The degree of degranulation of mice was evaluated by detecting the release rate of ?-hexosaminidase in the mast cells of the peritoneal cavity and observing the morphology of the cells under transmission electron microscope.The results showed that compared with wild-type mice(WT),the ?-hexosaminidase release rate of Mrgprb2 knockout mice(KO)was significantly decreased.(P<0.05).The transmission electron microscopy results were consistent with the ?-hexosaminidase release results,indicating that the knockout of Mrgprb2 gene could significantly inhibit the degranulation of mast cells.In vivo results reflected that compared with WT mice,Mrgprb2 KO mice significantly reduced serum histamine release(P<0.001),auricle vasodilation,pulmonary inflammatory response,and Evans blue leakage in hind PAWS were also reduced.Mrgprb2 gene was considered to be the key gene in the occurrence of anaphylaxis,and the KO model can be used for the evaluation of anaphylaxis.3.Preliminary study on the pathway of anaphylaxis induced by TCM injections.Transcriptome sequencing was performed on liver tissue samples of WT and Mrgprb2 KO mice to explore the mechanism of anaphylaxis induced by TCMI.Results show that the induced by Yuxingcao injection and C48/80 was not the same kind of allergic reaction mechanism,C48/80 mainly through MAPK signal pathway,TNF signaling pathways,toll-like receptors signaling pathways triggered allergic reactions,Mrgprb2 knockout,PI3K-Akt signaling pathways as its cause histamine,beta aminocaproic increased release of glycosidic enzyme(compared with the blank group),while the anaphylaxis induced by Yuxingcao injection may be mediated by the inflammatory mediator regulatory pathways of arachidonic acid and TRP channels.No inflammatory,anaphylaxis/anaphylaxis related pathways were found after the knockout of Mrgprb2 gene.In conclusion,it can be concluded that Mrgprb2 gene plays a key role in anaphylaxis induced by TCMI.