| The three major systems of nerve,endocrine and immune in the body are closely related,involving the communication of biological regulatory signals such as a variety of hormones,neurotransmitters and cytokines,forming a complex neuroendocrine-immune network(NEI)to maintain the homeostasis of the internal and external environment.The physiological processes of female animals are mainly regulated by hypothalamic-pituitary-ovarianaxis(HPOA),estrogen,as a key hormone in HPOA,plays an important role in regulating NEI homeostasis and maintaining body health.In order to control the number of animals and prevent the occurrence of specific diseases such as uterine accumulation,female animals raised in the family are usually sterilized.However,post-sterilization estrogen deficiency can lead to NEI dysfunction,resulting in a weakened immune response or imbalance in immune regulation.Soy isoflavone(SIF)is a natural active substance present in legumes,which can produce estrogen-like effects in the body,and has the functions of promoting animal growth,improving animal reproductive ability,and enhancing the body’s immunity.In order to explore whether SIF can regulate NEI and improve the immune function of female sterilized animals,this experiment gave different doses of SIF to sterilized rabbits,and explored the immunomodulatory effect of SIF by detecting serum immune indexes,blood routine indicators,calculating spleen index and observing changes in spleen tissue morphology.Through the results of immunohistochemical staining of different neurotransmitters in the spleen,the anatomical basis of SIF to achieve neuroimmune regulation of the spleen was explored.Finally,by sequencing the transcriptome of the celiac ganglia,the possible mechanism of SIF regulating immune function through sympathetic pathways was explored,which provided a theoretical basis for the application of SIF in production and clinical use to improve the immune function of sterilized animals.The main research contents and results are as follows:1.Effect of soy isoflavones on immune function of sterilized rabbits: The experiment was divided into 5 groups,namely sham surgery group(SHAM group),animal model group(MOD group),SIF low,medium and high dose groups(L-SIF group,M-SIF group,H-SIF group).All rabbits except the SHAM group were sterilized,recovered for 2 weeks after surgery,and the model of sterilized rabbits was successfully established by ELISA method to determine the level of peripheral blood estrogen,and then the corresponding drugs were given once a day for 14 days.(1)The results of serum ELISA showed that after sterilization,immunoglobulin Ig G in rabbit serum was significantly reduced(P < 0.05),pleiotropic cytokine IL-2 also showed a certain degree of decline,while the content of pro-inflammatory factors TNF-α and IFN-γ was significantly increased(P < 0.05)and very significantly increased(P < 0.01).After SIF,the content of Ig G in H-SIF group increased significantly(P < 0.05),the level of IL-2 in M-SIF and H-SIF groups increased significantly(P < 0.01),the serum TNF-α level in rabbits in the M-SIF and h-SIF groups decreased significantly(P < 0.05),and the serum IFN-γ levels in Rabbits in the H-SIF Group decreased significantly(P < 0.05).(2)The results of blood routine showed that the peripheral blood leukocytes of rabbits had a significant tendency to increase after sterilization,and the use of various doses of SIF could inhibit the increase of white blood cell number.(3)The results of spleen index showed that the spleen index of rabbits was significantly reduced after sterilization(P < 0.01),and after high-dose SIF administration,the spleen index was significantly increased(P < 0.05).(4)The results of spleen HE staining showed that after sterilization,the white pulp of the rabbit spleen was reduced,the cell arrangement was disordered,the density of red pulp tissue was reduced,and there was a large number of red blood cell infiltration,and all dose groups of SIF had obvious improvement effect on the spleen injury induced by sterilization.(4)The immunohistochemical results of spleen neurotransmitter receptor showed that the expression levels of β2-AR and NPY1 R in rabbits after sterilization were significantly reduced(P < 0.01),and the expression levels ofβ2-AR in the M-SIF and h-SIF groups were significantly increased after SIF administration(P < 0.01).The expression level of NPY1 R in the L-SIF group was significantly increased(P< 0.05),and the expression level of NPY1 R in the M-SIF and h-SIF groups was very significantly increased(P < 0.01),indicating that SIF could enhance the communication and interaction between the sympathetic nervous system and the immune system.2.Effect of soy isoflavones on the expression of neuronal m RNA in the abdominal ganglion of sterilized rabbits: the high-dose soy isoflavones(100 mg/kg)with the most significant effect in the above experiments were taken as the experimental group(SIF group),the conditions of the MOD group and the SHAM group remained unchanged,the celiac ganglion was taken,and m RNA was extracted for high-throughput sequencing.(1)The results of differential expression gene analysis showed that 2624 differentially expressed genes were detected in the HAM group compared with the MOD group;1179 differentially expressed genes were detected in the SIF group.There were 732 genes that were differentially expressed between the two comparison groups,and it was speculated that these genes with common differential expression were the targets of SIF regulation of the abdominal sympathetic ganglia in sterilized rabbits.(2)GO and KEGG enrichment analysis showed that these differential genes were mainly involved in biological processes such as inflammatory response,chemotaxis and innate immune response,and were mainly enriched in cytokines and their receptor interactions,intercellular adhesion factors and axon guidance and other signaling pathways.(3)The differential genes in these pathways were further compared with the target genes associated with the nervous system,and the results showed that SIF downregulated the expression of the neuroinflammation and nerve damage related genes PTPN22,CASP8,TLR3,STAT4,CD27,B2 M,PTPRC,CXCL10,CXCR4,and C4 A.The expression of axonal regeneration and axonal guide-related genes SERPINF1,FGF2,COL2A1,BMP4,NGEF and CHGA was upregulated.(4)19 differentially expressed genes were selected for verification by RT-q PCR,and the results showed that the sequencing results were consistent with the RTq PCR results,confirming the credibility of the sequencing data.In summary,SIF can inhibit inflammatory responses and regulate the immune function of sterilized rabbits;This effect may be mediated by SIF improving neural function and enhancing communication and interaction between the sympathetic nervous system and the immune system.In summary,SIF can inhibit the neuter-induced inflammatory response,maintain the homeostasis of the immune system of sterilized animals,and restore the spleen tissue damage caused by sterilization.This effect may be mediated by SIF improving neurological function and enhancing communication interactions between the sympathetic nervous system and the immune system. |
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