Effects Of Active Immunization Against GnRH On Immune Function Of Spleen In Male SD Rats

In animal husbandry production,castration of male animals can reduce the aggressiveness of the animals,increase the growth performance,and also eliminate the "taint" of meat,thus improving management efficiency and economic benefits in animal husbandry.Active immunization against GnRH(Gonadotrophin-releasing Hormone)is gradually replacing traditional surgical castration because of its advantages.such as it can not only improve the meat quality,but also decrease mortality and alleviate stress caused by surgical castration.Previous studies showed that there is interaction effects between the reproductive endocrine system and the immune system.Therefore,we speculate that the active immunization against GnRH may have an impact on the immune system of the animal.The spleen is the main immune organ of animals,It is rich in lymphocytes and macrophages.And it is the key to maintaining the cellular immunity and humoral immunity of animals.Active immunization against GnRH may affect the immune function of animals by regulating the content of GnRH and sex hormones in animals.But there are few related reports.Thus,this study used SD male rats as a model to explore the effect of active immunization against GnRH on animal spleen immune function.The study can Enrich the research content in this field and provide a more solid theoretical basis for the application of GnRH active immune castration technology in animal husbandry production.(1)Experiment one: 30 healthy male SD rats aged 4-5 weeks were used,and were divided into 3 groups randomly,10 animals for each group.The 3 groups were named Intact controls,GnRH-immunized and Surgical castrates.The Intact controls was not treated,and the Intact controls was injected with emulsified antigen(1ml,50 μg GnRH-TDK)into the leg muscle at 9 weeks of age.The Surgical castrates underwent surgical castration at 7 weeks of age.Blood samples were collected from the tail artery on the day of the initial immunized and recorded as 0wpv(weeks post vaccination).Thereafter,blood samples were collected every four weeks,3 times in total.The rats were slaughter at 14 wpv,the relevant tissues were taken and the relevant experimental data were recorded.Flow cytometry was used to detect the number of immune-related cells and T-lymphocyte grouping in the spleen of rats.The levels of anti-GnRH antibodies and Th1,Th2 related cytokines in the serum were detected by ELISA.The levels of testosterone and estradiol in the serum of male rats were analyzed by radioimmunoassay kit.The total RNA of the spleen was extracted,and after reverse transcription,the expression levels of GnRH receptor m RNA and immune-related genes were determined by fluorescence quantitative PCR.The results are as follows:Compared with the intact controls,the active immunization against GnRH male rats had atrophy of testicles and a significant reduction in testicular weight(P <0.05).Serum GnRH antibody titers increased significantly after the first injection and the booster injection(P <0.05).The levels of testosterone and estradiol in male rats were significantly reduced(P<0.05);there was no significant difference in spleen weight and spleen index among Intact controls,GnRH-immunized and Surgical castrates(P> 0.05).Compared with the Intact controls,the number of CD4 + lymphocytes and CD8 + lymphocytes in the spleen tissues of immunocastrated male rats significantly increased(P <0.05).There was no significant difference in the number of NK cells and B cells(P> 0.05).Compared with the Intact controls,the serum levels of IL-2 and IL-10 in GnRH-immunized and Surgical castrates male rats increased significantly(P <0.05),and the content of IL-6 significantly decreased(P<0.05).There was no significant difference in the levels of IL-1β,tumor necrosis factor,IFN-γ,and IL-4 in the serum of the Intact controls,the GnRH-immunized,and the Surgical castrates(P> 0.05).Compared with the Intact controls,the m RNA expression of IL-4 in the spleen tissues of the immune castrated male rats was significantly increased(P <0.05).The amount significantly increased(P <0.05).There were no significant differences in the m RNA expression levels of IL-1β,IFN-γ,and IL-6 in the spleen tissues among the Intact controls,the GnRH-immunized,and the Surgical castrates(P> 0.05).(2)Experiment two: 18 healthy male SD rats aged 4-5 weeks were randomly divided into 3 groups,6 animals for each group.The 6 group were named Intact controls+BSA,GnRH-immunized+BSA,Surgical castrates+BSA.In the GnRH immune castration group,emulsified antigen(1 ml,containing GnRH-TDK 50 μg)was injected intramuscularly at the age of 9 weeks in rats,and the same dose was immunized after 8 weeks.The surgical castration group were surgical castration at 7 weeks of age,while the control group did not get castration treatment.Record the day of the first injection as o wpv.At 12 wpv,All the groups of BSA-immunized rats were injected with emulsified antigen(1ml,50μg BSA).Blood samples were collected from the tail artery of rats at 18 wpv.The rats were slaughter at 20 wpv.The relevant tissues were taken and the relevant experimental data was recorded.The content of testosterone and estradiol in the serum of male rats was analyzed by radioimmunoassay.The content of anti-BSA antibody in serum was determined by ELISA.The results are as follows:Compared with the Intact controls,the active immunization against GnRH male rats had atrophy of testicles and a significant reduction in testicular weight(P <0.05).Serum GnRH antibody titers increased significantly after the first injection and the booster injection(P <0.05),The levels of testosterone and estradiol in male rats were significantly reduced(P<0.05)at slaughter,Compared with the control group,there was no significant change in the serum anti-BSA antibody level in the active immunization group of GnRH(P> 0.05).The serum anti-BSA antibody level in the two groups was basically the same after injection of BSA;The serum anti-BSA antibody level in the castration group was significantly increased(P <0.05).The above results showed that active immunization against GnRH in male SD rats can effectively inhibit the testes development,reduce the serum testosterone and estradiol content of male rats;significantly increase the number of total cells,helper T lymphocytes and Cytotoxic T lymphocytes in the spleen tissue of male rats.Serum concentration of IL-2and IL-10 are significantly increased;The expression of IL-2,IL-4 and IL-10 m RNA in the spleen tissues of male rats are significantly increased.In summary,active immunization against GnRH significantly increased the proportion of T cells in spleen tissue,significantly increased the expression of IL-2,IL-4,IL-10 m RNA in spleen tissue and the concentration of IL-2,IL-10 in serum.This may be caused by active immunization against GnRH reducing testosterone androgen synthesis in male rats.