Evaluation Of Castration Effect For A Novel GnRH Vaccine

Objective: To develop a novel GnRH vaccine and explore its castration effect on male SD rats.Methods:(1)GnRH-tandem(GnRH2)was synthesized on solid-phase peptide synthesizer,the purity and molecular weight was analysed by high performance liquid chromatography(HPLC)and mass spectrometry,respectively.Then the GnRH2 was modified with multiple antigen peptides(GnRH2-MAP)and detected its purity.(2)18 ~ 20 d male rats were randomly divided into three groups of the following treatments: GnRH2-MAP immunization(n=30),GnRH2 immunization(n=30)and control(n=30).The animals of GnRH2-MAP and GnRH2 were injected 100?g / vaccine,respectively.and 50% adjuvant saline solution,the booster injection was given at 2W intervals until immunion times were 3.(3)The samples were collected at 2W intervals after the first immunization,the serum was seperated,the body weight,testicular weight and size were recorded,and a small amount of testicular tissue was taken and fixed in 4% paraformaldehyde solution.(3)The levels of GnRH antibody and testosterone in the serum were measured by enzyme linked immunosorbent assay(ELISA).The testicular tissue was embedded with paraffin,hematoxylin-eosin(HE)staining,and observed and photographed under optical microscope.Results:(1)The purity of GnRH2-MAP was more than 95%.The body weight of immunized group showed no significance with the control group.The serum GnRH antibody in the immunized group was significantly higher than that of in the control group(P<0.05),and the testosterone concentration significantly decreased(P<0.05).The GnRH antibody of the GnRH2-MAP immunized animals significantly increased after the first immunization and was significantly higher than that of the GnRH2 immunized rats at several periods(P<0.05).No difference of testosterone concentration was found between GnRH2 and the control at several periods(P>0.05).The weight and sizes of the testes were significantly lower than those of the control(P<0.05).(3)Testicular tissue sections showed that the spermatogenic tubules in GnRH2-MAP treated animals had atrophy,the structure of the tube was loose,the number of cells was few and the spermatogonial differentiation was not obvious.The tubular lumen in GnRH2 immunized animals became large,a few cells and sperm were observed in the tube.Spermatogenic tubules were well developed in the control group.The perfect histological structure and cell layer were found.Spermatogonial cell differentiation were obvious,a large number of sperm were observed in the control.Conclusion:GnRH2-MAP with high purity is successful synthesed and the reproductive function is inhibited in GnRH2-MAP immunized rats.The effect of castration is satisfactory which suggests that GnRH2-MAP is a very effective immunogen.