Studies On Recombinant Follicle Stimulating Hormone-Fc Fusion Protein Of Safety

Follicular Stimulating Hormone(FSH)is a kind of hormone which is synthesized and secreted by animal pituitary gland.It belongs to glycosylated protein hormone and plays an important role in regulating the development,growth,sexual maturation and reproduction of the body,so the follicle stimulating hormone,whether in livestock production or in the treatment of human infertility have important application value.In actual production and application,recombinant follicle-stimulating hormone has a higher activity,a more stable structure and a lower price.To evaluate the toxicity of Recombinant folliclein-Fc Fusion Protein,and study the safety of the drug and provide a scientific basis for further clinical use,acute toxicity test、repeated exposure test、mutagenicity assay and teratogenicity test were conducted.1.Acute toxicity testTo evaluate the safety of Recombinant follicle stimulating Hormone-Fc fusion protein,20 ICR rats,half of male and female,were intramuscular treated with Recombinant follicle stimula-ting Hormone-Fc fusion protein.The results show that Recombinant follicle stimulating Horm-one-Fc fusion protein through the mouth,LD50 greaters than 2g/kg.Accoring to WTO relevant about chemicals acute toxicity grading standards to evaluate,there is no significant toxicity in ICR rats treated with Recombinant follicle stimulating Hormone-Fc fusion protein.2.Repeated exposure test80 Wister rats were randomly divided into the experimental group and the control group,20 Rats in each group and inramuseular given at 1000 U/kg bw/d,500 U/Kg bw/d and 50U/kg bw/d three dose groups;another set of saline control group.Once a day for 14 days to test the safety and if the drug has toxic or side effects.The results showed that subacute toxicity test were not significantly different from the control group.The drug had no poisonous effects on normall activities,physioloical,pathologica and biochemical index of rats after 14 days,the test suggested that Recombinant follicle stimulating Hormone-Fc fusion protein is no genotoxicity.3.Mutagenicity Assay3.1 Sperm deformity testSelect 60 KunMing male and female mice on the base of preliminary experiment,the selected mice were divided into five groups trabantly,there are 12 mice in each group.The male and female mice were exposed to Recombinant folliclein-Fc Fusion Protein through intramuscular use at a dose of 375 U/kg bw,187.5 U/kg bw,93.75 U/kg bw,there are also cytooxan positive control group and negative control group.The groups were given Recombinant folliclein-Fc Fusion Protein seven times for continually.Then killed all mice on the 35th day after the first exposure,and the rates of sperm deformity were detected.The result of each dose group were no significant difference as compared with control group.Recombinant Folliclein-Fc Fusion Protein can not cause sperm deformity.3.2 Micronucleus TestSelect 50 KunMing male and female mice on the base of preliminary experiment,the selected mice were divided into five groups trabantly,there are 10 mice in each group.The KunMing male and female mice were exposed to Recombinant folliclein-Fc Fusion Protein through intramuscular use at a dose of 375 U/kg bw,187.5 U/kg bw,93.75 U/kg bw,there are also Cytoxan positive control group and 0.9%N.S.negative control group.The mice were killed by cervical dislocation after 6h after intramuscular use,the number of micronucleus cell per 1000 polychromatic erythrocytes from stermum bone marrow was counted and the figures wee analysed.The result showed that micronucleus test was not significantly different from the control group.The micronucleus test is negative.3.3 Ames testThe experiment was conducted with a view to evaluate the potential mutageenicity of Re-combinant folliclein-Fc Fusion Protein.Suspension of echinacea herb was prepared for Ames test.In the flat-plate incorporation test of Salmonella typhimurium,the average number of spontaneous revertants did not increase significantly compared to that of spontaneous revertants.The Ames test was negative.It indicates that Recombinant folliclein-Fc Fusion Protein did not cause the increase of the back mutation in murine typhoid Salmonella and could not cause gene mutation.As a result,acute toxicity,repeated exposure and mutagenicity test showed that Recombinant follicle stimulating Hormone-Fc fusion protein was a low toxic and safe medicine.