Thymosin alpha-1 and FA-1 monoclonal antibody affect murine preimplantation embryo development by modulating protein phosphorylation.
The effects of thymosin alpha 1 (T alpha 1) and FA-1 monoclonal antibody (anti-FA-1 mAb) on murine preimplantation embryonic development were investigated by performing 2-cell embryo bioassay and by studying ova/embryos protein phosphorylation pattern (by 32P metabolic labeling and by in vitro kinase assay) and protein synthesis (by in vitro [35S]methionine labeling). T alpha 1 treatment (0.1, 0.5 and 5 ng/100 microliters) significantly increased blastulation rates (P < 0.01), blastocyst hatching rate (P < 0.0001), blastocyst diameter (P < 0.001) and number of cells per blastocyst (P < 0.0001) of the in vitro cultured 2-cell stage embryos. Anti-FA-1 mAb reduced blastulation rates (P < 0.001) primarily due to an arrest of development at morula stage. In vitro metabolic labeling of murine ova/embryos showed 32P incorporation into 4 major protein bands of murine ova (M(r) 125, 90, 68 and 31 kDa, respectively), 7 protein bands of 2-cell (M(r) 90, 68 and 31; and 145, 52, 38 and 32 kDa, respectively), 10 protein bands of morula (M(r) 150, 110, 92, 82, 70, 54, 39, 34, 30 and 29 kDa, respectively), and 15 protein bands of blastocyst (150, 110, 92, 70, 68, 54, 39, 34 and 30; and 131, 105, 52, 44, 43 and 33 kDa, respectively) stage embryos. T alpha 1 treatment (0.1-0.5 ng/100 microliters) resulted in a general increase in 32P labeling in all proteins of 2-cell, morula and blastocyst stage embryos. Anti-FA-1 mAb completely blocked 32P labeling of various proteins of murine ova, 2-cell, morula and blastocyst stage embryos, whereas control mouse myeloma IgG did not affect phosphorylation of these proteins. In vitro kinase assay performed directly on various ova/embryos extracts revealed 6 phosphoproteins (M(r) 105, 82, 55, 38, 34 and 33 kDa, respectively) that were common to ova and 2-cell embryos, besides a 43 kDa protein detected only in the ova extract. Of these phosphoproteins, T alpha 1 treatment specifically enhanced whereas anti-FA-1 mAb inhibited autophosphorylation of a 55 kDa protein of 2-cell embryos.(ABSTRACT TRUNCATED AT 250 WORDS)
Ahmad
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Department of Obstetrics and Gynecology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
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D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000941:Antigens / Q000502:physiology*; D001755:Blastocyst / Q000502:physiology*; D005314:Embryonic and Fetal Development / Q000187:drug effects; D005260:Female; D051379:Mice; D009924:Organ Culture Techniques; D010766:Phosphorylation; D011506:Proteins / Q000378:metabolism*; D000077596:Thymalfasin; D013947:Thymosin / Q000031:analogs & derivatives* / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D000941:Antigens; D011506:Proteins; C054151:fertilization antigen; D013947:Thymosin; D000077596:Thymalfasin