Epidermal growth factor-receptor tyrosine kinase activity regulates expansion of porcine oocyte-cumulus cell complexes in vitro.
We have recently shown that epidermal growth factor (EGF) strongly stimulates expansion of porcine oocyte-cumulus complexes (OCCs) isolated from large follicles (>6 mm) and does not promote expansion of OCCs from small (3-4-mm) follicles. In order to elucidate the role of EGF in OCCs expansion, in the present study, we first examined the presence of EGF receptors (EGFRs) in cumulus cells isolated from follicles of different sizes. Surprisingly, immunoblotting showed that cumulus cells obtained from all follicular size categories contained similar amounts of EGFR protein. On the other hand, we found a dramatic difference in the pattern of protein tyrosine phosphorylation in a comparison of cumulus cells isolated from small and large follicles treated by EGF. Furthermore, tyrosine-phosphorylated EGFR was specifically immunoprecipitated with antiphosphotyrosine antibodies from EGF-treated cumulus cells isolated from the large follicles. This result strongly indicates that only OCCs from the large follicles contain mature EGFRs that are capable of becoming activated by EGF. Remarkably, preincubation of cumulus cells from small follicles (3-4 mm) with FSH strongly increased EGF-stimulated tyrosine phosphorylation to levels comparable with OCCs from large follicles. The FSH-dependent activation of EGFRs was beneficial for expansion of OCCs isolated from the small follicles since OCCs treated sequentially by FSH (3 h) and EGF (1 h) underwent expansion significantly better then OCCs cultured in FSH or EGF alone. We conclude that a FSH-dependent pathway has an important role in the maturation of the EGFR in cumulus cells and that activation of EGFR-dependent signaling is sufficient to induce expansion.
Prochazka
Radek
R
Institute of Animal Physiology and Genetics, Academy of Sciences of the Czech Republic, 277 21 Libechov, Czech Republic. prochazka@iapg.cas.cz
Kalab
Petr
P
Nagyova
Eva
E
D000818:Animals; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000378:metabolism; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000378:metabolism*; D005260:Female; D005640:Follicle Stimulating Hormone / Q000378:metabolism; D006107:Granulosa Cells / Q000201:enzymology; D006820:Hyaluronic Acid / Q000378:metabolism; D015151:Immunoblotting / Q000662:veterinary; D009865:Oocytes / Q000201:enzymology* / Q000502:physiology; D006080:Ovarian Follicle / Q000201:enzymology* / Q000502:physiology; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D013552:Swine / Q000378:metabolism*; D020032:Tyrphostins / Q000494:pharmacology
D004791:Enzyme Inhibitors; D020032:Tyrphostins; C086599:tyrphostin A46; D004815:Epidermal Growth Factor; D005640:Follicle Stimulating Hormone; D006820:Hyaluronic Acid; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases