Stimulation of epidermal growth factor gene expression during the fetal mouse reproductive tract differentiation: role of androgen and its receptor.
We have shown previously that epidermal growth factor (EGF) plays a role in testosterone-dependent fetal Wolffian duct differentiation. To further assess the role for EGF, we determined whether EGF gene expression was modulated in response to male reproductive tract differentiation. The expression of EGF messenger RNA (mRNA) was measured by an RT-PCR assay using primer pairs spanning the coding sequence of 3228 nucleotide (nt) to 3401 nt. Using the RT-PCR reaction, an amplimer of the expected size, 173 bp, was detected in the fetal male reproductive tract. The amplimer hybridized with a radioactive probe representing an internal sequence of the amplified product and was digested by the restriction enzyme HaeIII, which has an unique cleavage site at 3365 nt. The level of EGF mRNA at different stages of sexual differentiation was measured by a newly developed quantitative competitive RNA PCR (QCPCR) assay for EGF mRNA. The assay was sensitive and reproducible within a linear range of amplification with 5 x 10(4) to 140 x 10(4) copies of mRNA. Using the quantitative competitive RNA PCR we found that the level of EGF mRNA was higher in the male reproductive tract than that in the female reproductive tract. Exposure to testosterone (40 mg/kg.day) during days 13-17 of gestation induced the Wolffian duct in the female fetuses and resulted in stimulation of EGF-mRNA expression. Similarly, an antiandrogen receptor, flutamide (100 mg/kg.day) exposure during days 13-17 of gestation inhibited male reproductive tract differentiation and resulted in inhibition of EGF-mRNA expression. Moreover, during the differentiation of the male reproductive tract there was a biphasic increase in the level of EGF-mRNA, first at day 14 of gestation, the period of onset of testicular activity and Wolffian duct morphogenesis, and second at day 18 of gestation, corresponding to onset of differentiation of the urogenital sinus, epididymal duct, and seminal vesicle. Thus, it appears that testosterone-induced male sexual differentiation is accompanied by an increase in EGF gene expression.
Gupta
C
C
Department of Pediatric Endocrinology, Children's Hospital of Pittsburgh, Rangos Research Center, Pennsylvania 15213, USA.
Singh
M
M
D000728:Androgens / Q000502:physiology*; D000818:Animals; D001483:Base Sequence; D005314:Embryonic and Fetal Development; D004815:Epidermal Growth Factor / Q000235:genetics*; D005260:Female; D005333:Fetus / Q000502:physiology*; D018507:Gene Expression Regulation, Developmental; D005835:Genitalia / Q000196:embryology*; D008297:Male; D051379:Mice / Q000196:embryology; D008815:Mice, Inbred Strains; D015335:Molecular Probes / Q000235:genetics; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D011944:Receptors, Androgen / Q000502:physiology*; D012727:Sex Characteristics
D000728:Androgens; D015335:Molecular Probes; D012333:RNA, Messenger; D011944:Receptors, Androgen; D004815:Epidermal Growth Factor