ABSTRACT
Testicular development occurs prenatally in mammals. The developmental underlying mechanism is only partially understood. The aim of the present investigation was to study the expression of the gene coding for insulin-like growth factor 1 (Igf-1) and Igf-1 type 1 receptor (Igf-1r) and their respective proteins in mouse Sertoli and Leydig cells at gestation day 12 (E12)-E18. Moreover, we sought to determine the effect of IGF-1 on the proliferation of both cell types and to establish the signal transduction mechanism involved in the IGF-1 pathway. Transcripts for the Igf-1 and Igf-1r genes were found in Sertoli and Leydig cells from E12-E18. Highest IGF-1 and IGF-Ir protein expression levels were found in both cell types at E18. Exogenous IGF-1 administration increased Sertoli and Leydig cell proliferation at E14-E18 in vitro. Inhibition of the pathway mitogen-activated extracellular signal-regulated protein kinase (MEK) 1/2 with UO126 diminished the proliferation of the Sertoli and Leydig cells in vitro. We propose that IGF-1 and IGF-1r regulate Sertoli and Leydig cell proliferation through the MEK/extracellular-signal-regulated kinase (ERK) 1/2 signal transduction pathway, leading to development and growth of the mouse embryonic testis.
Subject(s)
Cell Proliferation , Insulin-Like Growth Factor I/metabolism , Testis , Animals , Butadienes , Insulin-Like Growth Factor I/genetics , MAP Kinase Kinase 1/antagonists & inhibitors , MAP Kinase Kinase 1/metabolism , MAP Kinase Kinase 2/antagonists & inhibitors , MAP Kinase Kinase 2/metabolism , MAP Kinase Signaling System/physiology , Male , Mice , Nitriles , Protein Isoforms/genetics , Protein Isoforms/metabolism , Testis/cytology , Testis/embryology , Testis/metabolismABSTRACT
Insulin-like growth factor I (IGF-I) is important for gonadal and reproductive functions in mammals, although the physiological role of this growth factor during gonadal development in rodents remains largely unknown. Here, we examined the steady-state levels of IGF-I mRNA by the reverse transcriptase polymerase chain reaction (RT-PCR). IGF-I protein expression was also detected by Western blot. The effect of IGF-I as promoter of 17alpha-hydroxylase/C17-20 lyase and 17beta-hydroxysteroid dehydrogenase enzyme activity in vitro was evaluated by radioimmunoassay. Onset of IGF-I gene expression was on day E10 (urogenital ridge stage). IGF-I mRNA expression was markedly reduced on days E12 and E13 (testicular differentiation stage). IGF-I transcripts increased on day E14 and their transcription levels were maintained throughout the stages analyzed. Several IGF-I protein bands of 31-100 kDa were observed. Culture experiments demonstrated that 17alpha-hydroxyprogesterone and testosterone (T) secretion levels increased in the presence of IGF-I on days E11-E17. Additive effects of IGF-I plus (Bu)2cAMP were also seen during testicular development. It is proposed that IGF-I regulates the expression of key steroidogenic enzymes important for endocrine activity of the testis during prenatal development leading to establishment of the male phenotype and fertility.
Subject(s)
Endocrine System/metabolism , Insulin-Like Growth Factor I/physiology , Testis/embryology , 17-Hydroxysteroid Dehydrogenases/metabolism , Animals , Embryo, Mammalian , Endocrine System/embryology , Endocrine System/enzymology , Gestational Age , Gonads , Insulin-Like Growth Factor I/analysis , Male , Mice , Mice, Inbred Strains , RNA, Messenger/analysis , Steroid 17-alpha-Hydroxylase/metabolism , Testis/metabolism , Testosterone/biosynthesisABSTRACT
Developmental studies have shown that connexin43 (Cx43) is expressed in the ovary from the first day of life and throughout the rest of postnatal development. In both mouse embryonic ovaries and testes, target-directed deletion of Cx43 gene induces a significant decrease in germinal cells, but the exact mechanism determining this reduction remains unknown. Moreover, recently we found that Cx43 is abundantly expressed in mouse testes from the earliest stages of its fetal development. In the present work we investigate whether Cx43 transcript and protein are expressed in mouse embryonic ovaries. Total RNA was analyzed with specific Cx43 oligonucleotides in RT-PCR studies. A Cx43 PCR product was detected in ovaries at 16.5 and 18.5 days postcoitum (dpc). Bands of 43-45 kDa, characteristic of Cx43, were detected in immunoblots of total homogenates of ovaries at 14.5 and 18.5 dpc. Cell type-specific expression of Cx43 was investigated using double-labeled sections incubated with specific antibodies against Cx43 and the enzyme 3beta-hydroxysteroid dehydrogenase (3betaHSD) or a germ cell nuclear antigen (GCNA1), which are cell markers of steroidogenic and germinal cells, respectively. At 18.5 dpc, Cx43 was found in conglomerates of 3betaHSD-positive cells. Cx43 was also localized at homocellular junctions between parenchyma pregranulosa cells, and at heterocellular junctions between pregranulosa and germinal cells. At these two latter localizations, Cx43 was traced back to 12.5 dpc. In conclusion, this study demonstrates for the first time that from the earliest stages of embryonic ovary development, Cx43 is expressed in principal cell types involved in control of female fertility. These data suggest that the gap junctions formed with Cx43 between somatic and germinal cells may be necessary for prenatal expansion of germinal cells at initial stages of fetal gonadal development.