Epidermal growth factor mediates spermatogonial proliferation in newt testis

The complex processes of spermatogenesis are regulated by various factors. The aim of the current study is to determine the effect of epidermal growth factor (EGF) on spermatogonial proliferation and clarify the mechanism causing the proliferation in newt testis. In the organ culture, EGF stimulated spermatogonial proliferation, but not their differentiation into spermatocytes. cDNA cloning identified 3 members of the EGF receptors, ErbB1, ErbB2, and ErbB4, in the testis. RT-PCR showed that all the receptors cloned were expressed in both Sertoli and germ cells at the spermatogonial stage. In the organ cultures with inhibitors for the EGF receptors, mitogen-activated protein kinase (MAPK), and phosphoinositide 3-kinase (PI3K), the EGF-induced spermatogonial proliferation was suppressed. Furthermore, when the organ culture was exposed to EGF, the expressions of stem cell factor (SCF), immunoglobulin-like domain containing neuregulin1 (Ig-NRG1), and ErbB4 mRNA were increased. These results suggested that, since the spermatogonia are sequestered within cysts by the blood-testis barrier consisted of Sertoli cells, EGF possibly mediates spermatogonial proliferation in an endocrine manner through the receptors including ErbB1, ErbB2, and ErbB4 expressed on Sertoli cells via activation of MAPK cascade or/and PI3K cascade by elevating the expressions of SCF, Ig-NRG1, and ErbB4

Effect of EGF on spermatogonial proliferation and their differentiation to primary spermatocytes

(A and B) Stimulatory effect of EGF on spermatogonial proliferation. Testicular fragments containing speramatogonial stage were cultured for 1 week in the absence or presence of either FSH or various doses of EGF indicated. (A) Immunohistochemistry for BrdU incorporation in the sections of the fragments treated without (Control) or with either FSH or EGF. (B) Spermatogonial proliferation was determined by counting BrdU positive cysts among live ones in at least 3 sections. *, P < 0.05. (C) No effect of EGF on spermatogonial differentiation. Testicular fragments containing spermatogonial stage were cultured for 2 weeks in the absence (Control) or presence of either FSH or EGF, followed by staining with hematoxylin/eosin. Primary spermatocytes are surrounded by black dashes.<p><b>Copyright information:</b></p><p>Taken from "Epidermal growth factor mediates spermatogonial proliferation in newt testis"</p><p>http://www.rbej.com/content/6/1/7</p><p>Reproductive biology and endocrinology : RB&E 2008;6():7-7.</p><p>Published online 6 Feb 2008</p><p>PMCID:PMC2276507.</p><p></p

Dose-dependent effects of inhibitors for ErbB on EGF-stimulated spermatogonial proliferation

Testicular fragments containing spermatogonial stage treated without (0) or with various doses of pan ErbB4 inhibitor (PD153035) (A), an ErbB2-specific inhibitor (AG879) (B), or an ErbB1-specific inhibitor (AG1478) (C) were cultured for 1 week in the absence (Control) or presence of either FSH (200 ng/μl) or EGF (4000 ng/μl), followed by BrdU incorporation assay. *, P < 0.05; **, P < 0.01.<p><b>Copyright information:</b></p><p>Taken from "Epidermal growth factor mediates spermatogonial proliferation in newt testis"</p><p>http://www.rbej.com/content/6/1/7</p><p>Reproductive biology and endocrinology : RB&E 2008;6():7-7.</p><p>Published online 6 Feb 2008</p><p>PMCID:PMC2276507.</p><p></p

Expressions of mRNA for ErB1, ErbB2, and ErbB4 in various spermatogenic stages and testicular cell types

The expressions of ErbB1 (200 bp), ErbB2 (150 bp), and ErbB4 (150 bp) were analyzed by semi-quantitative RT-PCR. Total RNA was extracted from the testes containing early spermatogonial (1st – 4th generation), late spermatogonial (5th – 7th generation), and primary spermatocyte stages (A and B), and from the fractionated spermatogonia and Sertoli cells, the latter of which were cultured for 10 days and 1 month (C). The cycle numbers in PCR were 35 (A) and 33 (B), and 35 for ErbB1 and ErbB2 and 40 for ErbB4 (C). Elongation factor-1α (EF-1α, 550 bp, 25 cycles) is the internal control. SCF is a Sertoli cell-specific marker. Data are representative of at least 3 independent experiments.<p><b>Copyright information:</b></p><p>Taken from "Epidermal growth factor mediates spermatogonial proliferation in newt testis"</p><p>http://www.rbej.com/content/6/1/7</p><p>Reproductive biology and endocrinology : RB&E 2008;6():7-7.</p><p>Published online 6 Feb 2008</p><p>PMCID:PMC2276507.</p><p></p

Dose-dependent effects of inhibitors for MAPK and PI3K on EGF-stimulated spermatogonial proliferation

Testicular fragments containing spermatogonial stage treated without (0) or with various doses of a MAPK-specific inhibitor (PD98059) (A) or PI3K-specific inhibitors (Wortmannin (B) and LY294002 (C)), were cultured for 1 week in the absence (Control) or presence of either FSH (200 ng/μl) or EGF (4000 ng/μl), followed by BrdU incorporation assay. *, P < 0.05; **, P < 0.01.<p><b>Copyright information:</b></p><p>Taken from "Epidermal growth factor mediates spermatogonial proliferation in newt testis"</p><p>http://www.rbej.com/content/6/1/7</p><p>Reproductive biology and endocrinology : RB&E 2008;6():7-7.</p><p>Published online 6 Feb 2008</p><p>PMCID:PMC2276507.</p><p></p