RESUMO
The ability to mature human primordial follicles in vitro would assist fertility restoration. However, the signals initiating growth of primordial follicles are unknown. Growth factors such as nerve growth factor (NGF) may play a role in this process. To investigate the expression of NGF and its receptors, p75 and TrkA, in early developing follicles (mostly primordial, primary and secondary follicles), ten ovarian samples from adolescents/adults aged 13-39 and 33 ovaries from human fetuses aged 19-33 gestational weeks (GW) were obtained and immediately fixed or frozen. The fixed samples were prepared for a study of immunocytochemical staining of NGF and its two receptors. Total RNA was extracted from the frozen ovarian samples, and the expression of NGF, TrkA and p75 was investigated by RT-PCR. Products were resolved by 1% agarose gel electrophoresis and image analysis. Immunocytochemical staining revealed the expression of NGF in granulosa cells (GC) and oocytes; TrkA was mainly in oocytes and in GC in minority of the samples; and p75 was in some of the stroma cells from fetuses aged less than 22 GW. Transcripts of NGF and TrkA were identified by RT-PCR in all samples, while those for p75 were detected only in ovarian samples from fetuses aged less than 22 GW. To elucidate if NGF is indeed involved in growth initiation of human primordial follicles, it should be added to their culture medium. The immunocytochemical detection of p75 in some of the stroma cells and transcripts in ovarian samples of fetuses less than 22 GW may suggest its role in follicular assembly.
Assuntos
Fator de Crescimento Neural/metabolismo , Folículo Ovariano/embriologia , Folículo Ovariano/crescimento & desenvolvimento , Receptor trkA/metabolismo , Receptores de Fator de Crescimento Neural/metabolismo , Adolescente , Adulto , Feminino , Feto/citologia , Células da Granulosa/química , Células da Granulosa/metabolismo , Humanos , Fator de Crescimento Neural/análise , Fator de Crescimento Neural/genética , Oócitos/química , Oócitos/metabolismo , Reação em Cadeia da Polimerase , RNA Mensageiro/análise , RNA Mensageiro/metabolismo , Receptor de Fator de Crescimento Neural , Receptor trkA/análise , Receptor trkA/genética , Receptores de Fator de Crescimento Neural/análise , Receptores de Fator de Crescimento Neural/genética , Transcrição GênicaRESUMO
The ability to mature human primordial follicles in vitro would assist fertility restoration. However, the signals initiating growth of primordial follicles are unknown. Growth factors such as leukaemia inhibitory factor (LIF) may play a role in this process. To investigate the expression of LIF and its receptor in early developing follicles, nine ovarian samples from adolescents/adults aged 13-43 years and 23 ovaries from human fetuses aged 19-33 gestational weeks were immediately fixed or frozen. The fixed samples were prepared for a study of immunocytochemical staining of LIF and its two receptor units (LIF-R and gp 130). mRNA was extracted from the frozen ovarian samples, and the expression of LIF, LIF-R and gp 130 was investigated by RT-PCR. Products were resolved by 10% polyacrylamide gel electrophoresis and image analysis. There was strong to moderate immunocytochemical staining for LIF and LIF-R in oocytes from the primordial follicular stages onwards, and very weak to moderate staining for gp 130. LIF-R was also detected in granulosa cells of primary and secondary follicles from adolescents/adults. Transcripts of LIF, LIF-R and gp 130 RNA were identified by RT-PCR in all samples. The immunocytochemical staining and mRNA expression of LIF and its receptor are consistent with the concept that LIF might be involved in growth initiation of human primordial follicles through its receptor.
