MYO10 promotes transzonal projection-dependent germ line-somatic contact during mammalian folliculogenesis†.

Granulosa cells of growing ovarian follicles elaborate filopodia-like structures termed transzonal projections (TZPs) that supply the enclosed oocyte with factors essential for its development. Little is known, however, of the mechanisms underlying the generation of TZPs. We show in mouse and human that filopodia, defined by an actin backbone, emerge from granulosa cells in early stage primary follicles and that actin-rich TZPs become detectable as soon as a space corresponding to the zona pellucida appears. mRNA encoding Myosin10 (MYO10), a motor protein that accumulates at the base and tips of filopodia and has been implicated in their initiation and elongation, is present in granulosa cells and oocytes of growing follicles. MYO10 protein accumulates in foci located mainly between the oocyte and innermost layer of granulosa cells, where it colocalizes with actin. In both mouse and human, the number of MYO10 foci increases as oocytes grow, corresponding to the increase in the number of actin-TZPs. RNAi-mediated depletion of MYO10 in cultured mouse granulosa cell-oocyte complexes is associated with a 52% reduction in the number of MYO10 foci and a 28% reduction in the number of actin-TZPs. Moreover, incubation of cumulus-oocyte complexes in the presence of epidermal growth factor, which triggers a 93% reduction in the number of actin-TZPs, is associated with a 55% reduction in the number of MYO10 foci. These results suggest that granulosa cells possess an ability to elaborate filopodia, which when directed toward the oocyte become actin-TZPs, and that MYO10 increases the efficiency of formation or maintenance of actin-TZPs.

Immunocytochemical detection and reverse transcription polymerase chain reaction expression of oncostatin M (OSM) and its receptor (OSM-Rbeta) in human fetal and adult ovaries.

OBJECTIVE: To investigate the immunocytochemical expression and presence of mRNA transcripts of oncostatin M (OSM) and its exclusive receptor (OSM-Rbeta) in ovaries from human adults and fetuses. DESIGN: Immunocytochemical and reverse transcription polymerase chain reaction (RT-PCR) study. SETTING: Major tertiary care and referral academic centers. PATIENT(S): Ten women and girls undergoing laparoscopic ovarian biopsy and 30 women undergoing second-trimester and third-trimester pregnancy terminations. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Microscopic morphometric analysis, immunocytochemistry for OSM and OSM-Rbeta, and RT-PCR analyses. RESULT(S): There was strong to moderate immunocytochemical staining for OSM in both oocytes and granulosa cells of follicles from primordial stages onwards in ovaries from both fetuses and adults/adolescents. OSM-Rbeta was detected mainly in the oocytes. Transcripts of OSM and OSM-Rbeta RNA were detected by RT-PCR analyses. CONCLUSION(S): The expression of OSM and its receptor in ovarian tissue from fetuses and women suggests a possible role of OSM in growth initiation of human primordial follicles.

Expression of stem cell factor and its receptor in human fetal and adult ovaries.

OBJECTIVE: To investigate the immunocytochemical expression and presence of mRNA transcripts of stem cell factor (SCF) and its receptor (SCF-R) in ovaries from human adults and fetuses. DESIGN: Immunocytochemical and reverse transcription polymerase chain reaction (RT-PCR) study. SETTING: Major tertiary care and referral academic centers. PATIENT(S): Seven women and girls undergoing laparoscopic ovarian biopsy and 13 women undergoing second and third trimester pregnancy terminations. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Microscopic morphometric analysis, immunocytochemistry for SCF and SCF-R, and RT-PCR analyses. RESULT(S): There was strong to moderate immunocytochemical staining for SCF and its receptor in oocytes from primordial stages onward, but not in granulosa cells, in both fetal and adult ovarian samples. Transcripts of SCF and SCF-R RNA were detected by RT-PCR analyses for SCF and SCF-R. CONCLUSION(S): The expression of SCF and its receptor in ovarian tissue from fetuses and women suggests a possible role of SCF in growth initiation of human primordial follicles.

First successful pregnancy outcome after preimplantation genetic diagnosis for aneuploidy screening in embryos generated from natural-cycle in vitro fertilization combined with an in vitro maturation procedure.

OBJECTIVE: To report on the first successful pregnancy performing preimplantation genetic diagnosis (PGD) on embryos obtained after combining natural IVF with in vitro maturation procedure. DESIGN: Case report. SETTING: University hospital. PATIENT(S): A 35-year-old patient with polycystic ovaries who was attending the fertility clinic. INTERVENTION(S): In vitro maturation of immature oocytes and preimplantation genetic diagnosis. MAIN OUTCOME MEASURE(S): Preimplantation genetic diagnosis of aneuploidy screening in embryos produced from in vitro maturation procedure. RESULT(S): Thirteen immature and two mature oocytes were collected, and eight mature oocytes were normally fertilized, of which six embryos were biopsied for chromosome analysis. Two chromosomally normal embryos were transferred on day 5, resulting in the birth of a healthy child. CONCLUSION(S): This case report demonstrates that an acceptable number of embryos can be generated after in vitro maturation procedure to perform PGD for aneuploidy screening and that this approach can be extended to patients with polycystic ovaries or polycystic ovary syndrome who are undergoing PGD for other genetic diseases.

Diploid-aneuploid mosaicism in human embryos cultured to the blastocyst stage.

OBJECTIVE: To examine diploid-aneuploid mosaicism in human in vitro cultured blastocysts. DESIGN: A laboratory study on spare blastocysts from an IVF program. SETTING: University hospital laboratory. PATIENTS(S): Forty-three couples undergoing IVF or intracytoplasmic sperm injection. INTERVENTION(S): Ninety-one blastocysts were spread for fluorescence in situ hybridization using the HCl-Tween 20 method. A total of 6,664 nuclei were analyzed for aneuploidy using fluorescent DNA probes specific to chromosomes 2, 7, and 18. MAIN OUTCOME MEASURE(S): The proportion of aneuploid cells within each blastocyst. RESULTS(S): The incidence of diploid-aneuploid mosaicism among 91 blastocysts examined was 17.6%. All of the mosaic blastocysts were abnormal for only one of the three chromosomes tested, with the incidence of involvement of chromosomes 2, 7, and 18 being 3.3%, 8.8%, and 5.5%, respectively. The majority of the mosaic blastocysts had low proportions of aneuploid cells. Ten of the 16 (62.5%) affected blastocysts were of morphology compatible with implantation. CONCLUSION(S): A considerable proportion of human IVF blastocysts show a form of mosaicism that has been observed in fetal and placental tissues. This mosaicism often arises at the final stage of preimplantation development in vitro and is present in blastocysts of morphology compatible with implantation.

Leptin and its receptors in human fetal and adult ovaries.

The objective of the study was to investigate the immunocytochemical expression and presence of messenger RNA (mRNA) transcripts for leptin and its receptors in ovaries from human adults and adolescents and second- and third-trimester fetuses. Staining for leptin and the long form of its receptor was identified in oocytes of follicles from primordial stages onward, and for leptin only in granulosa cells of a minority of the samples. Expression of mRNA transcripts for both ligands was detected in all the samples tested.

Expression of basic fibroblast growth factor and its receptors in human ovarian follicles from adults and fetuses.

OBJECTIVE: To investigate the immunocytochemical expression and presence of mRNA transcripts for basic fibroblast growth factor (bFGF) and its four receptors (FGFR-1, -2, -3, and -4) in ovaries from human adults and fetuses. DESIGN: Immunocytochemical and reverse transcription polymerase chain reaction (RT-PCR) study. SETTING: Major tertiary care and referral academic centers. PATIENT(S): Nine women and girls undergoing laparoscopic ovarian biopsy and 26 women undergoing second- and third-trimester pregnancy terminations. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Microscopic morphometric analysis, immunocytochemistry for bFGF and its receptors, and RT-PCR analyses. RESULT(S): The proteins for bFGF, FGFR-2, FGFR-3, and FGFR-4 were identified in oocytes of all follicular classes. Immunocytochemical expression of bFGF and its receptors was detected in granulosa cells of follicles from adolescents/women but not from fetuses. There was no immunocytochemical expression of FGFR-1. Transcripts of bFGF and its four receptors were identified by RT-PCR in all samples. CONCLUSION(S): The expression of bFGF and its receptors in human ovaries suggests that bFGF might have a role in early folliculogenesis.