Direct reprogramming of human somatic cells into pluripotency has broad implications in generating patient-specific induced pluripotent stem (iPS) cells for disease modeling and cellular replacement therapies. However, the low efficiency and safety issues associated with generation of human iPS cells have limited their usage in clinical settings. Cell types can significantly influence reprogramming efficiency and kinetics. To date, human iPS cells have been obtained only from a few cell types. Here, we report for the first time rapid and efficient generation of iPS cells from human amniotic fluid-derived cells (hAFDCs) via ectopic expression of four human factors: OCT4/SOX2/KLF4/C-MYC. Significantly, typical single iPS cell colonies can be picked up 6 days after viral infection with high efficiency. Eight iPS cell lines have been derived. They can be continuously propagated in vitro and express pluripotency markers such as AKP, OCT4, SOX2, SSEA4, TRA-1-60 and TRA-1-81, maintaining the normal karyotype. Transgenes are completely inactivated and the endogenous OCT4 promoter is adequately demethylated in the established iPS cell lines. Moreover, various cells and tissues from all three germ layers are found in embryoid bodies and teratomas, respectively. In addition, microarray analysis demonstrates a high correlation coefficient between hAFDC-iPS cells and human embryonic stem cells, but a low correlation coefficient between hAFDCs and hAFDC-iPS cells. Taken together, these data identify an ideal human somatic cell resource for rapid and efficient generation of iPS cells, allowing us to establish human iPS cells using more advanced approaches and possibly to establish disease- or patient-specific iPS cells.

Meiotic chromosome segregation in human oocytes is notoriously error-prone, especially with ageing. Such errors markedly reduce the reproductive chances of increasing numbers of women embarking on pregnancy later in life. However, understanding the basis for these errors is hampered by limited access to human oocytes. Important new discoveries have arisen from molecular analyses of human female recombination and aneuploidy along with high-resolution analyses of human oocyte maturation and mouse models. Here, we review these findings to provide a contemporary picture of the key players choreographing chromosome segregation in mammalian oocytes and the cellular basis for errors. A search of PubMed was conducted using keywords including meiosis, oocytes, recombination, cohesion, cohesin complex, chromosome segregation, kinetochores, spindle, aneuploidy, meiotic cell cycle, spindle assembly checkpoint, anaphase-promoting complex, DNA damage, telomeres, mitochondria, female ageing and female fertility. We extracted papers focusing on mouse and human oocytes that best aligned with the themes of this review and that reported transformative and novel discoveries. Meiosis incorporates two sequential rounds of chromosome segregation executed by a spindle whose component microtubules bind chromosomes via kinetochores. Cohesion mediated by the cohesin complex holds chromosomes together and should be resolved at the appropriate time, in a specific step-wise manner and in conjunction with meiotically programmed kinetochore behaviour. In women, the stage is set for meiotic error even before birth when female-specific crossover maturation inefficiency leads to the formation of at-risk recombination patterns. In adult life, multiple co-conspiring factors interact with at-risk crossovers to increase the likelihood of mis-segregation. Available evidence support that these factors include, but are not limited to, cohesion deterioration, uncoordinated sister kinetochore behaviour, erroneous microtubule attachments, spindle instability and structural chromosomal defects that impact centromeres and telomeres. Data from mice indicate that cohesin and centromere-specific histones are long-lived proteins in oocytes. Since these proteins are pivotal for chromosome segregation, but lack any obvious renewal pathway, their deterioration with age provides an appealing explanation for at least some of the problems in older oocytes. Research in the mouse model has identified a number of candidate genes and pathways that are important for chromosome segregation in this species. However, many of these have not yet been investigated in human oocytes so it is uncertain at this stage to what extent they apply to women. The challenge for the future involves applying emerging knowledge of female meiotic molecular regulation towards improving clinical fertility management.

Self-renewal and differentiation of embryonic stem cells (ESCs) are controlled by intracellular transcriptional factors and extracellular factor-activated signaling pathways. Transcription factor Oct4 is a key player maintaining ESCs in an undifferentiated state, whereas the Erk/MAPK pathway is known to be important for ESC differentiation. However, the manner in which intracellular pluripotency factors modulate extracellular factor-activated signaling pathways in ESCs is not well understood. Here, we report identification of a target gene of Oct4, serine/threonine kinase 40 ( Stk40 ), which is able to activate the Erk/MAPK pathway and induce extraembryonic–endoderm (ExEn) differentiation in mouse ESCs. Interestingly, cells overexpressing Stk40 exclusively contribute to the ExEn layer of chimeric embryos when injected into host blastocysts. In contrast, deletion of Stk40 in ESCs markedly reduces ExEn differentiation in vitro. Mechanistically, Stk40 interacts with Rcn2, which also activates Erk1/2 to induce ExEn specification in mouse ESCs. Moreover, Rcn2 proteins are specifically located in the cytoplasm of the ExEn layer of early mouse embryos. Importantly, knockdown of Rcn2 blocks Stk40 -activated Erk1/2 and ESC differentiation. Therefore, our study establishes a link between the pluripotency factor Oct4 and the Erk/MAPK signaling pathway, and it uncovers cooperating signals in the Erk/MAPK activation that control ExEn differentiation.

The NAD⁺‐dependent sirtuin deacetylase, Sirt1, regulates key transcription factors strongly implicated in ageing and lifespan. Due to potential confounding effects secondary to loss of Sirt1 function from the soma in existing whole‐animal mutants, the in vivo role of Sirt1 in oocytes (oocyte‐Sirt1) for female fertility remains unknown. We deleted Sirt1 specifically in growing oocytes and study how loss of oocyte‐Sirt1 affects a comprehensive range of female reproductive parameters including ovarian follicular reservoir, oocyte maturation, oocyte mitochondrial abundance, oxidative stress, fertilization, embryo development and fertility during ageing. Surprisingly, eliminating this key sirtuin from growing oocytes has no effect in young females. During a 10‐month‐long breeding trial, however, we find that 50% of females lacking oocyte‐Sirt1 become prematurely sterile between 9 and 11 months of age when 100% of wild‐type females remain fertile. This is not due to an accelerated age‐related decline in oocyte numbers in the absence of oocyte‐Sirt1 but to reduced oocyte developmental competence or quality. Compromised oocyte quality does not impact in vivo oocyte maturation or fertilization but leads to increased oxidative stress in preimplantation embryos that inhibits cleavage divisions. Our data suggest that defects emerge in aged females lacking oocyte‐Sirt1 due to concurrent age‐related changes such as reduced NAD⁺ and sirtuin expression levels, which compromise compensatory mechanisms that can cover for Sirt1 loss in younger oocytes. In contrast to evidence that increasing Sirt1 activity delays ageing, our data provide some of the only in vivo evidence that loss of Sirt1 induces premature ageing.

Female meiotic divisions are extremely asymmetric, producing large oocytes and small polar bodies (PBs). In mouse oocytes, the spindle relocates to the cortex before anaphase of meiosis I (MI). It is presumed that by displacing the future midzone, pre-anaphase spindle repositioning alone ensures asymmetry. But how subsequent anaphase events might contribute to asymmetric PB extrusion (PBE) is unknown. Here, we find that inactivation of cyclin-dependent kinase 1 (Cdk1) induces anaphase and simultaneously triggers cytoplasmic formin-mediated F-actin polymerisation that propels the spindle into the cortex causing it to protrude while anaphase progresses. Significantly, if post-anaphase-onset spindle migration fails, protrusion and asymmetry are severely threatened even with intact pre-anaphase migration. Conversely, post-anaphase migration can completely compensate for failed pre-anaphase migration. These data identify a cell-cycle-triggered phase of spindle displacement occurring after anaphase-onset, which, by inducing protrusion, is necessary for extreme asymmetry in mouse oocytes and uncover a pathway for maximising unequal division.