Purification of germline stem cells from adult mammalian ovaries: a step closer towards control of the female biological clock?

For decades it was believed that a non-renewable pool of oocyte-containing follicles is established in female mammals at birth. This cornerstone of reproductive biology was challenged 5 years ago by a study reporting on the presence of mitotically-active germ cells in juvenile and adult mouse ovaries. Additional findings presented in this study and others that followed further suggested that mammals retain the capacity to generate oocytes during adulthood; however, isolation of oocyte-producing germline stem cells (GSC) as unequivocal proof of their existence remained elusive. This piece of information now appears to have been provided by Ji Wu and colleagues. In addition to showing that proliferative germ cells resembling male spermatogonial stem cells can be purified from neonatal or adult mouse ovaries and maintained in vitro for months, transplantation studies demonstrated that these cells generate oocytes in ovaries of chemotherapy-sterilized recipients that fertilize and produce viable offspring. Although these findings do not establish that oogenesis occurs in adult females under physiological conditions, they strongly support the existence of GSC in adult mouse ovaries. If equivalent cells can be found in human ovaries, stem cell-based rejuvenation of the oocyte reserve in ovaries on the verge of failure may one day be realized

Spatio-temporal remodelling of the composition and architecture of the human ovarian cortical extracellular matrix during in vitro culture

Funding: MRC grant MR/R003246/1 and Wellcome Trust Collaborative Award in Science: 215625/Z/19/Z.Study question How does in vitro culture alter the human ovarian cortical extracellular matrix (ECM) network structure? Summary answer The ECM composition and architecture vary in the different layers of the ovarian cortex and are remodelled during in vitro culture. What is known already The ovarian ECM is the scaffold within which follicles and stromal cells are organized. Its composition and structural properties constantly evolve to accommodate follicle development and expansion. Tissue preparation for culture of primordial follicles within the native ECM involves mechanical loosening; this induces undefined modifications in the ECM network and alters cell–cell contact, leading to spontaneous follicle activation. Study design, size, duration Fresh ovarian cortical biopsies were obtained from six women aged 28–38 years (mean ± SD: 32.7 ± 4.1 years) at elective caesarean section. Biopsies were cut into fragments of ∼4 × 1 × 1 mm and cultured for 0, 2, 4, or 6 days (D). Participants/materials, settings, methods Primordial follicle activation, stromal cell density, and ECM-related protein (collagen, elastin, fibronectin, laminin) positive area in the entire cortex were quantified at each time point using histological and immunohistological analysis. Collagen and elastin content, collagen fibre characteristics, and follicle distribution within the tissue were further quantified within each layer of the human ovarian cortex, namely the outer cortex, the mid-cortex, and the cortex–medulla junction regions. Main results and the role of chance Primordial follicle activation occurred concomitantly with a loosening of the ovarian cortex during culture, characterized by an early decrease in stromal cell density from 3.6 ± 0.2 × 106 at day 0 (D0) to 2.8 ± 0.1 × 106 cells/mm3 at D2 (P = 0.033) and a dynamic remodelling of the ECM. Notably, collagen content gradually fell from 55.5 ± 1.7% positive area at D0 to 42.3 ± 1.1% at D6 (P = 0.001), while elastin increased from 1.1 ± 0.2% at D0 to 1.9 ± 0.1% at D6 (P = 0.001). Fibronectin and laminin content remained stable. Moreover, collagen and elastin distribution were uneven throughout the cortex and during culture. Analysis at the sub-region level showed that collagen deposition was maximal in the outer cortex and the lowest in the mid-cortex (69.4 ± 1.2% versus 53.8 ± 0.8% positive area, respectively, P < 0.0001), and cortical collagen staining overall decreased from D0 to D2 (65.2 ± 2.4% versus 60.6 ± 1.8%, P = 0.033) then stabilized. Elastin showed the converse distribution, being most concentrated at the cortex–medulla junction (3.7 ± 0.6% versus 0.9 ± 0.2% in the outer cortex, P < 0.0001), and cortical elastin peaked at D6 compared to D0 (3.1 ± 0.5% versus 1.3 ± 0.2%, P < 0.0001). This was corroborated by a specific signature of the collagen fibre type across the cortex, indicating a distinct phenotype of the ovarian cortical ECM depending on region and culture period that might be responsible for the spatio-temporal and developmental pattern of follicular distribution observed within the cortex. Large scale data N/A. Limitations, reasons for caution Ovarian cortical biopsies were obtained from women undergoing caesarean sections. As such, the data obtained may not accurately reflect the ECM distribution and structure of non-pregnant women. Wider implications of the findings Clarifying the composition and architecture signature of the human ovarian cortical ECM provides a foundation for further exploration of ovarian microenvironments. It is also critical for understanding the ECM–follicle interactions regulating follicle quiescence and awakening, leading to improvements in both in vitro activation and in vitro growth techniques.Publisher PDFPeer reviewe

Activin promotes oocyte development in ovine preantral follicles in vitro

Activins have been implicated as important regulating factors for many reproductive processes. The aim of this study was to determine the effect of activin A on the development of ovine preantral follicles in vitro. Mechanically isolated preantral follicles (161 ± 2 microm) were cultured for 6 days in the presence of human recombinant activin A (0, 10 and 100 ng/ml). Half of the medium was replaced every second day and follicle diameters were measured. Conditioned medium was subsequently analysed for oestradiol content using a delayed enhancement lanthanide fluorometric immunoassay (DELFIA). At the end of the culture period, follicles were fixed and processed for histology, after which oocyte diameter and granulosa cell death were measured. There was significant follicle growth over 6 days in all groups (p < 0.001). Activin, at both concentrations, increased follicle growth over control levels by Day 6 (p < 0.05). Oocyte diameters were also significantly increased by Day 6 of culture in all groups (p < 0.05), with 100 ng/ml activin increasing oocyte diameter over control levels (p < 0.05). Activin, at both concentrations, increased oestradiol production on Day 2 of culture, but this increase was not sustained during the culture period. Moreover, activin did not have any effect on antrum formation or follicle survival. In conclusion, activin promoted ovine preantral follicle and oocyte growth in vitro, but did not accelerate follicle differentiation over a six-day culture period. These results support a paracrine role for activin A during early oocyte and follicular development

Dynamic plantar loading index detects altered foot function in individuals with rheumatoid arthritis but not changes due to orthotic use

Background Altered foot function is common in individuals with rheumatoid arthritis. Plantar pressure distributions during gait are regularly assessed in this patient group; however, the association between frequently reported magnitude-based pressure variables and clinical outcomes has not been clearly established. Recently, a novel approach to the analysis of plantar pressure distributions throughout stance phase, the dynamic plantar loading index, has been proposed. This study aimed to assess the utility of this index for measuring foot function in individuals with rheumatoid arthritis.Methods Barefoot plantar pressures during gait were measured in 63 patients with rheumatoid arthritis and 51 matched controls. Additionally, 15 individuals with rheumatoid arthritis had in-shoe plantar pressures measured whilst walking in standardized footwear for two conditions: shoes-only; and shoes with prescribed custom foot orthoses. The dynamic plantar loading index was determined for all participants and conditions. Patient and control groups were compared for significant differences as were the shod and orthosis conditions.Findings The patient group was found to have a mean index of 0.19, significantly lower than the control group's index of 0.32 (p > 0.001, 95% CI [0.054, 0.197]). No significant differences were found between the shoe-only and shoe plus orthosis conditions. The loading index was found to correlate with clinical measures of structural deformity.Interpretation The dynamic plantar loading index may be a useful tool for researchers and clinicians looking to objectively assess dynamic foot function in patients with rheumatoid arthritis; however, it may be unresponsive to changes caused by orthotic interventions in this patient group.</p

A Novel Approach to Power Circuit Breaker Design for Replacement of SF6

This contribution explores the role of PTFE ablation in enhancing current interruption for various background gases in high voltage circuit breakers. An assessment of the current interruption capability has been made in terms of the arcing duration and the contact gap length at which critical arc extinction is achieved. These observations are supported by measurements of the magnitude of extinction and re-ignition voltage peaks. Most previous and other current experimental work on gas filled circuit breaker design follows conventional wisdom in investigating arcing behaviours at elevated gas pressures (usually up to 6 bar). But in this work we concentrate on the effects of using low gas pressures (less than 1 bar) in the presence of a close-fitting shield of ablatant polymer material (PTFE) that surrounds the electrode assembly of an experimental high power circuit breaker. We demonstrate that for several different gases, arc extinction capability compares well under these conditions with SF6, suggesting that SF6 could be replaced entirely in this novel system by more environmentally friendly gases. Moreover, the critical contact gap lengths at extinction are only slightly greater than when using SF6 at 6 bar. Weight loss measurements from the ablatant shield suggest that a chemical puffer action is the most likely mechanism for achieving the observed arc extinctions in this system