Bone marrow as a source of stem cells and germ cells? Perspectives for transplantation

Recent publications have suggested the existence of germ stem cells in the mouse at postnatal stages. The mechanism of de novo oocyte formation is proposed to involve a contribution from the bone marrow to the germ cell pool, via the bloodstream. Critical examination of the data underpinning these contentious claims is under way from a reproductive biology perspective but little has been said about the nature of this elusive bone marrow population with germ cell potential. Furthermore, whereas the prospect of marrow-derived germ cells may appear propitious for fertility applications, its wider impact on transplantation medicine remains to be considered. This paper examines the evidence leading to the current debate and considers the implications of such findings for the field of bone marrow transplantation. © 2007 Springer-Verlag

Dynamic distribution and stem cell characteristics of Sox1-expressing cells in the cerebellar cortex

Bergmann glia cells are a discrete radial glia population surrounding Purkinje cells in the cerebellar cortex. Although Bergmann glia are essential for the development and correct arborization of Purkinje cells, little is known about the regulation of this cell population after the developmental phase. In an effort to characterize this population at the molecular level, we have analyzed marker expression and established that adult Bergmann glia express Sox1, Sox2 and Sox9, a feature otherwise associated with neural stem cells (NSCs). In the present study, we have further analyzed the developmental pattern of Sox1-expressing cells in the developing cerebellum. We report that before becoming restricted to the Purkinje cell layer, Sox1-positive cells are present throughout the immature tissue, and that these cells show characteristics of Bergmann glia progenitors. Our study shows that these progenitors express Sox1, Sox2 and Sox9, a signature maintained throughout cerebellar maturation into adulthood. When isolated in culture, the Sox1-expressing cerebellar population exhibited neurosphere-forming ability, NSC-marker characteristics, and demonstrated multipotency at the clonal level. Our results show that the Bergmann glia population expresses Sox1 during cerebellar development, and that these cells can be isolated and show stem cell characteristics in vitro, suggesting that they could hold a broader potential than previously thought. © 2009 IBCB, SIBS, CAS. All rights reserved

A high-capacity screen for adipogenic differentiation

Glycerol-3-phosphate dehydrogenase (GPDH) is highly expressed in mature adipocytes. Activity of this enzyme is therefore routinely measured to assess adipogenic differentiation in cell cultures. Existing protocols for GPDH assays require relatively large amounts of cells, and throughput is limited due to multiple steps needed for cell harvest and enzyme extraction. We present here a new protocol allowing GPDH determinations to be performed in a 96-well-plate format. From the start of cell culture to the final readout all steps are carried out using the same multiwell plate, with a minimum of handling required. Our method is suitable for setting up high-throughput assays of adipogenic differentiation. © 2001 Academic Press

The chick embryo: Hatching a model for contemporary biomedical research

Animal models play a crucial role in fundamental and medical research. Progress in the fields of drug discovery, regenerative medicine and cancer research among others are heavily dependent on in vivo models to validate in vitro observations, and develop new therapeutic approaches. However, conventional rodent and large animal experiments face ethical, practical and technical issues that limit their usage. The chick embryo represents an accessible and economical in vivo model, which has long been used in developmental biology, gene expression analysis and loss/ gain of function experiments. It is also an established model for tissue/ cell transplantation, and because of its lack of immune system in early development, the chick embryo is increasingly recognised as a model of choice for mammalian biology with new applications for stem cell and cancer research. Here, we review novel applications of the chick embryo model, and discuss future developments of this in vivo model for biomedical research. © 2009 Wiley Periodicals, Inc

Consumer attitudes and preference exploration towards fresh-cut salads using best–worst scaling and latent class analysis

This research explored the preferences and buying habits of a sample of 620 consumers of fresh-cut, ready-to-eat salads. A best–worst scaling approach was used to measure the level of preference stated by individuals regarding 12 attributes for quality (intrinsic, extrinsic and credence) of fresh-cut salads. The experiment was carried out through direct interviews at several large-scale retail outlets in the Turin metropolitan area (north-west of Italy). Out of the total number of questioned consumers, 35% said they did not consume fresh-cut salads. On the contrary, the rest of the involved sample expressed the highest degree of preference towards the freshness/appearance attribute, followed by the expiration date and the brand. On the contrary, attributes such as price, organic certification and food safety did not emerge as discriminating factors in consumer choices. Additionally, five clusters of consumers were identified, whose preferences are related both to purchasing styles and socio-demographic variables. In conclusion, this research has highlighted the positive attitude of consumers towards quality products backed by a brand, providing ideas for companies to improve within this sector and implement strategies to answer the needs of a new segment of consumers, by determining market opportunities that aim to strengthen local brands

In vitro osteogenic differentiation of human ES cells

Since their isolation in 1998, human embryonic stem (hES) cells have been shown to be capable of adopting various cell fates in vitro. Here, we present in vitro data demonstrating the directed commitment of human embryonic stem cells to the osteogenic lineage. Human ES cells are shown to respond to factors that promote osteogenesis, leading to activation of the osteogenic markers osteocalcin, parathyroid hormone receptor, bone sialoprotein, osteopontin, cbfa1, and collagen 1. Moreover, the mineralized nodules obtained are composed of hydroxyapatite, further establishing the similarity of osteoblasts in culture to bone. These results show that osteoblasts can be derived from human ES cultures in vitro and provide the basis for comparison of adult and embryonic-derived osteogenesis, and for an investigation of potential applications for hES cells in orthopaedic tissue repair

Exposure to the ROCK inhibitor fasudil promotes gliogenesis of neural stem cells in vitro

Fasudil is a clinically approved Rho-associated protein kinase (ROCK) inhibitor that has been used widely to treat cerebral consequences of subarachnoid hemorrhage. It is known to have a positive effect on animal models of neurological disorders including Parkinson's disease and stroke. However, its cellular effect on progenitor populations and differentiation is not clearly understood. While recent studies suggest that fasudil promotes the mobilization of neural stem cells (NSCs) from the subventricular zone in vivo and promotes the differentiation of the C17.2 cerebellar neuroprogenitor line in vitro, it is unclear whether fasudil is involved in the differentiation of primary NSCs. Here, we tested the effect of fasudil on mouse NSCs in vitro, and observed increased gliogenesis in NSCs derived from lateral ventricles. Upon treatment, fasudil promoted characteristics of neurogenesis including phenotypic changes in neural outgrowth and interkinetic nuclear-like movements as an immediate response, while Sox2 expression was maintained and GFAP expression increased. Moreover, the gliogenic response to fasudil medium was observed in both early postnatal and adult NSC cultures. Taken together, our results show that fasudil promotes the differentiation of NSCs into astroglial lineage, suggesting that it could be used to develop novel vitro gliogenesis models and regulate differentiation for neural repair

Expression of a SOX1 overlapping transcript in neural differentiation and cancer models

SOX1 is a member of the SOXB1 subgroup of transcription factors involved in early embryogenesis, CNS development and maintenance of neural stem cells. The structure and regulation of the human SOX1 locus has been less studied than that of SOX2, another member of the SOXB1 subgroup for which an overlapping transcript has been reported. Here we report that the SOX1 locus harbours a SOX1 overlapping transcript (SOX1-OT), and describe expression, splicing variants and detection of SOX1-OT in different stem and cancer cells. RT-PCR and RACE experiments were performed to detect and characterise the structure of SOX1-OT in neuroprogenitor cultures and across different cancer cell lines. SOX1-OT was found to present a complex structure including several unannotated exons, different transcript variants and at least two potential transcription start sites. SOX1-OT was found to be highly expressed in differentiated neural stem cells across different time points of differentiation, and its expression correlated with SOX1 gene expression. Concomitant expression of SOX1 and SOX1-OT was further observed in several cancer cell models. While the function of this transcript is unknown, the regulatory role reported for other lncRNAs strongly suggests a possible role for SOX1-OT in regulating SOX1 expression, as previously observed for SOX2. The elucidation of the genetic and regulatory context governing SOX1 expression will contribute to clarifying its role in stem cell differentiation and tumorigenesis

Impact of serum source on human mesenchymal stem cell osteogenic differentiation in culture

Human mesenchymal stem cells (MSCs) show promise for musculoskeletal repair applications. Animal-derived serum is extensively used for MSC culture as a source of nutrients, extracellular matrix proteins and growth factors. However, the routine use of fetal calf serum (FCS) is not innocuous due to its animal antigens and ill-defined composition, driving the development of alternatives protocols. The present study sought to reduce exposure to FCS via the transient use of human serum. Transient exposure to animal serum had previously proved successful for the osteogenic differentiation of MSCs but had not yet been tested with alternative serum sources. Here, human serum was used to support the proliferation of MSCs, which retained surface marker expression and presented higher alkaline phosphatase activity than those in FCS-based medium. Addition of osteogenic supplements supported strong mineralisation over a 3-week treatment. When limiting serum exposure to the first five days of treatment, MSCs achieved higher differentiation with human serum than with FCS. Finally, human serum analysis revealed significantly higher levels of osteogenic components such as alkaline phosphatase and 25-Hydroxyvitamin D, consistent with the enhanced osteogenic effect. These results indicate that human serum used at the start of the culture offers an efficient replacement for continuous FCS treatment and could enable short-term exposure to patient-derived serum in the future

THE ROLE OF DIVERSITY AND DIVERSIFICATION FOR RESILIENT AGRICULTURAL SYSTEMS

Biodiversity is a unique and precious heritage: generic but also cultural, social and economic. Its drastic curtailment, however, puts at risk the survival of local farming systems, and this is even more so in fragile socio-economic contexts where it risks translating into conditions of food insecurity and poverty. From the elementary level of the gene, rising in complexity up to the ecosystem, it is therefore a central element in defining first the resistance and then the resilience of the system, and by the first term meaning the degree of resistance to a disruption that distances it from the initial state of equilibrium and by the second the capacity of a system to return to guaranteeing minimum standards following a disturbance, the capacity to get back on ones feet after a fall. It seems to be crucial, then, at a time when cooperation development projects that operate in various ways to safeguard and promote biodiversity are far more numerous, to intervene to preserve and restore the local biodiversity in order to avert future problems, and even curb them ahead of time, using resilience as an approach for managing the system we are dealing with (natural or heavily affected by human activity). In any event, this is a passage that is not routine, which makes it necessary to look at the ecosystem, at its various components, both natural and human. In the light of these preliminary remarks, the article will analyse the potentiality for applying, also in the field of development cooperation, the theoretical approach including empirical methods and instruments represented by the Diversified Farming Systems DFS), where the starting point is diversity and diversification as functional elements in the construction of resilient farming systems