No abstract

The neurosphere assay has been used to maintain neural progenitor cells (NPCs) in the undifferentiated state. These cells are multipotent and gave rise to neurons and glial cells. Here we show that within 10 days of culture, neurospheres contained precursors and differentiated progeny of all three major central nervous system (CNS) cell lineages and these occupied distinct zones. The microenvironment of the inner zone supported cell differentiation. Cells of oligodendroglial lineage generated within the neurosphere were frequently observed. Of these cells, A2B5(+) cells were homogeneously distributed in the neurospheres, NG2(+) cells preferentially occupied the outer zone and O4(+) cells were localized at the inner zone of 10 day-old neurospheres. We prevented a massive cell death of dissociated neurosphere cells seen after differentiation triggered with adhesion and fetal calf serum by adding epidermal growth factor and basic fibroblast growth factor to the culture medium. Under these conditions, less than one third of cells did not express cell specific markers, glial fibrillary acidic protein-positive astroglia represented 43.4%, NG2(+) and/or O4(+) oligodendroglia represented 24.3%, and betaIII-tubulin(+) neurons 3.1% of cells recovered after neurosphere differentiation. We present evidence that oligodendroglial cells differentiate in a stepwise process as a result of their distribution in subsets that represent distinct developmental stages according to antigenic and morphological criteria. These include oligodendrocyte progenitors, preoligodendrocytes, and oligodendrocytes. The highly complex morphology of mature oligodendrocytes was compatible with functional cells.

The purpose of the study was to compare the stability of the plate osteosyntheses of intra-articular calcaneal fractures using various types of a sustentacular screw insertions. A geometrical model of a calcaneal fracture was created. The fracture was fixed with a plate and screws with a uniform distribution. The individual models differed regarding the position of the sustentacular screw. The screw was inserted using three different variants: Model A: into the tip of sustentaculum tali, Model B: under the sustentaculum tali, and Model C: into the inferior peripherial rim of the sustentacular fragment. In all three variants, the screw was either locked into the plate via threads or unlocked. The model was loaded with force in the vertical direction. The stiffness of individual models was evaluated using the finite element method, which was expressed as the maximum force (Fmax) that the system was able to transmit and by determining the magnitude and distribution of reduced stress (σred) on the individual parts of the model of a fixed calcaneal fracture. The greatest stiffness of the system was observed in the Model B (Fmax = 335.8 N). The least stiffness was observed in Model C (Fmax = 296.3 N). This model also produced the greatest load on bone tissue was observed (σmaxred = 67.5 MPa). The least load on bone tissue was measured in Model B (σmaxred = 53.7 MPa). The load on the plate was similar in all three models (814.0–820.0 MPa). The analyses suggest that in a plate osteosynthesis of a calcaneal fracture, the insertion of a sustentacular screw under the tip of the sustentaculum tali is acceptable in terms of osteosynthesis stability. This sustentacular screw position reduces the risk of the screw penetrating into the talocalcaneal joint.

We described three different conditions that induce differentiation of dissociated neural stem cells derived from mouse embryonic CNS. In the first set of experiments, where the cell differentiation was triggered by cell adhesion, removal of growth factors and serum-supplemented medium, only sporadic neuronal and astroglial cells survived longer than two weeks and the latter formed a monolayer. When differentiation was induced in serum-free medium supplemented with retinoic acid, rapid and massive cell death occurred. A prolonged survival was observed in cultivation medium supplemented with serum and growth factors EGF plus FGF-2. One third of the cells did not express cell differentiation markers and were responsible for an increase in cell numbers. The remaining cells differentiated and formed the astrocytic monolayer on which occasional neuronal cells grew. One third of the differentiated phenotypes were represented by cells of oligodendroglial lineage. Differentiation of oligodendroglial cells occurred in a stepwise mechanism because the culture contained all successive developmental stages, including oligodendrocyte progenitors, preoligodendrocytes and immature and mature oligodendrocytes. Maturing oligodendrocytes displayed immunocytochemical and morphological features characteristic of cells that undergo physiological development. The cultivation conditions that supported growth and differentiation of neural stem cells were optimal for in vitro developmental studies and the production of oligodendroglial cells.