Development of the cerebellum in turbot (Psetta maxima): Analysis of cell proliferation and distribution of calcium binding proteins.

The morphogenesis, cell proliferation and neuronal differentiation of the turbot (Psetta maxima) cerebellum has been studied using conventional histological techniques and immunohistochemical methods for proliferating cell nuclear antigen and calcium binding proteins. As in other vertebrates, the cerebellar anlage emerges as proliferative plates of neural tissue during the embryonic period. The anlage of the cerebellum persists without morphological changes until the end of the larval life when the mantle zone is differentiated. The major ontogenetic changesthat drive the formation of the cerebellar subdivisions begin in late premetamorphic larvae when cerebellar plates growth and merge medially. This transformation is accomplished by the reorganization of proliferative zones as well as by the onset of cell differentiation. The cerebellum becomes fully differentiated during metamorphosis when parvalbumin and calretinin were detected in Purkinje and eurydendroid cells. Sustained proliferation is maintained in all subdivisions of the cerebellum and this support the robust growth of this part of the brain that takes place during the metamorphic and juvenile periods.The location and histological organization of the proliferative activity in the turbot mature cerebellum are described and their functional significance was analyzed in light of the information available for other teleosts.

Analysis of the morphogenesis and cell proliferation in the retina of a pleuronectiform fish, the turbot Psetta maxima (Pleuronectiformes: Teleostei).

The morphogenesis and cell proliferation in the retina of turbot (Psetta maxima, Pleuronectiformes: Teleostei) from embryo through metamorphosis have been examined by using proliferating cell nuclear antigen (PCNA)-immunohistochemistry and general histological procedures. In the embryonic retina, cell proliferation and spatial cell reorganization form the anlage of the pigment epithelium and neural retina. Neurogenesis begins around hatching in the temporal retina, dorsal to the optic nerve exit, and then a wave of cell differentiation spreads to the nasal retina to yield a laminated retina by the end of the prolarval turbot stage. Germinal zones in the differentiated retina persist as a rim at the retinal margin, as well as surrounding the optic fissure in premetamorphic and metamorphic turbot larvae. In these zones, progenitor cells with different morphologies show a similar spatial arrangement, which suggests that they have a similar retinogenic potential. During metamorphosis, asymmetric proliferative activity in turbot germinal zones is associated with a marked expansion of the retinal tissue. Scattered stem cells in the laminated retina, related to the lineage of rod photoreceptors, were also observed both in large premetamorphic larvae and metamorphic turbots. The proliferative activity of these cells increases considerably during metamorphosis, when rod photoreceptors become morphologically differentiated.

Assessing methods for blood cell cytotoxic responses to inorganic nanoparticles and nanoparticle aggregates.

Inorganic nanoparticles (NPs) show great potential for medicinal therapy. However, biocompatibility studies are essential to determine if they are safe. Here, five different NPs are compared for their cytotoxicity, internalization, aggregation in medium, and reactive oxygen species (ROS) production, using tumoral and normal human blood cells. Differences depending on the cell type are analyzed, and no direct correlation between ROS production and cell toxicity is found. Results are discussed with the aim of standardizing the procedures for the evaluation of the toxicity.

Activity and properties of alpha-L-fucosidase are dependent on the state of enterocytic differentiation of HT-29 colon cancer cells.

Previously we have demonstrated an impairment in the activity of alpha-L-fucosidase in colon tumours. In order to establish an in vitro model to study this enzyme in colon cancer, we have determined the activity and properties of the enzyme during the differentiation of HT-29 colon cancer cells. Cultures were committed to differentiate into enterocyte-like cells by placing them in a culture medium without glucose for 18-21 days. The state of differentiation was evaluated by assaying the activity of enterocytic marker enzymes, and the acquisition of enterocyte morphology was assessed by electron microscopy. The alpha-L-fucosidase activity was determined using a fluorometric method. Intracellular levels of alpha-L-fucosidase activity are lower in non-differentiated cells (3.0 +/- 1.01 U/mg) than in differentiated ones (9.2 +/- 4.09 U/mg) (P < 0.001). This variation is not due to a greater secretion of the enzyme to the culture medium, and properties such as pH optimum or the affinity towards substrate are not dependent on differentiation. The enzyme however, is more stable at acidic pH and at high temperatures, and V(max) is higher in differentiated cells. Moreover, in undifferentiated cells the enzyme is mainly in a monomeric form whereas multimeric forms of the enzyme appear only upon differentiation. Most of these changes are very similar to those previously observed between normal colon tissue and colon tumours. Thus, we suggest that differentiation of HT-29 colon cancer cells could be used as a model to study the alterations of the enzyme alpha-L-fucosidase during the progression of the tumoural process.