Molecular organization of rat prolactin granules. I. In vitro stability of intact and "membraneless" granules.

Studies carried out on a number of secretory cell systems suggest that the specific cytoplasmic granules in which the secretion products are stored before their release are complex organelles which can possess a distinct molecular organization. For instance, it has been reported that in some granules the segregated secretion products are organized into crystalline structures (1-3) or large intermolecular aggregates (4-8). It is likely that all phenomena of this type are favorable to the economy of the cell, in the sense that they reduce the energy required for storage of the secretion products. The prolactin (LTH) granules of the rat pituitary possess a number of morphological features which strongly suggest that the molecules(s) of their content might be arranged in a relatively stable structure. Thus, these granules are remarkably polymorphic in shape, and their membrane is usually separated from their content by a clear space. Furthermore, identifiable LTH granules devoid of their membrane are often seen in the pericapillary space, suggesting that upon discharge by exocytosis they are dissolved only slowly (9). However, no studies specifically concerned with the mechanisms of LTH storage have been reported so far. In order to obtain some information on this question, we have studied the behavior of isolated granule fractions incubated in vitro under a variety of carefully controlled experimental conditions.     

The interplay between local and landscape scales on the density of pond-dwelling anurans in subtropical grasslands

We investigated the role of local and landscape environmental variables on anurans density classified as habitat specialists and generalists in grassland landscapes, known as South Brazilian grasslands (SBG). In this region, we surveyed 187 ponds distributed over 40 landscape sampling units. For each pond, 31 local environmental variables were measured. Each landscape sampling unit was embedded within a larger regional sampling unit with different landscape properties. For each landscape and regional sampling units, 16 landscape metrics were extracted from a land cover and use map. We recorded 35 species, eleven of which are specialists in the SBG. The specialists were affected by 11 local and 2 landscape environmental variables, while generalists were affected by 14 local and one landscape environmental variable. Thus, specialists and generalists presented different relationships with local and landscape variables, but in general local variables had a greater influence on the density of anurans than the landscape variables. However, the landscape indirectly influenced local variables because higher quality ponds were in landscapes with higher percentages of natural habitat. In conclusion, reproductive sites with higher local quality and located within landscapes with higher percentages of natural grasslands are essential to conserve anurans in this habitat. Effective conservation of such sites would benefit from further studies that assess effects of land use and biotic integrity of ponds, which can help to determine (a) the relative effects of local habitat quality of ponds and (b) the effectiveness of protecting ponds and their local surroundings for anuran conservation in SBG. Abstract in Portuguese is available with online material.     

Evaluation of the ratio of ω6 : ω3 fatty acids and vitamin e levels in the diet on the reproductive performance of cockerels

Three hundred and twenty 30-week old White Leghorn cockerels were housed in individual cages and distributed in a completely randomized factorial design of 5?×?3, with five oil sources (sunflower, soybean, canola, linseed and fish/soybean) and three levels of antioxidant (30, 200 and 400?mg of vitamin E/kg). The aim of this study was to evaluate the effect of the ratio of ω6?:?ω3 fatty acids by the inclusion of different oil sources and of dietary supplementation with vitamin E on the reproductive performance of cockerels. The use of the fish/soybean combination determined the lowest total antioxidant status of the semen. However, the addition of vitamin E to the fish/soybean-oil-based diet resulted in a linear increase in semen volume, motility and sperm vigour in the 38th week and again in the 52nd week for motility and for sperm vigour and fertility rate in the periods from 50?–?53 and 41?–?53 weeks of age. The use of canola oil in the diet resulted in the highest fertility rate during 50?–?53 and 41?–?53 weeks of life. Animals receiving the soybean oil based diet showed the smallest fertility rate in the range from 50?–?53 weeks of age and concomitantly the highest level of cholesterol in the spermatozoa in the range from 47?–?51 weeks. An interaction between the vitamin E level and soybean oil was verified by the linear increase in motility and sperm vigour at 38 weeks of age. Later, the contrary was shown by a linear reduction in fertility in the periods from 44?–?46, 47?–?49 and 41?–?53 weeks of age. Cockerels that had been fed on the sunflower-oil-based diet showed the highest content of saturated fatty acids in the spermatozoa from 48?–?51 weeks. An interaction effect was observed between the vitamin E level and sunflower oil shown by a linear increase in the content of saturated fatty acids in the spermatozoa and a linear reduction in the C18?:?1ω9, C18?:?2ω6 and PUFA (C18?:?2ω6?+?C20?:?4ω6) contents in the spermatozoa at 48?–?51 weeks and in sperm volume at 52 weeks of age.     

Reproductive biology of Philodryas patagoniensis (Snakes: Dipsadidae) in south Brazil: Female reproductive cycle

In this study, we describe the female reproductive cycle of Philodryas patagoniensis in south Brazil, which was described through morpho‐anatomical and histological analyses. The peak of secondary vitellogenesis occurred during winter–spring (July–December), ovulation in spring (October–December), mating and fertilization in spring–summer (October–February), oviposition in spring–autumn (October–May) and births from late spring to autumn (December–July). The diameter of vitellogenic follicles/eggs was larger in winter–spring than in other seasons. The diameter of the shell glands was also larger in winter–spring. In spite of the clear reproductive peak, gonads only showed reduced activity in the autumn. Therefore, at the individual level, females have a discontinuous cyclical reproduction; in the populational level, the reproductive cycle is seasonal semisynchronous. We support the hypothesis that P. patagoniensis have the ability to produce multiple clutches with long‐term stored sperm. Sexual dimorphism in body size was evident, and females are significantly larger and heavier than males. Larger females were able to produce follicles and eggs in larger amount and size. The maternal body size was positively related to the reproductive effort and fecundity. To conclude, we deliberated about the proximal and distal causes that influence the reproductive traits and patterns of P. patagoniensis.     

Immunolocalization of secretogranin II, chromogranin A, and chromogranin B in differentiating human neuroblastoma cells.

In order to obtain further insights into the expression of the known markers of secretory neuroendocrine dense core organelles, secretogranin II (SgII), chromogranin A (CgA), and chromogranin B (CgB) during neuronal differentiation, the immunolocalization of these proteins was studied by means of double immunofluorescence in both undifferentiated and retinoic acid-differentiated SH-SY5Y human neuroblastoma cells. The majority of undifferentiated cells was not immunolabeled for all three proteins. In the majority of differentiated cells, a clearly punctate SgII immunolabeling indicative of the presence of secretory organelles was present in the Golgi region, at the cell periphery, along the neurites and in growth cones. Only relatively few of the SgII-immunolabeled cells were also immunolabeled for CgA and CgB, and in a single cell the three proteins were not always present in the same organelles. These results, obtained in a cultured cell line, confirm the not necessarily parallel distribution of SgII, CgA, and CgB observed in different neuroendocrine tissues and suggest that SgII may be the best marker of human neuroblastoma cell differentiation.     

Microhabitat and morphometric variation of two Chaetophoracean (Chaetophorales, Chlorophyta) species in tropical streams of southeastern Brazil

Two populations of Chaetophora elegans (Roth) C. Agardh and two of Stigeoclonium helveticum Vischer were investigated for microhabitat characteristics and morphological variation in streams of São Paulo State, southeastern Brazil. Different patterns of microhabitat distribution were found between species investigated. Populations of C, elegans were distributed under relatively narrow microhabitat conditions (high irradiance, low depth, moderate to high current velocity, rocky substrata and lower values of niche width) and showing little morphometric variation (colony diameter, main axis cell size, and apical branch number), Stigeoclonium helveticum occurred under more diverse microhabitat conditions, revealed by lack of significant difference between sampling units with and without the alga and wider niche width, but also exhibited relatively narrow morphometric variation (plant length, main axis cell and ateral branch cell sizes). The narrow microhabitat conditions and smaller niche width of C. elegans can explain its low abundance (percentage cover) in streams from the area studied as well as in other regions of São Paulo State, In contrast, the wider variation of microhabitat conditions and the higher niche widths of S. helveticum suggest that this green alga is able to grow in a high number of stream ecosystems in the region investigated, ranging from undisturbed to highly disturbed habitats. Thus. the results suggest that S. helveticum is a generalist species.     

Differentiation of mesenchymal stem cells derived from pancreatic islets and bone marrow into islet-like cell phenotype

Background

Regarding regenerative medicine for diabetes, accessible sources of Mesenchymal Stem Cells (MSCs) for induction of insular beta cell differentiation may be as important as mastering the differentiation process itself.

Methodology/Principal Findings

In the present work, stem cells from pancreatic islets (human islet-mesenchymal stem cells, HI-MSCs) and from human bone marrow (bone marrow mesenchymal stem cells, BM-MSCs) were cultured in custom-made serum-free medium, using suitable conditions in order to induce differentiation into Islet-like Cells (ILCs). HI-MSCs and BM-MSCs were positive for the MSC markers CD105, CD73, CD90, CD29. Following this induction, HI-MSC and BM-MSC formed evident islet-like structures in the culture flasks. To investigate functional modifications after induction to ILCs, ultrastructural analysis and immunofluorescence were performed. PDX1 (pancreatic duodenal homeobox gene-1), insulin, C peptide and Glut-2 were detected in HI-ILCs whereas BM-ILCs only expressed Glut-2 and insulin. Insulin was also detected in the culture medium following glucose stimulation, confirming an initial differentiation that resulted in glucose-sensitive endocrine secretion. In order to identify proteins that were modified following differentiation from basal MSC (HI-MSCs and BM-MSCs) to their HI-ILCs and BM-ILCs counterparts, proteomic analysis was performed. Three new proteins (APOA1, ATL2 and SODM) were present in both ILC types, while other detected proteins were verified to be unique to the single individual differentiated cells lines. Hierarchical analysis underscored the limited similarities between HI-MSCs and BM-MSCs after induction of differentiation, and the persistence of relevant differences related to cells of different origin.

Conclusions/Significance

Proteomic analysis highlighted differences in the MSCs according to site of origin, reflecting spontaneous differentiation and commitment. A more detailed understanding of protein assets may provide insights required to master the differentiation process of HI-MSCs to functional beta cells based only upon culture conditioning. These findings may open new strategies for the clinical use of BM-MSCs in diabetes.