Percolation and diffusion in three-component lipid bilayers: effect of cholesterol on an equimolar mixture of two phosphatidylcholines.

The lateral diffusion of a phospholipid probe is studied in bilayers of binary mixtures of dimyristoylphosphatidylcholine (DMPC)/cholesterol and distearoylphosphatidylcholine (DSPC)/cholesterol and in the ternary system DMPC/DSPC/cholesterol using fluorescence recovery after photobleaching. An approximate phase diagram for the ternary system, as a function of temperature and cholesterol concentration, was obtained using differential scanning calorimetry and the phase diagrams of the binary systems. This phase diagram is similar to those of the phospholipid/cholesterol binary mixtures. In bilayers where solid and liquid phases coexist, the diffusion results are interpreted in terms of phase percolation. The size of the liquid-phase domains is estimated using percolation theory. In the ternary system, addition of cholesterol up to approximately 20 mol% shifts the percolation threshold to lower area fractions of liquid, but the size of the liquid-phase domains does not change. Above approximately 20 mol% cholesterol, the liquid phase is always connected. The size of solid-phase domains clusters is estimated using a model recently developed (Almeida, P.F.F., W.L.C. Vaz, and T.E. Thompson. 1992. Biochemistry. 31:7198-7210). For cholesterol concentrations up to 20 mol%, the size of solid-phase domain units does not change. Beyond 20 mol%, cholesterol causes the size of the solid units to decrease.     

Ecological integrity of tropical secondary forests: concepts and indicators

Naturally regenerating forests or secondary forests (SFs) are a promising strategy for restoring large expanses of tropical forests at low cost and with high environmental benefits. This expectation is supported by the high resilience of tropical forests after natural disturbances, yet this resilience can be severely reduced by human impacts. Assessing the characteristics of SFs and their ecological integrity (EI) is essential to evaluating their role for conservation, restoration, and provisioning of ecosystem services. In this study, we aim to propose a concept and indicators that allow the assessment and classification of the EI of SFs. To this end, we review the literature to assess how EI has been addressed in different ecosystems and which indicators of EI are most commonly used for tropical forests. Building upon this knowledge we propose a modification of the concept of EI to embrace SFs and suggest indicators of EI that can be applied to different successional stages or stand ages. Additionally, we relate these indicators to ecosystem service provision in order to support the practical application of the theory. EI is generally defined as the ability of ecosystems to support and maintain composition, structure and function similar to the reference conditions of an undisturbed ecosystem. This definition does not consider the temporal dynamics of recovering ecosystems, such as SFs. Therefore, we suggest incorporation of an optimal successional trajectory as a reference in addition to the old-growth forest reference. The optimal successional trajectory represents the maximum EI that can be attained at each successional stage in a given region and enables the evaluation of EI at any given age class. We further suggest a list of indicators, the main ones being: compositional indicators (species diversity/richness and indicator species); structural indicators (basal area, heterogeneity of basal area and canopy cover); function indicators (tree growth and mortality); and landscape proxies (landscape heterogeneity, landscape connectivity). Finally, we discuss how this approach can assist in defining the value of SF patches to provide ecosystem services, restore forests and contribute to ecosystem conservation.     

Meiotic abnormalities in sugarcane (Saccharum spp.) and parental species: Evidence for peri- and paracentric inversions

The modern cultivars of sugarcane (Saccharum spp.) are highly polyploid and accumulate aneuploidies due to their history of domestication, genetic improvement and interspecific hybrid origin involving the domesticated sweet species Saccharum officinarum (‘noble cane’) and the wild Saccharum spontaneum, both with an evolutionary history of polyploidy. The first hybrids were backcrossed with S. officinarum, and selection from progenies in subsequent generations established the genetic basis of modern cultivars. Saccharum genome complexity has inspired several molecular studies that have elucidated aspects of sugarcane genome constitution, architecture and cytogenetics. Herein, we conducted a comparative analysis of the meiotic behaviour of representatives of the parentals S. officinarum and S. spontaneum, and the commercial variety, SP80-3280. S. officinarum, an octoploid species, exhibited regular meiotic behaviour. In contrast, S. spontaneum and SP80-3280 exhibited several abnormalities from metaphase I to the end of division. We reported and typified, for the first time, the occurrence of peri- and paracentric inversions. Using in-situ hybridisation techniques, we were able to determine how pairing association occurred at diakinesis, the origin of lagging chromosomes and, in particular, the mitotic chromosome composition of SP80-3280. Interestingly, S. spontaneum and recombinant chromosomes showed the most marked tendency to produce laggards in both divisions. Future attempts to advance knowledge on sugarcane genetics and genomics should take meiotic chromosome behaviour information into account.     

Hypomethylation of classical satellite DNA and chromosome instability in lymphoblastoid cell lines

To determine possible relationships between DNA hypomethylation and chromosome instability, human lymphoblastoid cell lines from different genetic constitutions were studied with regard to 1) uncoiling and rearrangements, which preferentially affect the heterochromatic segments of chromosomes 1 and 16; 2) the methylation status of the tandemly repetitive sequences (classical satellite and alphoid DNAs) from chromosomes 1 and 16, and of the L1Hs interspersed repetitive sequences. The methylation status largely varied from cell line to cell line, but for a given cell line, the degree of methylation was similar for all the repetitive DNAs studied. Two cell lines, one obtained from a Fanconi anemia patient and the other from an ataxia telangiectasia patient were found to be heavily hypomethylated. The heterochromatic segments of their chromosomes 1 and 16 were more frequently elongated and rearranged than those from other cell lines, which were found to be less hypomethylated. Thus, in these lymphoblastoid cell lines, alterations characterized by uncoiling and rearrangements of heterochromatic segments from chromosomes 1 and 16 seem to correlate with the hypomethylation of their repetitive DNAs. Two-color in situ hybridizations demonstrated that these elongations and rearrangements involved only classical satellite-DNA-containing heterochromatin. This specificity may be related to the excess of breakages affecting the chromosomes carrying these structures in a variety of pathological conditions.     

Alterations of bovine nucleus pulposus cells with aging

Aging is one of the major etiological factors driving intervertebral disc (IVD) degeneration, the main cause of low back pain. The nucleus pulposus (NP) includes a heterogeneous cell population, which is still poorly characterized. Here, we aimed to uncover main alterations in NP cells with aging. For that, bovine coccygeal discs from young (12 months) and old (10–16 years old) animals were dissected and primary NP cells were isolated. Gene expression and proteomics of fresh NP cells were performed. NP cells were labelled with propidium iodide and analysed by flow cytometry for the expression of CD29, CD44, CD45, CD146, GD2, Tie2, CD34 and Stro-1. Morphological cell features were also dissected by imaging flow cytometry. Elder NP cells (up-regulated bIL-6 and bMMP1 gene expression) presented lower percentages of CD29+, CD44+, CD45+ and Tie2+ cells compared with young NP cells (upregulated bIL-8, bCOL2A1 and bACAN gene expression), while GD2, CD146, Stro-1 and CD34 expression were maintained with age. NP cellulome showed an upregulation of proteins related to endoplasmic reticulum (ER) and melanosome independently of age, whereas proteins upregulated in elder NP cells were also associated with glycosylation and disulfide bonds. Flow cytometry analysis of NP cells disclosed the existence of 4 subpopulations with distinct auto-fluorescence and size with different dynamics along aging. Regarding cell morphology, aging increases NP cell area, diameter and vesicles. These results contribute to a better understanding of NP cells aging and highlighting potential anti-aging targets that can help to mitigate age-related disc disease.     

Mitochondrial growth and division during the cell cycle in HeLa cells

The growth and division of mitochondria during the cell cycle was investigated by a morphometric analysis of electron micrographs of synchronized HeLa cells. The ratio of total outer membrane contour length to cytoplasmic area did not vary significantly during the cell cycle, implying a continuous growth of the mitochondrial outer membrane. The mean fraction of cytoplasmic area occupied by mitochondrial profiles was likewise found to remain constant, indicating that the increase in total mitochondrial volume per cell occurs continuously during interphase, in such a way that the mitochondrial complement occupies a constant fraction( approximately 10-11(percent)) of the volume of the cytoplasm. The mean area, outer membrane contour length, and axis ratio of the mitochondrial profiles also did not vary appreciably during the cell cycle; furthermore, the close similarity of the frequency distributions of these parameters for the six experimental time-points suggested a stable mitochondrial shape distribution. The constancy of both the mean mitochondrial profile area and the number of mitochondrial profiles per unit of cytoplasmic area was interpreted to indicate the continuous division of mitochondria at the level of the cell population. Furthermore, no evidence was found for the occurrence of synchronous mitochondrial growth and division within individual cells. Thus, it appears that, in HeLa cells, there is no fixed temporal relationship between the growth and division of mitochondria and the events of the cell cycle. A number of statistical methods were developed for the purpose of making numerical estimates of certain three-dimensional cellular and mitochondrial parameters. Mean cellular and cytoplasmic volumes were calculated for the six time-points; both exhibited a nonlinear, approx. twofold increase. A comparison of the axis ratio distributions of the mitochondrial profiles with theoretical distributions expected from random sectioning of bodies of various three-dimensional shapes allowed the derivation of an "average" mitochondrial shape. This, in turn, permitted calculations to be made which expressed the two-dimensional results in three-dimensional terms. Thus, the estimated values for the number of mitochondria per unit of cytoplasmic volume and for the mean mitochondrial volume were found to remain constant during the cell cycle, while the estimated number of mitochondria per cell increase approx. twofold in an essentially continuous manner.     

Microtubules and cyclic amp in human leukocytes: on the order of things

We have shown previously that the β-adrenergic agonist isoproterenol (2μM) and the phosphodiesterase inhibitor isobutylmethylxanthine (1 mM) produce a much greater increase in cyclic AMP in human leukocytes that have been pretreated with colchicine (or with other agents that affect microtubule assembly) than in control leukocytes. The effects of colchicines were both time- and dose-dependant. These and other data suggested that the generation of cyclic AMP is normally restricted by an intact system of cytoplasmic microtubules. If so, then the same time and dose dependencies might apply to other colchicines-induced changes in leukocyte function. We have now assayed the distribution of concanavalin A (Con A)-receptor complexes on the leukocyte membrane, taking into account that leukocytes competent to assemble microtubules show a uniform distribution of surface- bound Con A whereas microtubule-deficient cells accumulate Con A in surface caps. We have found that the effect of colchicine on capping is also both time- and dose dependent, and that the dose-response relationships conform to those required to increase cyclic AMP levels. These findings provide further evidence that both colchicine-induced Con-A capping and colchicine- induced cyclic AMP generation depend upon the relaxation of constraints normally imposed by cytoplasmic microtubules upon the plasma membrane, which limit, respectively, lateral mobility of the lectin-receptor complexes, and expression of hormone-sensitive adenylate cyclase. Moreover, colchicine-induced Con-A cap formation is not affected even by very large changes in leukocyte cyclic AMP levels. Thus, elevated cyclic AMP levels do not appear to promote the dissolution of microtubules; rather, the dissolution of microtubules permits the generation of increased amounts of cyclic AMP.     

Chromosome banding in Amphibia. XV. Two types of Y chromosomes and heterochromatin hypervariabilty inGastrotheca pseustes (Anura,Hylidae)

The chromosomes of the newly discovered South American marsupial frogGastrotheca pseustes were analyzed by conventional methods and by various banding techniques. This species is characterized by XY/XX sex chromosomes and the existence of two different morphs of Y chromosomes. Whereas in type A males the XY^A chromosomes are still homomorphic, in type B males the Y^B chromosome displays a large heterochromatic region at the long arm telomere which is absent in the X. In male meiosis, the homomorphic XY^A chromosomes exhibit the same pairing configuration as the autosomal bivalents. On the other hand, the heteromorphic XY^B chromosomes form a sex bivalent by pairing their short arm telomeres in a characteristic end-to-end arrangement. Analysis of the karyotypes by C-banding and DNA base pair-specific fluorochromes reveals enormous interindividual size variability of the autosomal heterochromatin.