In vitro culture of primary human myoblasts by using the dextran microcarriers Cytodex3®

Introduction. Primary cells in vitro culture scale-up is a crucial issue in cell-based tissue and organ regeneration therapy. Reducing costs and space occupied by the cells cultured in vitro has been an important target. Cells cultured in vitro with the use of bioreactor with dextran microcarriers (Cytodex®) have potentially a chance to meet many of the cell therapy requirements. Material and methods. We used collagen-coated carriers (Cytodex3®) and a spinner flask bioreactor to develop environment suitable for human myoblast proliferation. In parallel, standard adherent in vitro culture conditions for myoblasts propagation (T-flask) were conducted. Cell cycle characterization, senescence, myogenic gene ex­pression and cell apoptosis were evaluated in order to find differences between two culture systems under study. Results. The number of cells obtained in bioreactor per 106 of starting cells population was approximately ten times lower in comparison with T-flask culture system. The microcarriers cultured adult myoblasts in compari­son with the regular T-flask culture showed faster and more advanced replicative aging and lower proliferative potential. Moreover, the percentage of the cells that entailed an irreversible cell arrest (G0 phase) was also significantly (p < 0.0001) increased. Conclusions. Our results suggest that population of primary human myoblasts obtained from adult individuals and propagated on dextran microcarriers did not meet the requirements of the regenerative medicine regarding quantity and quality of the cells obtained. Nonetheless, further optimization of the cell scaling up process including both microcarriers and/or bioreactor program is still an important option

Volume modulus inflection point inflation and the gravitino mass problem

Several models of inflection point inflation with the volume modulus as the inflaton are investigated. Non-perturbative superpotentials containing two gaugino condensation terms or one such term with threshold corrections are considered. It is shown that the gravitino mass may be much smaller than the Hubble scale during inflation if at least one of the non-perturbative terms has a positive exponent. Higher order corrections to the Kahler potential have to be taken into account in such models. Those corrections are used to stabilize the potential in the axion direction in the vicinity of the inflection point. Models with only negative exponents require uplifting and in consequence have the supersymmetry breaking scale higher than the inflation scale. Fine-tuning of parameters and initial conditions is analyzed in some detail for both types of models. It is found that fine-tuning of parameters in models with heavy gravitino is much stronger than in models with light gravitino. It is shown that recently proposed time dependent potentials can provide a solution to the problem of the initial conditions only in models with heavy gravitino. Such potentials can not be used to relax fine tuning of parameters in any model because this would lead to values of the spectral index well outside the experimental bounds.Comment: 27 pages, 9 figures, comments and references added, version to be publishe

Inflation with racetrack superpotential and matter field

Several models of inflation with the racetrack superpotential for the volume modulus coupled to a matter field are investigated. In particular, it is shown that two classes of racetrack inflation models, saddle point and inflection point ones, can be constructed in a fully supersymmetric framework with the matter field F-term as a source of supersymmetry breaking and uplifting. Two models of F-term supersymmetry breaking are considered: the Polonyi model and the quantum corrected O'Raifeartaigh model. In the former case, both classes of racetrack inflation models differ significantly from the corresponding models with non-supersymmetric uplifting. The main difference is a quite strong dominance of the inflaton by the matter field. In addition, fine-tuning of the parameters is relaxed as compared to the original racetrack models. In the case of the racetrack inflation models coupled to the O'Raifeartaigh model, the matter field is approximately decoupled from the inflationary dynamics. In all of the above models the gravitino mass is larger than the Hubble scale during inflation. The possibility of having the gravitino much lighter than the Hubble scale is also investigated. It is very hard to construct models with light gravitino in which the volume modulus dominates inflation. On the other hand, models in which the inflationary dynamics is dominated by the matter field are relatively simple and seem to be more natural.Comment: 40 pages, 13 figures, references added, typos corrected, version to be publishe

Sterol biosensor reveals LAM-family Ltc1-dependent sterol flow to endosomes upon Arp2/3 inhibition.

Sterols are crucial components of biological membranes, which are synthetized in the ER and accumulate in the plasma membrane (PM). Here, by applying a genetically encoded sterol biosensor (D4H), we visualize a sterol flow between PM and endosomes in the fission yeast Schizosaccharomyces pombe. Using time-lapse and correlative light-electron microscopy, we found that inhibition of Arp2/3-dependent F-actin assembly promotes the reversible relocalization of D4H from the PM to internal sterol-rich compartments (STRIC) labeled by synaptobrevin Syb1. Retrograde sterol internalization to STRIC is independent of endocytosis or an intact Golgi, but depends on Ltc1, a LAM/StARkin-family protein localized to ER-PM contact sites. The PM in ltc1Δ cells over-accumulates sterols and upon Arp2/3 inhibition forms extended ER-interacting invaginations, indicating that sterol transfer contributes to PM size homeostasis. Anterograde sterol movement from STRIC is independent of canonical vesicular trafficking but requires Arp2/3, suggesting a novel role for this complex. Thus, transfer routes orthogonal to vesicular trafficking govern the flow of sterols in the cell

Bacterial diversity in snow on North Pole ice floes

The microbial abundance and diversity in snow on ice floes at three sites near the North Pole was assessed using quantitative PCR and 454 pyrosequencing. Abundance of 16S rRNA genes in the samples ranged between 43 and 248 gene copies per millilitre of melted snow. A total of 291,331 sequences were obtained through 454 pyrosequencing of 16S rRNA genes, resulting in 984 OTUs at 97 % identity. Two sites were dominated by Cyanobacteria (72 and 61 %, respectively), including chloroplasts. The third site differed by consisting of 95 % Proteobacteria. Principal component analysis showed that the three sites clustered together when compared to the underlying environments of sea ice and ocean water. The Shannon indices ranged from 2.226 to 3.758, and the Chao1 indices showed species richness between 293 and 353 for the three samples. The relatively low abundances and diversity found in the samples indicate a lower rate of microbial input to this snow habitat compared to snow in the proximity of terrestrial and anthropogenic sources of microorganisms. The differences in species composition and diversity between the sites show that apparently similar snow habitats contain a large variation in biodiversity, although the differences were smaller than the differences to the underlying environment. The results support the idea that a globally distributed community exists in snow and that the global snow community can in part be attributed to microbial input from the atmosphere. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00792-014-0660-y) contains supplementary material, which is available to authorized users

Gene banks for wild and cultivated sunflower genetic resources

Modern breeding of sunflower (Helianthus annuus L.), which started 100 years ago, increased the number and the diversity of cultivated forms. In addition, for more than 50 years, wild sunflower and other Helianthus species have been collected in North America where they all originated. Collections of both cultivated and wild forms are maintained in gene banks in many countries where sunflower is an important crop, with some specificity according to the availability of germplasm and to local research and breeding programmes. Cultivated material includes land races, open pollinated varieties, synthetics and inbred lines. The majority of wild accessions are ecotypes of wild Helianthus annuus, but also 52 other species of Helianthus and a few related genera. The activities of three gene banks, in USA, France and Serbia, are described in detail, supplemented by data from seven other countries. Past and future uses of the genetic resources for environmental adaptation and breeding are discussed in relation to genomic and improved phenotypic knowledge of the cultivated and wild accessions available in the gene banks.L’amélioration moderne du tournesol (Helianthus annuus L.) a débuté il y a un siècle, diversifiant et augmentant le nombre des formes cultivées du tournesol. De plus, des collectes de tournesols sauvages et d’espèces du genre Helianthus ont lieu depuis 50 ans en Amérique du Nord d’où ils sont tous originaires. Ainsi, des collections de tournesols cultivés et sauvages sont conservées par des centres de ressources génétiques dans de nombreux pays où le tournesol est une culture importante. Chacun d’eux présente des spécificités par rapport aux ressources génétiques maintenues, en fonction des programmes de recherche ou de sélection variétale locales. Le matériel génétique cultivé comprend des écotypes, des populations et des lignées tandis que les accessions sauvages correspondent eux écotypes d’Helianthus annuus sauvages et des 52 autres espèces apparentées du genre Helianthus. Les activités de trois centres de ressources génétiques des États-Unis, de la France et de la Serbie sont décrites en détail, complétées par des données provenant des centres de sept autres pays. L’historique de l’utilisation des ressources génétiques et les perspectives futures pour l’adaptation des variétés à l’environnement sont discutés ainsi que leur caractérisation au niveau génomique et phénotypique.The Supplementary Material is available at [https://www.ocljournal.org/10.1051/ocl/2020004/olm]

RTX proteins: a highly diverse family secreted by a common mechanism

Repeats-in-toxin (RTX) exoproteins of Gram-negative bacteria form a steadily growing family of proteins with diverse biological functions. Their common feature is the unique mode of export across the bacterial envelope via the type I secretion system and the characteristic, typically nonapeptide, glycine- and aspartate-rich repeats binding Ca2+ ions. In this review, we summarize the current state of knowledge on the organization of rtx loci and on the biological and biochemical activities of therein encoded proteins. Applying several types of bioinformatic screens on the steadily growing set of sequenced bacterial genomes, over 1000 RTX family members were detected, with the biological functions of most of them remaining to be characterized. Activities of the so far characterized RTX family members are then discussed and classified according to functional categories, ranging from the historically first characterized pore-forming RTX leukotoxins, through the large multifunctional enzymatic toxins, bacteriocins, nodulation proteins, surface layer proteins, up to secreted hydrolytic enzymes exhibiting metalloprotease or lipase activities of industrial interest

GrassPlot - a database of multi-scale plant diversity in Palaearctic grasslands

GrassPlot is a collaborative vegetation-plot database organised by the Eurasian Dry Grassland Group (EDGG) and listed in the Global Index of Vegetation-Plot Databases (GIVD ID EU-00-003). GrassPlot collects plot records (releves) from grasslands and other open habitats of the Palaearctic biogeographic realm. It focuses on precisely delimited plots of eight standard grain sizes (0.0001; 0.001;... 1,000 m(2)) and on nested-plot series with at least four different grain sizes. The usage of GrassPlot is regulated through Bylaws that intend to balance the interests of data contributors and data users. The current version (v. 1.00) contains data for approximately 170,000 plots of different sizes and 2,800 nested-plot series. The key components are richness data and metadata. However, most included datasets also encompass compositional data. About 14,000 plots have near-complete records of terricolous bryophytes and lichens in addition to vascular plants. At present, GrassPlot contains data from 36 countries throughout the Palaearctic, spread across elevational gradients and major grassland types. GrassPlot with its multi-scale and multi-taxon focus complements the larger international vegetationplot databases, such as the European Vegetation Archive (EVA) and the global database " sPlot". Its main aim is to facilitate studies on the scale-and taxon-dependency of biodiversity patterns and drivers along macroecological gradients. GrassPlot is a dynamic database and will expand through new data collection coordinated by the elected Governing Board. We invite researchers with suitable data to join GrassPlot. Researchers with project ideas addressable with GrassPlot data are welcome to submit proposals to the Governing Board