Identification of senescent cells in multipotent mesenchymal stromal cell cultures: Current methods and future directions

Regardless of their tissue of origin, multipotent mesenchymal stromal cells (MSCs) are commonly expanded in vitro for several population doublings to achieve a sufficient number of cells for therapy. Prolonged MSC expansion has been shown to result in phenotypical, morphological and gene expression changes in MSCs, which ultimately lead to the state of senescence. The presence of senescent cells in therapeutic MSC batches is undesirable because it reduces their viability, differentiation potential and trophic capabilities. Additionally, senescent cells acquire senescence-activated secretory phenotype, which may not only induce apoptosis in the neighboring host cells following MSC transplantation, but also trigger local inflammatory reactions. This review outlines the current and promising new methodologies for the identification of senescent cells in MSC cultures, with a particular emphasis on non-destructive and label-free methodologies. Technologies allowing identification of individual senescent cells, based on new surface markers, offer potential advantage for targeted senescent cell removal using new-generation senolytic agents, and subsequent production of therapeutic MSC batches fully devoid of senescent cells. Methods or a combination of methods that are non-destructive and label-free, for example, involving cell size and spectroscopic measurements, could be the best way forward because they do not modify the cells of interest, thus maximizing the final output of therapeutic-grade MSC cultures. The further incorporation of machine learning methods has also recently shown promise in facilitating, automating and enhancing the analysis of these measured data.     

Automated binning of microsatellite alleles: problems and solutions

As genotyping methods move ever closer to full automation, care must be taken to ensure that there is no equivalent rise in allele‐calling error rates. One clear source of error lies with how raw allele lengths are converted into allele classes, a process referred to as binning. Standard automated approaches usually assume collinearity between expected and measured fragment length. Unfortunately, such collinearity is often only approximate, with the consequence that alleles do not conform to a perfect 2‐, 3‐ or 4‐base‐pair periodicity. To account for these problems, we introduce a method that allows repeat units to be fractionally shorter or longer than their theoretical value. Tested on a large human data set, our algorithm performs well over a wide range of dinucleotide repeat loci. The size of the problem caused by sticking to whole numbers of bases is indicated by the fact that the effective repeat length was within 5% of the assumed length only 68.3% of the time.     

A multiplex set of microsatellite markers for the scarlet rosefinch (Carpodacus erythrinus)

Cardueline finches have become important models in studies of sexual selection and evolution of carotenoid‐based ornamentation. Here, we describe eight new polymorphic microsatellites isolated from the Scarlet rosefinch (Carpodacus erythrinus) and four from the House finch (Carpodacus mexicanus). Together with the cross‐species amplification of additional loci, originally published for two species of songbirds, we optimized a multiplex panel for C. erythrinus allowing genotyping of 22 polymorphic loci. Number of alleles and heterozygosity per locus in a sample of 34 individuals ranged from three to 38 and from 0.27 to 0.94, respectively.     

Assessment of diet choice by the yellow fever mosquito Aedes aegypti

To investigate feeding‐related decisions in Aedes aegypti (L.), adults are presented with simple diets of paired gustatory stimuli conveying information concerning energy content, nutrient richness, osmotic balance and food toxicity in a two‐diet matrix assay. Assessment of mosquito gut contents indicates that both sexes accept single sugar diets in a dose‐dependent manner. When presented with a choice between two different yet equimolar sugar solutions, more individuals of both sexes accept the disaccharides, sucrose and trehalose, than the monosacharrides, fructose and glucose. The combination of pyranose and furanose sugars in solution, either physically associated (as in sucrose) or present as monomers (as glucose and fructose), is accepted over solutions containing a single sugar moiety. Using the two‐diet matrix assay, mosquito diet‐choice is also tested between two equimolar sucrose ‘driver’ solutions in which one is presented with various concentrations of another potential feeding cue ‘test’ compound (i.e. each of the 20 naturally‐occurring amino acids, sodium chloride, quinine or caffeine). Diet‐choice between the ‘driver’ sucrose‐only solution and the solution of the ‘driver’ sucrose containing a ‘test’ amino acid is influenced by sex, amino acid concentration and sucrose concentration. There is also an example of synergism between the diet components, leucine and sucrose. Mosquitoes demonstrate a dose‐dependent acceptance of sucrose‐only diets over sodium chloride‐containing sucrose when presented together. Interestingly, the sucrose‐only diet is accepted by more mosquitoes than all concentrations of the saline‐containing sucrose diets except those approximately isotonic to mosquito haemolymph, at which concentration mosquitoes show no clear choice between the diets. More individuals of both sexes accept sucrose‐only diets than the diets of caffeine‐containing sucrose in a dose‐dependent manner. Only females, however, respond to quinine‐containing sucrose diets and modulate this behaviour in relation to the energetic reward: more females imbibed quinine‐containing sucrose at the higher sucrose concentration (1 m ). A systematic characterization of diet selection behaviour of A. aegypti is presented for 27 putative feeding cues potentially involved in nectar/honeydew feeding. This study will be used as a basis from which to investigate further the mosquito's assessment of food quality and ultimately host choice.     

Can filter‐feeding Asian carp invade the Laurentian Great Lakes? A bioenergetic modelling exercise

1. There is much concern that filter‐feeding Asian carp will invade the Laurentian Great Lakes and deplete crucial plankton resources. We developed bioenergetic models, using parameters from Asian carp and other fish species, to explore the possibility that planktonic food resources are insufficient to support the growth of silver carp (Hypophthalmichthys molitrix) and bighead carp (H. nobilis) in the Great Lakes. 2. The models estimated basic metabolic requirements of silver and bighead carp under various body sizes, swimming speeds and reproductive stages. These requirements were then related to planktonic food resources and environmental temperature to predict when and where silver and bighead carp may survive in the Great Lakes, and how far they may travel. 3. Parameter values for respiration functions were derived experimentally in a coordinated study of silver and bighead carp, while consumption parameters were obtained from the literature on silver carp. Other model parameters lacking for Asian carp, such as those for egestion and excretion, were obtained from the literature on other fish species. 4. We found that full‐sized bighead carp required 61.0 kJ d^?1 just to maintain their body mass at 20 °C, approximately equivalent to feeding in a region with 255 μg L^?1 macrozooplankton (dry) or 10.43 μg L^?1 chlorophyll a. Silver carp energy requirements were slightly higher. 5. When applied to various habitats in the Great Lakes, our results suggest that silver and bighead carp will be unable to colonise most open‐water regions because of limited plankton availability. However, in some circumstances, carp metabolism at lower temperatures may be low enough to permit positive growth even at very low rations. Positive growth is even more likely in productive embayments and wetlands, and the modelled swimming costs in some of these habitats suggest that carp could travel >1 km d^?1 without losing biomass. 6. The simulation (and firmly hypothetical) results from this modelling study suggest when and where Asian carp could become established in the Great Lakes. Given the potential consequences to Great Lakes ecosystems if these filter feeders do prove capable of establishing reproducing populations, efforts to keep Asian carp out of the Great Lakes must not be lessened. However, we do encourage the use of bioenergetic modelling in a holistic approach to assessing the risk of Asian carp invasion in the Great Lakes region.     

The scales of salticid spiders

The form and location of the scales of salticid spiders, as revealed by light and scanning electron microscopy, provide useful diagnostic characters for the separation of species, the assignment of species to genera, and a further understanding of the relationships between salticid genera. Partly on the basis of distinctions provided by scale structure, the new genus Platycryptus , type-species Aranea undata De Geer, 1778, is defined. Metaphidippus vitis Cockerell, 1894, is placed in the genus Sassacus Peckham, 1895, and Paromaevia michelsoni (Barnes), 1955, is returned to the genus Maevia . The placement of Eris, Hentzia, Icius, Metaphidippus, Phidippus , and Sassacus in the subfamily Dendryphantinae is substantiated; Tutelina and Zygoballus are added to this group. On the basis of common scale structure, Evarcha, Habrocestum, Menememerus, Phlegra, Platycryptus , and Sitticus are tentatively assigned to the subfamily Habrocestinae. Scales of Anasntis, Corythalia, Cosmophasis, Hyllus, Marpissa, Metacyrba, Pellenes, Plexippus, Salticus, Sarinda, and Thiodina are also described.     

Pollen source affects female reproductive success and early offspring traits in the rare endemic plant Polemonium vanbruntiae (Polemoniaceae)

Understanding the relative magnitudes of inbreeding and outbreeding depression in rare plant populations is increasingly important for effective management strategies. There may be positive and negative effects of crossing individuals in fragmented populations. Conservation strategies may include introducing new genetic material into rare plant populations, which may be beneficial or detrimental based on whether hybrid offspring are of increased or decreased quality. Thus, it is important to determine the effects of pollen source on offspring fitness in rare plants. We established pollen crosses (i.e. geitonogamous‐self, autonomous‐self, intrasite‐outcross, intersite‐outcross and open‐pollinated controls) to determine the effects of pollen source on fitness (seeds/fruit and seed mass) and early offspring traits (probability of germination, number of leaves, leaf area and seedling height) in the rare plant Polemonium vanbruntiae. Open‐pollinated, intrasite‐outcross and geitonogamous‐self treatments did not differ in fitness. However, plants receiving autonomous‐self pollen had the lowest fitness and the lowest probability of seed germination. Intersite‐outcross plants contained fewer seeds/fruit, but seeds germinated at higher frequencies and seedlings were more vigorous. We also detected heterosis at the seed germination stage. These data may imply that natural populations of P. vanbruntiae exhibit low genetic variation and little gene flow. Evidence suggests that deleterious alleles were not responsible for reduced germination; rather environmental factors, dichogamy, herkogamy and/or lack of competition among pollen grains may have caused low germinability in selfed offspring. Although self‐pollination may provide some reproductive assurance in P. vanbruntiae, the result is a reduction in germination and size‐related early traits for selfed offspring.