A Pillar-Based High-Throughput Myogenic Differentiation Assay to Assess Drug Safety

High-throughput, pillar-strip-based assays have been proposed as a drug-safety screening tool for developmental toxicity. In the assay described here, muscle cell culture and differentiation were allowed to occur at the end of a pillar strip (eight pillars) compatible with commercially available 96-well plates. Previous approaches to characterize cellular differentiation with immunostaining required a burdensome number of washing steps; these multiple washes also resulted in a high proportion of cellular loss resulting in poor yield. To overcome these limitations, the approach described here utilizes cell growth by easily moving the pillars for washing and immunostaining without significant loss of cells. Thus, the present pillar-strip approach is deemed suitable for monitoring high-throughput myogenic differentiation. Using this experimental high-throughput approach, eight drugs (including two well-known myogenic inhibitory drugs) were tested at six doses in triplicate, which allows for the generation of dose–response curves of nuclei and myotubes in a 96-well platform. As a result of comparing these F-actin (an actin-cytoskeleton protein), nucleus, and myotube data, two proposed differentiation indices—curve-area-based differentiation index (CA-DI) and maximum-point-based differentiation index (MP-DI) were generated. Both indices successfully allowed for screening of high-myogenic inhibitory drugs, and the maximum-point-based differentiation index (MP-DI) experimentally demonstrated sensitivity for quantifying drugs that inhibited myogenic differentiation.

Comparison between QST and ΦST indices in an endangered Boswellia serrata Roxb: Implications for conservation

Boswellia serrata, an economically important indigenous tree of dry deciduous forests, provides oleoresin gum of pharmaceutical significance and excellent pulp for paper industries, but faces threat to extinction due to poor natural regeneration and commercial exploitation. 240 individuals of the species representing 12 locations of its natural distribution in central India were investigated to compare the genetic differentiation indices, QST for GBH and wood fiber length and ϕST for neutral (RAPD+ISSR) markers. The comparison for paired locations was more informative than for metapopulation. The most paired locations were either under the stabilizing selection (QST (L) < ΦST (L)) or in the genetic drift (QST(L) = ΦST (L)) whereas a relatively small number of paired locations was under the divergent selection (QS T(L) > ΦST (L)). The comparison for the metapopulation generating only a single trend of QST (P) > ΦST (P) is, therefore, misleading. For conservation, the genetically deficit locations (QST (L) < ΦST (L) and QST (L) = ΦST (L)) of B. serrata warrant for reinforcement of their genetic diversity by introduction of genotypes from other genetically divergent locations (QST (L) > ΦST (L)), which would check the fragmentation and genetic drift, resulting in reproductive vigour, natural regeneration and reverse the endangered status of the species.

Influence of historical events and contemporary estuarine circulation on the genetic structure of the banded killifish (Fundulus diaphanus) in the St. Lawrence River (Quebec, Canada)

Eight microsatellite markers were used to examine the historical and contemporary factors influencing the distribution of genetic variation within and among populations of Fundulus diaphanus (Lesueur, 1817) in the St. Lawrence River. Our results show that the contemporary hydrodynamics of the river affect levels of diversity and differentiation in this species. Genetic diversity increased towards downstream sites and levelled off at the upstream end of the tidal section, in the vicinity of Lake St-Pierre outlet. Likewise, differentiation and isolation-by-distance were most pronounced in the upstream, strictly fluvial section of the river. Surprisingly, however, we did not detect any significant intershore differentiation in this riparian fish species. Historical influence was also clearly detected and transcended the contemporary pattern, as revealed by an extremely strong isolation-by-distance pattern along the entire river stretch. This pattern likely results from the historical, postglacial secondary contact between two glacial races, as indicated by the clinal variation of constrasting allele size frequency distributions along the river and the significantly greater values of differentiation indices considering mutational information.