An integrated nano-scale approach to profile miRNAs in limited clinical samples

Profiling miRNA expression in cells that directly contribute to human disease pathogenesis is likely to aid the discovery of novel drug targets and biomarkers. However, tissue heterogeneity and the limited amount of human diseased tissue available for research purposes present fundamental difficulties that often constrain the scope and potential of such studies. We established a flow cytometry-based method for isolating pure populations of pathogenic T cells from bronchial biopsy samples of asthma patients, and optimized a high-throughput nano-scale qRT-PCR method capable of accurately measuring 96 miRNAs in as little as 100 cells. Comparison of circulating and airway T cells from healthy and asthmatic subjects revealed asthma-associated and tissue-specific miRNA expression patterns. These results establish the feasibility and utility of investigating miRNA expression in small populations of cells involved in asthma pathogenesis, and set a precedent for application of our nano-scale approach in other human diseases. The microarray data from this study (Figure 7) has been submitted to the NCBI Gene Expression Omnibus (GEO; http://ncbi.nlm.nih.gov/geo) under accession no. GSE31030.

The influence of nano-scale second-phase particles on deformation of fine grained calcite mylonites

Grey and white carbonate mylonites were collected along thrust planes of the Helvetic Alps. They are characterised by very small grain sizes and non-random grain shape (SPO) and crystallographic preferred orientation (CPO). Presumably they deformed in the field of grain size sensitive flow by recrystallisation accommodated intracrystalline deformation in combination with granular flow. Both mylonites show a similar mean grain size, but in the grey mylonites the grain size range is larger, the grain shapes are more elongate and the dynamically recrystallised calcite grains are more often twinned. Grey mylonites have an oblique CPO, while the CPO in white mylonites is symmetric with respect to the shear plane. Combustion analysis and TEM investigations revealed that grey mylonites contain a higher amount of highly structured kerogens with particle sizes of a few tens of nanometers, which are finely dispersed at the grain boundaries. During deformation of the rock, nano-scale particles reduced the migration velocity of grain boundaries by Zener drag resulting in slower recrystallisation rates of the calcite aggregate. In the grey mylonites, more strain increments were accommodated by individual grains before they became refreshed by dynamic recrystallisation than in white mylonites, where grain boundary migration was less hindered and recrystallisation cycles were faster. Consequently, grey mylonites represent ‘deformation’ microfabrics while white mylonites are characterised by ‘recrystallisation’ microfabrics. Field geologists must utilise this different deformation behavior when applying the obliquity in CPO and SPO of the respective mylonites as reliable shear sense indicators.

Implementing the plasma-lasing potential for tabletop nano-imaging

Implementing the plasma-lasing potential for tabletop nano-imaging on across a hot plasma medium drives short-wavelength lasing, promising for "turnkey" nano-imaging setups. A systematic study of the illumination characteristics, combined with design-adapted objectives, is presented. It is shown how the ultimate nano-scale feature is dictated by either the diffraction-limited or the wavefront-limited resolution, which imposed a combined study of both the source and the optics. For nano-imaging, the spatial homogeneity of the illumination (spot noise) was shown as critical. Plasma-lasing from a triple grazing-incidence pumping scheme compensated for the missing spot homogeneity in classical schemes. We demonstrate that a collimating mirror pre-conditions both the pointing stability and the divergence below half a mrad.

Grain coarsening in contact metamorphic carbonates: effects of second-phase particles, fluid flow and thermal perturbations

Under contact metamorphic conditions, carbonate rocks in the direct vicinity of the Adamello pluton reflect a temperature-induced grain coarsening. Despite this large-scale trend, a considerable grain size scatter occurs on the outcrop-scale indicating local influence of second-order effects such as thermal perturbations, fluid flow and second-phase particles. Second-phase particles, whose sizes range from nano- to the micron-scale, induce the most pronounced data scatter resulting in grain sizes too small by up to a factor of 10, compared with theoretical grain growth in a pure system. Such values are restricted to relatively impure samples consisting of up to 10 vol.% micron-scale second-phase particles, or to samples containing a large number of nano-scale particles. The obtained data set suggests that the second phases induce a temperature-controlled reduction on calcite grain growth. The mean calcite grain size can therefore be expressed in the form D 1⁄4 C2 eQ*/RT(dp/fp)m*, where C2 is a constant, Q* is an activation energy, T the temperature and m* the exponent of the ratio dp/fp, i.e. of the average size of the second phases divided by their volume fraction. However, more data are needed to obtain reliable values for C2 and Q*. Besides variations in the average grain size, the presence of second-phase particles generates crystal size distribution (CSD) shapes characterized by lognormal distributions, which differ from the Gaussian-type distributions of the pure samples. In contrast, fluid-enhanced grain growth does not change the shape of the CSDs, but due to enhanced transport properties, the average grain sizes increase by a factor of 2 and the variance of the distribution increases. Stable d18O and d13C isotope ratios in fluid-affected zones only deviate slightly from the host rock values, suggesting low fluid/rock ratios. Grain growth modelling indicates that the fluid-induced grain size variations can develop within several ka. As inferred from a combination of thermal and grain growth modelling, dykes with widths of up to 1 m have only a restricted influence on grain size deviations smaller than a factor of 1.1.To summarize, considerable grain size variations of up to one order of magnitude can locally result from second-order effects. Such effects require special attention when comparing experimentally derived grain growth kinetics with field studies.

Differences in grain growth of calcite: a field-based modeling approach

Normal grain growth of calcite was investigated by combining grain size analysis of calcite across the contact aureole of the Adamello pluton, and grain growth modeling based on a thermal model of the surroundings of the pluton. In an unbiased model system, i.e., location dependent variations in temperature-time path, 2/3 and 1/3 of grain growth occurs during pro- and retrograde metamorphism at all locations, respectively. In contrast to this idealized situation, in the field example three groups can be distinguished, which are characterized by variations in their grain size versus temperature relationships: Group I occurs at low temperatures and the grain size remains constant because nano-scale second phase particles of organic origin inhibit grain growth in the calcite aggregates under these conditions. In the presence of an aqueous fluid, these second phases decay at a temperature of about 350 °C enabling the onset of grain growth in calcite. In the following growth period, fluid-enhanced group II and slower group III growth occurs. For group II a continuous and intense grain size increase with T is typical while the grain growth decreases with T for group III. None of the observed trends correlate with experimentally based grain growth kinetics, probably due to differences between nature and experiment which have not yet been investigated (e.g., porosity, second phases). Therefore, grain growth modeling was used to iteratively improve the correlation between measured and modeled grain sizes by optimizing activation energy (Q), pre-exponential factor (k0) and grain size exponent (n). For n=2, Q of 350 kJ/mol, k0 of 1.7×1021 μmns−1 and Q of 35 kJ/mol, k0 of 2.5×10-5 μmns−1 were obtained for group II and III, respectively. With respect to future work, field-data based grain growth modeling might be a promising tool for investigating the influences of secondary effects like porosity and second phases on grain growth in nature, and to unravel differences between nature and experiment.