Emerging Single-Cell Technological Approaches to Investigate Chromatin Dynamics and Centromere Regulation in Human Health and Disease

Epigenetic regulators play a crucial role in establishing and maintaining gene expression states. To date, the main efforts to study cellular heterogeneity have focused on elucidating the variable nature of the chromatin landscape. Specific chromatin organisation is fundamental for normal organogenesis and developmental homeostasis and can be affected by different environmental factors. The latter can lead to detrimental alterations in gene transcription, as well as pathological conditions such as cancer. Epigenetic marks regulate the transcriptional output of cells. Centromeres are chromosome structures that are epigenetically regulated and are crucial for accurate segregation. The advent of single-cell epigenetic profiling has provided finer analytical resolution, exposing the intrinsic peculiarities of different cells within an apparently homogenous population. In this review, we discuss recent advances in methodologies applied to epigenetics, such as CUT&RUN and CUT&TAG. Then, we compare standard and emerging single-cell techniques and their relevance for investigating human diseases. Finally, we describe emerging methodologies that investigate centromeric chromatin specification and neocentromere formation.

How high to ultra-high temperature terranes form

<p>Long-lived high- to ultra-high temperature (HT-UHT) terranes formed mostly during the Paleo-Proterozoic and are often associated to supercontinent cycles. Yet the detailed processes and conditions involved in their formation remain largely unresolved. Here we highlight the importance of the specific geothermal conditions necessary to form migmatitic to granulitic crusts. An analytical resolution of the heat equation highlights the interdependency of the thermal parameters controlling the crustal geotherm, i.e. the Moho temperature, when deformation occurs at thermal equilibrium. We further perform thermo-mechanical experiments mimicking an orogenic cycle, from shortening to gravitational collapse, to study the effect of deformation velocity that affects the crustal thermal equilibrium. We show that the formation of HT-UHT terranes is promoted by an elevated radiogenic heat production in the crust. Finally, the interplay between the thermal parameters and the orogenic cycle duration explain the difference in orogenic style through time and why some terranes are preferentially granulitic or migmatitic.</p>

Determination of Ozone or Hypochlorite in Waters Based on Digital Images Analysis Using Same Reagent

Ozone and hypochlorite are used for the disinfection of water and is essential to monitor these. In this paper, methods based on digital images were developed to determine these analytes in waters using the malachite green dye. The oxidation of the reagent with ozone was very fast and with hypochlorite occurred in 10 min in pseudo-first-order kinetics. New software called ICCA was developed for the capture and processing of red, green and blue (RGB) data, which is planned for digital image analysis. In addition, the recently proposed parameters analytical resolution and N9 were used to define some study conditions. An evaluation of different response functions and white correction of the webcam were done, having thrown some light on the subject to this theme. Their optimization showed a linear range from 0.6 mg L-1 to 5.0/6.0 mg L-1 (O3/Cl, respectively). The figures of merit were similar for both. The methods were validated and the recoveries achieved were between 85.0 and 96.0% (ozone) and 101.0 and 119.0% (hypochlorite).

On-Line Gaseous Formaldehyde Detection Based on a Closed-Microfluidic-Circuit Analysis

This paper describes a compact microfluidic analytical device in a closed-circuit developed for the detection of low airborne formaldehyde levels. The detection is based on the passive trapping of gaseous formaldehyde through a microporous tube into the acetylacetone solution, the derivative reaction of formaldehyde with acetylacetone to form 3,5-Diacetyl-1,4-dihydrolutidine (DDL) and the detection of DDL by fluorescence. The recirculation mode of the analytical device means that the concentration measurement is carried out by quantification of the signal increase in the liquid mixture over time, the instantaneous signal increase rate being proportional to the surrounding gaseous formaldehyde concentration. The response of this novel microdevice is found to be linear in the range 0–278 µg m−3. The reagent volume needed is flexible and depends on the desired analytical resolution time and the concentration of gaseous formaldehyde in the environment. Indeed, if either the gaseous concentration of formaldehyde is high or the reagent volume is low, the fluorescence signal of this recirculating liquid solution will increase very rapidly. Consequently, the sensitivity simultaneously depends on both the reagent volume and the temporal resolution. Considering a reagent volume of 6 mL, the hourly and daily detection limits are 2 and 0.08 µg m−3, respectively, while the reagent autonomy is more than 4 days the airborne formaldehyde concentration does not exceed 50 µg m−3 as it is usually the case in domestic or public indoor environments.

Simplified Micro-Mechanical Approach for Coated Viscoelastic Inclusion

The present work deals with the analytical resolution of the problem of viscoelastic coated inclusion embedded in a viscoelastic matrix.In a first step, we will study the problem of a linear viscoelastic inclusion, without coating, embedded in a linear viscoelastic matrix.Then, the problem of coated viscoelastic inclusion considering the coating as a thin layer whose viscoelastic properties are different from those of the inclusion and the matrix is performed.The resolution of this problem will be based simultaneously on the Green function technique as well as the interface operator. The analytical expression of the solution is obtained by assuming the isotropy of the matrix as well as the spherical shape of the coatedinclusion.These results are used to determine the effective properties of a heterogeneous medium from a self-consistent approach taking into accountthe interactions between coated inclusions and the equivalent homogeneous medium.

Analytical resolution of the Poisson–Boltzmann equation for a cylindrical polyion immersed in an electrolyte solution: using the optimal linearization method

By using the optimal linearization method (OLM), the potential of the electrical double layer created by a highly charged cylindrical polyion immersed in an electrolyte reservoir, which is represented by the so-called Poisson–Boltzmann equation (PBE), has been solved analytically under general potential conditions. For this system, three regions must be considered. The first one is in the near-neighborhood of the polyion and it is deprived of coions because of the repulsion phenomenon between the polyion and the coions, as proposed by Fuoss et al. (Proc. Natl. Acad. Sci. 37, 579 (1951). doi: 10.1073/pnas.37.9.579 ). For the second region, where the potential is slightly lower, we propose an OLM for solving the PBE. In the last region, where the potential is sufficiently low, the approximation of Debye–Hückel is adopted. This method allowed us to overcome some shortcomings in the analytical calculation of the electrostatic potential created by a polyion in an electrolyte solution.

The influence of cell morphology on microfluidic single cell analysis

Analytical resolution is influenced by cell morphology in microfluidic single cell analysis.

Application of Turbidity Curve Analysis to New Coagulation Factor VIII and IX Assays with Superior Analytical Resolution in Low-Level Range

Background: The needs for sensitive coagulation factor assays able to measure factor VIII (FVIII) and factor IX (FIX) in the range of 0.0 to 1.0 %, are continuously growing with diversification of hemophilia management. However, practical methods with sufficient analytical sensitivity available in clinical laboratory have not yet been introduced. We developed new coagulation factor assays applying various parameters derived from a turbidity based coagulometer and examined their ability to measure low-level FVIII and FIX and analytical resolution in that range. Method: We prepared 12 spiked samples with FVIII and FIX levels from 0.0 to 2.4 % and conducted conventional one-stage coagulation factor assays in repeat. We collected measured values of APTT, velocity and acceleration peaks of coagulation (peak 1 and peak 2) from each measurement. We also calculated values of peak 1 and peak 2 from the mathematical model of turbidity curves. From the measured values of these parameters we derived calibration formulae for coagulation factor assays, FVIIICT, FVIIIpeak1, FVIIIpeak2, FVIIIcalc1, FVIIIcalc2, FIXCT, FIXpeak1, FIXpeak2, FIXcalc1, and FIXcalc2. Results: The reliability interval (range of FVIII levels producing unequivocal results) of FVIIICT (the conventional FVIII assay) covered only 9 % of 0.0 to 1.0 % range. For new assays, the coverages were 54, 31, 55, and 65 % for FVIIIpeak1, FVIIIpeak2, FVIIIcalc1, and FVIIIcalc2 respectively. The resolution between immediate levels of spiked samples could be determined from modeled distributions or be checked simply by inspecting the actual assay result distributions. For FVIIIpeak1, 0.2 % and 0.6 % results stood apart from each other. For FVIIIcalc1 and FVIIIcalc2, 0.2, 0.4, and 0.6 % were distinguished from each other. When we measured recombinant human (rh) FVIII, the coverages were 7, 64, 52, 73, and 79 % for rhFVIIICT, rhFVIIIpeak1, rhFVIIIpeak2, rhFVIIIcalc1, and rhFVIIIcalc2 respectively. (rh)FVIIIpeak1, (rh)FVIIIcalc1, and (rh)FVIIIcalc2 particularly showed wide measurable ranges of guarantee. For FVIIIpeak1, 0.2 % and 0.6 % results stood apart from each other. For FVIIIcalc1 and FVIIIcalc2, 0.2, 0.4, and 0.6 % were distinguished from each other. rhFVIIIpeak1 and rhFVIIIcalc1 showed slightly better resolution than the former. rhFVIIIcalc2 was notable in that every 0.1, 0.2, 0.4, 0.6, 0.8 % result stood apart from each immediate level result. We could not determine certainty interval (the range of unequivocal values) of FIXCT and FIXpeak2 because the 0.0 % and 1.0 % ranges overlapped. Thus, the conventional FIX assay cannot measure between 0.0 and 1.0 %. FIXpeak1, FIXcalc1, and FIXcalc2 worked better and the certainty interval of unequivocal results could be determined between 0.0 and 1.0 %. The reliability interval was not available for any FIX assay. Results from rhFIX measurements were similar those of plasma FIX assays. Conclusion: We introduce new FVIII and FIX assays with superior analytical resolution in the range of 0.0 to 1.0 % in comparison to the conventional assays. Figure. Figure. Disclosures No relevant conflicts of interest to declare.