Recent revelations and future directions using single-cell technologies in chronic lymphocytic leukemia

Chronic lymphocytic leukemia (CLL) is a clinically and biologically heterogeneous disease with varying outcomes. In the last decade, the application of next-generation sequencing technologies has allowed extensive mapping of disease-specific genomic, epigenomic, immunogenetic, and transcriptomic signatures linked to CLL pathogenesis. These technologies have improved our understanding of the impact of tumor heterogeneity and evolution on disease outcome, although they have mostly been performed on bulk preparations of nucleic acids. As a further development, new technologies have emerged in recent years that allow high-resolution mapping at the single-cell level. These include single-cell RNA sequencing for assessment of the transcriptome, both of leukemic and non-malignant cells in the tumor microenvironment; immunogenetic profiling of B and T cell receptor rearrangements; single-cell sequencing methods for investigation of methylation and chromatin accessibility across the genome; and targeted single-cell DNA sequencing for analysis of copy-number alterations and single nucleotide variants. In addition, concomitant profiling of cellular subpopulations, based on protein expression, can also be obtained by various antibody-based approaches. In this review, we discuss different single-cell sequencing technologies and how they have been applied so far to study CLL onset and progression, also in response to treatment. This latter aspect is particularly relevant considering that we are moving away from chemoimmunotherapy to targeted therapies, with a potentially distinct impact on clonal dynamics. We also discuss new possibilities, such as integrative multi-omics analysis, as well as inherent limitations of the different single-cell technologies, from sample preparation to data interpretation using available bioinformatic pipelines. Finally, we discuss future directions in this rapidly evolving field.</p

Recent revelations and future directions using single-cell technologies in chronic lymphocytic leukemia

Chronic lymphocytic leukemia (CLL) is a clinically and biologically heterogeneous disease with varying outcomes. In the last decade, the application of next-generation sequencing technologies has allowed extensive mapping of disease-specific genomic, epigenomic, immunogenetic, and transcriptomic signatures linked to CLL pathogenesis. These technologies have improved our understanding of the impact of tumor heterogeneity and evolution on disease outcome, although they have mostly been performed on bulk preparations of nucleic acids. As a further development, new technologies have emerged in recent years that allow high-resolution mapping at the single-cell level. These include single-cell RNA sequencing for assessment of the transcriptome, both of leukemic and non-malignant cells in the tumor microenvironment; immunogenetic profiling of B and T cell receptor rearrangements; single-cell sequencing methods for investigation of methylation and chromatin accessibility across the genome; and targeted single-cell DNA sequencing for analysis of copy-number alterations and single nucleotide variants. In addition, concomitant profiling of cellular subpopulations, based on protein expression, can also be obtained by various antibody-based approaches. In this review, we discuss different single-cell sequencing technologies and how they have been applied so far to study CLL onset and progression, also in response to treatment. This latter aspect is particularly relevant considering that we are moving away from chemoimmunotherapy to targeted therapies, with a potentially distinct impact on clonal dynamics. We also discuss new possibilities, such as integrative multi-omics analysis, as well as inherent limitations of the different single-cell technologies, from sample preparation to data interpretation using available bioinformatic pipelines. Finally, we discuss future directions in this rapidly evolving field

Turbulent Flow over Confined Backward-Facing Step: PIV vs. DNS

Particle Image Velocimetry measurements of the liquid velocity fields in the flow over the backward-facing step were performed in the same flow configuration as in the existing Direct Numerical Simulation (DNS). The experiment and the simulation were performed in an identical cross-section geometry with step expansion rate 2.25 and the square shape of the outlet duct at the Reynolds number in an inlet part of the section 7100. The experiment was performed in transparent test section, 1.2 m long, with 20 × 45 mm2 cross-section upstream and 45 × 45 mm2 downstream, while a domain that was three times shorter was used in the DNS. A 2D-2C PIV system with a single high-speed camera and a pulse laser was used for a series of two-dimensional measurements of the velocity field at several cross-sections from two different perspectives. Variables analyzed in the experiment are time-averaged fluid velocities, velocity RMS fluctuations and two components of the Reynolds stress tensor. The key novelty is the comparison of two very accurate approaches, PIV and DNS, in the same cross-section geometry. Comparison of the similarities, and especially the differences between the two approaches, elucidates uncertainties of both studies and answers the question on what kind of agreement is expected when two very accurate approaches are compared