Cell Homogenates Research Articles

Development of a programmable magnetic agitation device to maintain colloidal suspension of cells during microfluidic syringe pump perfusion.

Droplet-based microfluidic devices have been used to achieve homogeneous cell encapsulation, but cells sediment in a solution, leading to heterogeneous products. In this technical note, we describe automated and programmable agitation device to maintain colloidal suspensions of cells. We demonstrate that the agitation device can be interfaced with a syringe pump for microfluidic applications. Agitation profiles of the device were predictable and corresponded to device settings. The device maintains the concentration of cells in an alginate solution over time without implicating cell viability. This device replaces manual agitation, and hence is suitable for applications that require slow perfusion for a longer period of time in a scalable manner.

CD73-Positive Cell Spheroid Transplantation Attenuates Colonic Atrophy

The incidence of inflammatory bowel diseases (IBD) is increasing worldwide. Mesenchymal stem/stromal cells (MSCs) have immunomodulatory functions and are a promising source for cell transplantation therapy for IBD. However, owing to their heterogeneous nature, their therapeutic efficacy in colitis is controversial and depends on the delivery route and form of transplanted cells. Cluster of differentiation (CD) 73 is widely expressed in MSCs and used to obtain a homogeneous MSC population. Herein, we determined the optimal method for MSC transplantation using CD73+ cells in a colitis model. mRNA sequencing analysis showed that CD73+ cells exhibited a downregulation of inflammatory gene expression and an upregulation of extracellular matrix-related gene expression. Furthermore, three-dimensional CD73+ cell spheroids showed enhanced engraftment at the injured site through the enteral route, facilitated extracellular matrix remodeling, and downregulated inflammatory gene expression in fibroblasts, leading to the attenuation of colonic atrophy. Therefore, the interaction between intestinal fibroblasts and exogenous MSCs via tissue remodeling is one mechanism that can be exploited for colitis prevention. Our results highlight that the transplantation of homogeneous cell populations with well-characterized properties is beneficial for IBD treatment.

Inferring density-dependent population dynamics mechanisms through rate disambiguation for logistic birth-death processes.

Density dependence is important in the ecology and evolution of microbial and cancer cells. Typically, we can only measure net growth rates, but the underlying density-dependent mechanisms that give rise to the observed dynamics can manifest in birth processes, death processes, or both. Therefore, we utilize the mean and variance of cell number fluctuations to separately identify birth and death rates from time series that follow stochastic birth-death processes with logistic growth. Our nonparametric method provides a novel perspective on stochastic parameter identifiability, which we validate by analyzing the accuracy in terms of the discretization bin size. We apply our method to the scenario where a homogeneous cell population goes through three stages: (1) grows naturally to its carrying capacity, (2) is treated with a drug that reduces its carrying capacity, and (3) overcomes the drug effect to restore its original carrying capacity. In each stage, we disambiguate whether the dynamics occur through the birth process, death process, or some combination of the two, which contributes to understanding drug resistance mechanisms. In the case of limited sample sizes, we provide an alternative method based on maximum likelihood and solve a constrained nonlinear optimization problem to identify the most likely density dependence parameter for a given cell number time series. Our methods can be applied to other biological systems at different scales to disambiguate density-dependent mechanisms underlying the same net growth rate.

Underwater cylindrical sandwich meta-structures composed of graded semi re-entrant zero Poisson's ratio metamaterials with pre-strained wave propagation properties

Zero Poisson's ratio (ZPR) mechanical metamaterials can yield no transverse displacements when unidirectionally compressed, and cylindrical sandwich meta-structures composed of semi re-entrant (SRE) ZPR metamaterials are thus explored for applications on cylindrical shells of underwater equipment or submersible structures. A group of ZPR unit cells with specified pre-strained wave propagation characteristics and adequate load-bearing capabilities is optimally designed based on the periodic boundary condition (PBC) and Bloch's Theorem. The sound transmission and pressure-resistant performance of cylindrical sandwich meta-structures comprising the homogeneous and graded SRE ZPR unit cells are then investigated. The results show that the designed meta-structures can perfectly yield better vibroacoustic attenuation behavior within the specified frequency regions corresponding to the pre-strained band gaps and safely bear the hydrostatic pressure equivalent to 1000 m depth with low weight-bulk ratios. In addition, the functionally graded metamaterial core can boost vibroacoustic performance within broader frequency ranges.

Homogeneity of XEN Cells Is Critical for Generation of Chemically Induced Pluripotent Stem Cells.

In induced pluripotent stem cells (iPSCs), pluripotency is induced artificially by introducing the transcription factors Oct4, Sox2, Klf4, and c-Myc. When a transgene is introduced using a viral vector, the transgene may be integrated into the host genome and cause a mutation and cancer. No integration occurs when an episomal vector is used, but this method has a limitation in that remnants of the virus or vector remain in the cell, which limits the use of such iPSCs in therapeutic applications. Chemical reprogramming, which relies on treatment with small-molecule compounds to induce pluripotency, can overcome this problem. In this method, reprogramming is induced according to the gene expression pattern of extra-embryonic endoderm (XEN) cells, which are used as an intermediate stage in pluripotency induction. Therefore, iPSCs can be induced only from established XEN cells. We induced XEN cells using small molecules that modulate a signaling pathway and affect epigenetic modifications, and devised a culture method in which can be produced homogeneous XEN cells. At least 4 passages were required to establish morphologically homogeneous chemically induced XEN (CiXEN) cells, whose properties were similar to those of XEN cells, as revealed through cellular and molecular characterization. Chemically iPSCs derived from CiXEN cells showed characteristics similar to those of mouse embryonic stem cells. Our results show that the homogeneity of CiXEN cells is critical for the efficient induction of pluripotency by chemicals.

Preparation of compact alginate films for 2D in vitro studies: Challenges and strategies for improvement

Alginate-based hydrogels show high biocompatibility and therefore are broadly used for cell cultivation and tissue engineering approaches in general. When hydrogel films for cell seeding were prepared by following published protocols, attached but inhomogeneously distributed cells were observed, which was attributed to the fact that casted hydrogel films were uneven and irregularly shaped. The causes of these irregularities resulting in an inhomogeneous cell distribution, e.g., due to accumulation of cells in cracks, were investigated. This study established a universal protocol for preparing planar hydrogel films that allow a homogenous cell distribution by varying several parameters such as alginate type, freezing, and crosslinking conditions. This finding can be applied to investigations of two-dimensional cell-material interactions of advanced alginate-based materials, e.g., when coupled with peptides.

Fabrication of 3D Bioprinted Bi-Phasic Scaffold for Bone-Cartilage Interface Regeneration.

Treatments for osteochondral defects (OCDs) are mainly palliative and, with the increase in this pathology seen among both young and elderly people, an alternative treatment modality is sought. Many tissue-engineered strategies have been explored for regenerating the cartilage-bone interface; however, they generally fall short of being ideal. Although cell-laden hydrogel scaffolds are a common approach for bone and cartilage tissue regeneration, they usually lack homogenous cell dispersion and patient specificity. In this study, a biphasic 3D bioprinted composite scaffold was fabricated for cartilage-bone interface regeneration. To overcome the shortcoming of both materials, alginate-gelatin (A-G) hydrogel was used to confer a naturally occurring environment for the cells and polycaprolactone (PCL) was used to enhance mechanical stability, thus maximizing the overall performance. Hydroxyapatite fillers were added to the PCL in the bone phase of the scaffold to improve its bioactivity. Physical and biological evaluation of scaffolds in both phases was assessed. The scaffolds demonstrated a desirable biological response both singly and in the combined PCL/A-G scaffolds, in both the short term and longer term, showing promise as an interfacial material between cartilage and bone.

Optimizing the homogeneity and efficiency of a solid oxide electrolysis cell based on multiphysics simulation and data-driven surrogate model

Inhomogeneous current and temperature distributions are harmful to the durability of solid oxide electrolysis cells (SOECs). A high steam utilization is favorable for system efficiency, but also enhances the inhomogeneity. This study combines segmented SOEC experiments, multiphysics simulation, and neural network to optimize the inhomogeneity and efficiency jointly. A three-dimensional (3D) cell model is built and experimental validation shows that the model correctly predicts the decreased down-stream current after the steam utilization exceeds 0.8. Fast surrogate models are trained with the simulation data and integrated into a multi-objective optimization problem for numerical solution. Its solutions form a Pareto front quantifying the conflicting relationship between the steam utilization, inhomogeneity, voltage, hydrogen production and working temperature, from which optimal solutions are chosen to achieve a trade-off. Under a power density of 1.11W cm−2, the ratio between the down-stream and up-stream currents drops from 63.1% to 55.2% when the steam utilization increases from 0.72 to 0.82. The Pareto fronts can enhance the collaboration between stack manufacturers and system operators by enabling the latter to optimize the operating point for a balance between system efficiency and inhomogeneity.

Organ-on-a-chip for dynamic tumor drug resistance investigation

Drug resistance continues to be the main limiting factor in the cure of cancer patients. Herein, an organ-on-a-chip platform based on inverse opal scaffold was proposed to generate tumor cell spheroids and study resistance dynamically. The relevant inverse opal scaffold with desired tridimensional porous structure was prepared by negatively replicating the assembled microfluidic droplet template. Benefitting from the ordered and uniform porous structure of the inverse opal scaffolds, hepatoma cells could aggregate in the pores of the scaffold and form large amounts of homogeneous hepatoma cell spheroids. The proposed liver tumor-on-a-chip was established by integrating the inverse opal scaffolds with a well-designed gradient microfluidic chip. The liver tumor models with varying degrees of drug-resistant phenotypes were realized by simply gradient injection of hepatoma parental and resistant cells into the microfluidic chip. Based on the resultant system, we have obtained the dynamic drug resistance performance of different tumor stages and diverse medication periods, which will provide extremely significant preclinical data for drug therapy of tumors. These features make the proposed organ-on-a-chip platform a valuable candidate for preclinical drug resistance evaluation.

Hypoxia increases cellular levels of phosphatidic acid and lysophospholipids in undifferentiated Caco-2 cells.

Cancer cells are known to survive in a hypoxic microenvironment by altering their lipid metabolism as well as their energy metabolism. In this study, Caco-2 cells derived from human colon cancer, were found to have elevated intracellular levels of phosphatidic acid and its lysoform, lysophosphatidic acid (LPA), under hypoxic conditions. Our results suggested that the elevation of LPA in Caco-2 cells was mainly due to the combined increases in cellular levels of lysophosphatidylcholine and lysophosphatidylethanolamine by phospholipase A2 and subsequent hydrolysis to LPA by lysophospholipase D. We detected the Ca2+ -stimulated choline-producing activities toward exogenous lysophosphatidylcholines in whole Caco-2 cell homogenates, indicating their involvement in the LPA production in intact Caco-2 cells.

Fast and safe heating-up control of a planar solid oxide cell stack: A three-dimensional model-in-the-loop study

A fast heating-up rate generally conflicts with the temperature homogeneity of solid oxide cell (SOC) stacks. This study designs an adaptive model predictive controller (MPC) to regulate the heating-up process of SOC stacks. The controller solves an optimization problem in real time to obtain optimal control that shortens the heating-up time by fully using the safety limit. The controller is verified with a three-dimensional (3D) hotbox–stack model calibrated with experiments. 3D model-in-the-loop (MiL) simulations show that the controller is effective under two heating modes despite of model deviations. Under the electrical heating mode, the controller achieves a 33% acceleration compared with a reference control strategy. Comparisons of electrical and gas heating reveal that the former is 63% faster but the latter consumes 41.6% less energy if the waste heat is fully recovered. The hotbox heat capacity is a major limiting factor for the heating-up process, particularly under the gas heating mode. An 80% reduction in the hotbox heat capacity, which can be achieved by reducing the thickness of the insulation layer, improves the heating-up time and the net energy consumption by 69% and 65% under the gas heating mode, respectively.

Mitochondrial transplant after ischemia reperfusion promotes cellular salvage and improves lung function during ex-vivo lung perfusion

In lung transplantation, ischemia-reperfusion injury associated with mitochondrial damage can lead to graft rejection. Intact, exogenous mitochondria provide a unique treatment option to salvage damaged cells within lung tissue. We developed a novel method to freeze and store allogeneic mitochondria isolated from porcine heart tissue. Stored mitochondria were injected into a model of induced ischemia-reperfusion injury using porcine ex-vivo lung perfusion. Treatment benefits to immune modulation, antioxidant defense, and cellular salvage were evaluated. These findings were corroborated in human lungs undergoing ex-vivo lung perfusion. Lung tissue homogenate and primary lung endothelial cells were then used to address underlying mechanisms. Following cold ischemia, mitochondrial transplant reduced lung pulmonary vascular resistance and tissue pro-inflammatory signaling and cytokine secretion. Further, exogenous mitochondria reduced reactive oxygen species by-products and promoted glutathione synthesis, thereby salvaging cell viability. These results were confirmed in a human model of ex-vivo lung perfusion wherein transplanted mitochondria decreased tissue oxidative and inflammatory signaling, improving lung function. We demonstrate that transplanted mitochondria induce autophagy and suggest that bolstered autophagy may act upstream of the anti-inflammatory and antioxidant benefits. Importantly, chemical inhibitors of the MEK autophagy pathway blunted the favorable effects of mitochondrial transplant. These data provide direct evidence that mitochondrial transplant improves cellular health and lung function when administered during ex-vivo lung perfusion and suggest the mechanism of action may be through promotion of cellular autophagy. Data herein contribute new insights into the therapeutic potential of mitochondrial transplant to abate ischemia-reperfusion injury during lung transplant, and thus reduce graft rejection.

Customizable 3D printed perfusion bioreactor for the engineering of stem cell microenvironments.

Faithful modeling of tissues and organs requires the development of systems reflecting their dynamic 3D cellular architecture and organization. Current technologies suffer from a lack of design flexibility and complex prototyping, preventing their broad adoption by the scientific community. To make 3D cell culture more available and adaptable we here describe the use of the fused deposition modeling (FDM) technology to rapid-prototype 3D printed perfusion bioreactors. Our 3D printed bioreactors are made of polylactic acid resulting in reusable systems customizable in size and shape. Following design confirmation, our bioreactors were biologically validated for the culture of human mesenchymal stromal cells under perfusion for up to 2weeks on collagen scaffolds. Microenvironments of various size/volume (6-12mm in diameter) could be engineered, by modulating the 3D printed bioreactor design. Metabolic assay and confocal microscopy confirmed the homogenous mesenchymal cell distribution throughout the material pores. The resulting human microenvironments were further exploited for the maintenance of human hematopoietic stem cells. Following 1 week of stromal coculture, we report the recapitulation of 3D interactions between the mesenchymal and hematopoietic fractions, associated with a phenotypic expansion of the blood stem cell populations.Our data confirm that perfusion bioreactors fit for cell culture can be generated using a 3D printing technology and exploited for the 3D modeling of complex stem cell systems. Our approach opens the gates for a more faithful investigation of cellular processes in relation to a dynamic 3D microenvironment.

Functional mass spectrometry imaging maps phospholipase-A2 enzyme activity during osteoarthritis progression.

Background: Enzymes are central components of many physiological processes, and changes in enzyme activity are linked to numerous disease states, including osteoarthritis (OA). Assessing changes in enzyme function can be challenging because of difficulties in separating affected tissue areas that result in the homogenisation of healthy and diseased cells. Direct correlation between spatially-resolved enzyme distribution(s) and diseased cells/tissues can thus lead to advances in our understanding of OA pathophysiology. Herein, we present a method that uses mass spectrometry imaging (MSI) to visualise the distribution of lipase enzymes and their downstream lipid products in fresh bone and cartilage tissue sections. Immunohistostaining of adjacent tissue sections was then used to identify OA cells/tissues, which were then statistically correlated with molecular-level images. Methods: MSI was used to image lipase enzymes, their substrates, and their metabolic products to validate enzymatic activity and correlate to OA regions determined by immunohistochemistry (IHC). Based on the modified Mankin score, six non-OA and OA patient-matched osteochondral samples were analysed by matrix-assisted laser desorption/ionisation mass spectrometry imaging (MALDI-MSI). Due to the involvement of phospholipase A2 (PLA2) in inflammatory pathways, explant tissues were treated with IL-1β to mimic inflammation observed in OA. Bovine explant tissues were then subject to MSI methods to observe the spatial distribution of PLA2. Results: Compared with non-OA samples, OA samples showed an elevated level of multiple arachidonic acid (AA)-containing phospholipids (P < 0.001), in which the elevation in the surface and deep layer cartilage of OA tissues is correlated to elevated PLA2 activity (P < 0.001). Bovine explant tissues treated with IL-1β to mimic OA pathophysiology validated these results and displayed elevated PLA2 levels in OA mimic samples relative to the controls (P < 0.001). It was established that the PLA2G2A isoform specifically was responsible for PLA2 enzyme activity changes in OA tissues (P < 0.001). Conclusion: Our results present a reliable method for imaging enzyme dynamics in OA cartilage, which sets up the foundation for future spatial enzyme dynamics in the OA field. We demonstrated that OA patients exhibit increased expression of PLA2G2A at the superficial and deep cartilage zone that degrades cartilage differently at the spatial level. A tissue-specific PLA2G2A precision inhibition may be the potential target for OA.

Reversibly Tuning Electrochemiluminescence with Stimulated Emission Route for Single-Cell Imaging

Electrochemiluminescence (ECL) has established itself as an excellent transduction technique in biosensing and light-emitting device, while conventional ECL mechanism depending on spontaneous emission of luminophores lacks reversibility and tunable emission characters, limiting the universality of ECL technique in the fields of fundamental research and clinical applications. Here, we report the first observation of stimulated emission route in ECL and thus establish a reversible tuning ECL microscopy for single-cell imaging. This microscopy uses a focused red-shifted beam to transfer spontaneous ECL into stimulated ECL, which enables selective and reversible tuning of ECL emission from homogeneous solution, single particles, and single cells. After excluding other possible competitive routes, the stimulated ECL emission route is confirmed by a dual-objective system in which the suppressed spontaneous ECL is accompanied by the enhanced stimulated ECL. By incorporating a commercial donut-shaped beam, the sharpness of single-cell matrix adhesion is improved 2 to 3 times compared with the counterpart in confocal ECL mode. The successful establishment of this stimulated emission ECL will greatly advance the development of light-emitting device and super-resolution ECL microscopy.

In Vitro Generation of Murine Dendritic Cells from Hoxb8-Immortalized Hematopoietic Progenitors.

Mouse dendritic cells (DCs) are routinely generated based on cells isolated form the bone marrow (BM) and cultured in the presence of growth factors that support DC development, such as FMS-like tyrosine kinase 3 ligand (FLT3L) and granulocyte-macrophage colony-stimulating factor (GM-CSF) (Guo et al., J Immunol Methods 432:24-29, 2016). In response to these growth factors, DC progenitors expand and differentiate, while other cell types die during the in vitro culture period, ultimately leading to relatively homogenous DC populations. An alternative method, which is discussed in detail in this chapter, relies on conditional immortalization of progenitor cells with DC potential in vitro using an estrogen-regulated form of Hoxb8 (ERHBD-Hoxb8). Such progenitors are established by retroviral transduction of largely unseparated BM cells with a retroviral vector expressing ERHBD-Hoxb8. Treatment of ERHBD-Hoxb8-expressing progenitors with estrogen results in Hoxb8 activation, which blocks cell differentiation and allows for expansion of homogenous progenitor cell populations in the presence of FLT3L. These cells, referred to as Hoxb8-FL cells, retain lineage potential for lymphocyte and myeloid lineages, including the DC lineage. Upon removal of estrogen (inactivation of Hoxb8), Hoxb8-FL cells differentiate into highly homogenous DC populations in the presence of GM-CSF or FLT3L akin to their endogenous counterparts. Given their unlimited proliferative capacity and amenability for genetic manipulation, for example, by CRISPR/Cas9, these cells provide a large number of options to investigate DC biology. Here, I am describing the method to establish Hoxb8-FL cells from mouse BM, as well as procedures for DC generation and gene deletion using lentivirally delivered CRISPR/Cas9.

Imaging Flow Cytometry of Legionella-Containing Vacuoles in Intact and Homogenized Wild-Type and Mutant Dictyostelium.

The causative agent of asevere pneumonia termed"Legionnaires' disease", Legionella pneumophila, replicates within protozoan and mammalian phagocytes in a specialized intracellular compartment called the Legionella-containing vacuole (LCV). This compartment does not fuse with bactericidal lysosomes but communicates extensively with several cellular vesicle trafficking pathways and eventually associates tightly with the endoplasmic reticulum. In order to comprehend in detail the complex process of LCV formation, the identification and kinetic analysis of cellular trafficking pathway markers on the pathogen vacuole are crucial. This chapter describes imaging flow cytometry (IFC)-based methods for the objective, quantitative and high-throughput analysis of different fluorescently tagged proteins or probes on the LCV. To this end, we use the haploid amoeba Dictyostelium discoideum as an infection model for L. pneumophila, to analyze either fixed intact infected host cells or LCVs from homogenized amoebae. Parental strains and isogenic mutant amoebae are compared in order to determine the contribution of a specific host factor to LCV formation. The amoebae simultaneously produce two different fluorescently tagged probes enabling tandem quantification of two LCV markers in intact amoebae or the identification of LCVs using one probe and quantification of the other probe in host cell homogenates. The IFC approach allows rapid generation of statistically robust data from thousands of pathogen vacuoles and can be applied to other infection models.

Human iPSC-derived midbrain organoids functionally integrate into striatum circuits and restore motor function in a mouse model of Parkinson's disease.

Rationale: Parkinson's disease (PD) is a prevalent neurodegenerative disorder that is characterized by degeneration of dopaminergic neurons (DA) at the substantia nigra pas compacta (SNpc). Cell therapy has been proposed as a potential treatment option for PD, with the aim of replenishing the lost DA neurons and restoring motor function. Fetal ventral mesencephalon tissues (fVM) and stem cell-derived DA precursors cultured in 2-dimentional (2-D) culture conditions have shown promising therapeutic outcomes in animal models and clinical trials. Recently, human induced pluripotent stem cells (hiPSC)-derived human midbrain organoids (hMOs) cultured in 3-dimentional (3-D) culture conditions have emerged as a novel source of graft that combines the strengths of fVM tissues and 2-D DA cells. Methods: 3-D hMOs were induced from three distinct hiPSC lines. hMOs at various stages of differentiation were transplanted as tissue pieces into the striatum of naïve immunodeficient mouse brains, with the aim of identifying the most suitable stage of hMOs for cellular therapy. The hMOs at Day 15 were determined to be the most appropriate stage and were transplanted into a PD mouse model to assess cell survival, differentiation, and axonal innervation in vivo. Behavioral tests were conducted to evaluate functional restoration following hMO treatment and to compare the therapeutic effects between 2-D and 3-D cultures. Rabies virus were introduced to identify the host presynaptic input onto the transplanted cells. Results: hMOs showed a relatively homogeneous cell composition, mostly consisting of dopaminergic cells of midbrain lineage. Analysis conducted 12 weeks post-transplantation of day 15 hMOs revealed that 14.11% of the engrafted cells expressed TH+ and over 90% of these cells were co-labeled with GIRK2+, indicating the survival and maturation of A9 mDA neurons in the striatum of PD mice. Transplantation of hMOs led to a reversal of motor function and establishment of bidirectional connections with natural brain target regions, without any incidence of tumor formation or graft overgrowth. Conclusion: The findings of this study highlight the potential of hMOs as safe and efficacious donor graft sources for cell therapy to treat PD.

High-throughput adjustable deformability cytometry utilizing elasto-inertial focusing and virtual fluidic channel.

Cell mechanical properties provide a label-free marker for indicating cell states and disease processes. Although microfluidic deformability cytometry has demonstrated great potential and successes in mechanical phenotyping in recent years, its universal applicability for characterizing multiple sizes of cells using a single device has not been realized. Herein, we propose high-throughput adjustable deformability cytometry integrated with three-dimensional (3D) elasto-inertial focusing and a virtual fluidic channel. By properly adjusting the flow ratio of the sample and sheath, the virtual fluidic channel in a wide solid channel can generate a strong shear force in the normal direction of the flow velocity and simultaneously squeeze cells from both sides to induce significant cell deformation. The combination of elasto-inertial focusing and a virtual fluidic channel provides a great hydrodynamic symmetrical force for inducing significant and homogeneous cell deformation. In addition, our deformability cytometry system not only achieves rapid and precise cell deformation, but also allows the adjustable detection of multiple sizes of cells at a high throughput of up to 3000 cells per second. The mini-bilateral segmentation network (mini-BiSeNet) was developed to identify cells and extract features quickly. The classification of different cell populations (A549, MCF-7, MDA-MB-231, and WBCs) was carried out based on the cell size and deformation. By applying deep learning to cell classification, a high accuracy reaching approximately 90% was achieved. We also revealed the potential of our deformability cytometry for characterizing pleural effusions. The flexibility of our deformability cytometry holds promise for the mechanical phenotyping and detection of various biological samples.

SimCH: simulation of single-cell RNA sequencing data by modeling cellular heterogeneity at gene expression level.

Single-cell ribonucleic acid (RNA) sequencing (scRNA-seq) has been a powerful technology for transcriptome analysis. However, the systematic validation of diverse computational tools used in scRNA-seq analysis remains challenging. Here, we propose a novel simulation tool, termed as Simulation of Cellular Heterogeneity (SimCH), for the flexible and comprehensive assessment of scRNA-seq computational methods. The Gaussian Copula framework is recruited to retain gene coexpression of experimental data shown to be associated with cellular heterogeneity. The synthetic count matrices generated by suitable SimCH modes closely match experimental data originating from either homogeneous or heterogeneous cell populations and either unique molecular identifier (UMI)-based or non-UMI-based techniques. We demonstrate how SimCH can benchmark several types of computational methods, including cell clustering, discovery of differentially expressed genes, trajectory inference, batch correction and imputation. Moreover, we show how SimCH can be used to conduct power evaluation of cell clustering methods. Given these merits, we believe that SimCH can accelerate single-cell research.