Cell Size Heterogeneity Research Articles

Single-cell imaging of the Mycobacterium tuberculosis cell cycle reveals linear and heterogenous growth.

Difficulties in antibiotic treatment of Mycobacterium tuberculosis (Mtb) are partly thought to be due to heterogeneity in growth. Although the ability of bacterial pathogens to regulate growth is crucial to control homeostasis, virulence and drug responses, single-cell growth and cell cycle behaviours of Mtb are poorly characterized. Here we use time-lapse, single-cell imaging of Mtb coupled with mathematical modelling to observe asymmetric growth and heterogeneity in cell size, interdivision time and elongation speed. We find that, contrary to Mycobacterium smegmatis, Mtb initiates cell growth not only from the old pole but also from new poles or both poles. Whereas most organisms grow exponentially at the single-cell level, Mtb has a linear growth mode. Our data show that the growth behaviour of Mtb diverges from that of model bacteria, provide details into how Mtb grows and creates heterogeneity and suggest that growth regulation may also diverge from that in other bacteria.

Red cell distribution width as a predictor of cardiovascular events in high-risk patients with statin intolerance: post-hoc analysis of the CLEAR Outcomes trial

Abstract Background Despite optimal lipid-lowering therapy there is up to 70% residual risk of recurrent atherosclerotic cardiovascular disease (ASCVD) events, motivating further testing of alternative risk factors and disease pathways. Red cell distribution width (RDW) is a simple and reproducible marker of heterogeneity of cell size in the peripheral blood. High RDW is associated with an increased risk of ASCVD and death, independent of established risk factors. Recent data suggest no significant impact of PCSK9 inhibitors on hsCRP or RDW. In CLEAR Outcomes, baseline hsCRP predicted risk for future cardiovascular (CV) events and death more strongly than elevated LDL-C. It has been shown that bempedoic acid (BA) reduces hsCRP, but its effects on RDW are unknown. Purpose To analyze the effect of BA on a 4-component composite of incident myocardial infarction, stroke, coronary revascularization, or cardiovascular death (MACE-4) and for CV death by baseline RDW, and to confirm the association of increased RDW with elevated risk of MACE. Methods A total of 13,970 individuals with statin intolerance, with or at high risk for ASCVD, were randomized in the multinational CLEAR Outcomes trial with a median follow up of 3.4 years. Compared to placebo, BA reduced mean LDL-C by 21% and median hsCRP by 22% at 6 months. Quartiles of increasing baseline RDW were assessed as predictors of future adverse events after adjustment for traditional risk factors, hsCRP, and randomized treatment assignment. Results Compared to placebo, BA reduced mean RDW by 0.28% at 6 months and 0.13% at 36 months (p<0.0001). There was a weak positive correlation between hsCRP and RDW at baseline (Figure 1). In the placebo group, higher baseline RDW was associated with an increase in the likelihood of MACE-4 and CV death, and the hazard ratios (HR) associated with a 1% increase in RDW after adjusting for covariates and hsCRP were 1.06 (95% CI 1.02-1.11) and 1.22 (1.15-1.29), respectively. When grouped by quartiles of baseline RDW, compared with a reference group with lowest RDW (<13.7%), there was a notable increase in the risk of MACE-4 and CV death among the higher quartiles in the placebo group (Table 1). Those in the 4th quartile demonstrated a 3-fold increase in the risk of CV death (HR, 3.03, 2.00-4.60). The adjusted HRs (95% CI) for BA vs placebo for MACE-4 were generally lower in individuals with elevated baseline RDW than in those in the first quartile [Q1: RDW≤13.7%; 0.96 (0.79-1.17), Q2: RDW 13.7%-14.4%; 0.81 (0.67-0.97), Q3: RDW 14.4%-15.2%; 0.90 (0.75-1.09), and Q4: RDW>15.2%; 0.84 (0.7-1.01)]. Conclusions In a large randomized controlled clinical trial setting, we confirmed an association between elevated RDW and MACE-4. Compared with placebo, BA demonstrated efficacy in reducing CV risk in patients with moderately increased RDW.Figure 1.Correlation of hsCRP and RDWTable 1.Risk of Outcome by RDW Quartile

Cell-to-cell signaling in cell populations with large cell size variability.

Sizes of multiple cells vary when they communicate with each other. Differences in cell size result in variations in the cell surface area and volume, as well as the number of enzymes and receptors involved in signal transduction. Although heterogeneity in cell size may inhibit uniformity in signaling, cell-to-cell signaling is still possible. The outcome when cell size changes to an extreme degree remains unclear. Hence, we inhibited cell division in Dictyostelium cells, a model organism for signal transduction, to gain insights into the consequences of extreme cell size variations. Measurements of cell signals in this population using fluorescence microscopy indicated that the giant cells can communicate with normal-sized cells by suppressing the signal level. Simulations of signal transduction based on the FitzHugh-Nagumo model also suggested similar results.Our findings suggest that signaling mechanism homogenizes cell-to-cell signaling in response to cell size.

Sepal shape variability is robust to cell size heterogeneity in Arabidopsis.

How organisms produce organs with robust shapes and sizes is still an open question. In recent years, the Arabidopsis sepal has been used as a model system to study this question because of its highly reproducible shape and size. One interesting aspect of the sepal is that its epidermis contains cells of very different sizes. Previous reports have qualitatively shown that sepals with more or less giant cells exhibit comparable final size and shape. Here, we investigate this question using quantitative approaches. We find that a mixed population of cell size modestly contribute to the normal width of the sepal but is not essential for its shape robustness. Furthermore, in a mutant with increased cell and organ growth variability, the change in final sepal shape caused by giant cells is exaggerated but the shape robustness is not affected. This formally demonstrates that sepal shape variability is robust to cell size heterogeneity.

Integrated molecular and multiparametric MRI mapping of high-grade glioma identifies regional biologic signatures

Sampling restrictions have hindered the comprehensive study of invasive non-enhancing (NE) high-grade glioma (HGG) cell populations driving tumor progression. Here, we present an integrated multi-omic analysis of spatially matched molecular and multi-parametric magnetic resonance imaging (MRI) profiling across 313 multi-regional tumor biopsies, including 111 from the NE, across 68 HGG patients. Whole exome and RNA sequencing uncover unique genomic alterations to unresectable invasive NE tumor, including subclonal events, which inform genomic models predictive of geographic evolution. Infiltrative NE tumor is alternatively enriched with tumor cells exhibiting neuronal or glycolytic/plurimetabolic cellular states, two principal transcriptomic pathway-based glioma subtypes, which respectively demonstrate abundant private mutations or enrichment in immune cell signatures. These NE phenotypes are non-invasively identified through normalized K2 imaging signatures, which discern cell size heterogeneity on dynamic susceptibility contrast (DSC)-MRI. NE tumor populations predicted to display increased cellular proliferation by mean diffusivity (MD) MRI metrics are uniquely associated with EGFR amplification and CDKN2A homozygous deletion. The biophysical mapping of infiltrative HGG potentially enables the clinical recognition of tumor subpopulations with aggressive molecular signatures driving tumor progression, thereby informing precision medicine targeting.

Abstract 5621: Multi-parametric MRI maps regional heterogeneity of high grade glioma phenotypes

Abstract High grade glioma (HGG) represents a group of devastating diseases with dismal prognosis. Surgical resection of the contrast enhancing (CE) region of HGG remains the mainstay of treatment, but recurrence inevitably arises from the unresected non-contrast enhancing (NE) region, surgically inaccessible due to cancer cell invasion into healthy brain tissue. Due to its critical role in recurrence, understanding of the NE region is central to the improvement of clinical outcomes. We reveal the biological characteristics of this region through image localized multi-regional sampling. We linked microenvironmental characteristics measured by multi-parametric MRI to genomic mutations and transcriptional phenotypes using mixed effect modeling which allowed us to control for individualized patient effects. We first confirmed that T2 is a significant indicator of IDH mutation status in the NE region, being the first description of such a relationship in a HGG cohort. We found the combination of EGFR amplification and CDKN2A homozygous loss was associated with a significantly lower mean diffusivity (MD) compared to double wild type tumors in the NE region, indicating the presence of greater cellular packing and proliferation in EGFR amplification/CDKN2A loss regions. Finally, using single cell pathway based tumor classifications, we showed that nK2, a DSC-MRI metric representing cell size heterogeneity, correlated positively with neuronal signature and negatively with glycolytic/plurimetabolic signature within the NE tumor, indicating that glycolytic/plurimetobolic tumors possessed a high amount of cell size heterogeneity compared to neuronal samples. This hypothesis was supported using digital reference object (DRO) modeling which confirmed that cell size and heterogeneity drove the differential nK2 signal between neuronal and glycolytic/plurimetabolic samples. We identified immune cell infiltrate as one possible mechanism of increased cell size heterogeneity using transcriptomic signature analysis which found more immune cell signatures within glycolytic/plurimetobolic tumors compared to neuronal. Collectively this study demonstrates the central role of multi-parametric MRI as a non-invasive measure of tumor biology and a tool for understanding the clinically critical NE region which can then inform new therapies targeting this region of HGG recurrence. Citation Format: Matthew Flick, Taylor Weiskittel, Kevin Meng-Lin, Fulvio D'Angelo, Francesca Caruso, Shannon Ensign, Mylan Blomquist, Luija Wang, Christopher Sereduk, Gustavo De Leon, Ashley Nespodzany, Javier Urcuyo, Ashlynn Gonzalez, Lee Curtin, Kyle Singleton, Aliya Anil, Natenael Simmineh, Erika Lewis, Teresa Noviello, Reyna Patel, Panwen Wang, Junwen Wang, Jennifer Eschbacher, Andrea Hawkins-Daarud, Pamela Jackson, Kris Smith, Peter Nakaji, Bernard Bendok, Richard Zimmerman, Chandan Krishna, Devi Patra, Naresh Patel, Mark Lyons, Matthew Neal, Kliment Donev, Maciej Mrugala, Alyx Porter, Scott Beeman, Yuxiang Zhou, Leslie Baxter, Christopher Plaisier, Jing Li, Hu Li, Anna Lasorella, Chad Quarles, Kristin Swanson, Michele Ceccarelli, Antonio Iavarone, Nhan Tran, Leland Hu. Multi-parametric MRI maps regional heterogeneity of high grade glioma phenotypes. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5621.

Relation of Red Cell Distribution Width to Glucose Metabolism and Adverse Long-Term Prognosis in Patients with Acute Coronary Syndrome

Red cell distribution width (RDW) reflects the heterogeneity of red blood cell size. However, few studies examined whether RDW is related to glucose metabolism indices, such as fasting blood glucose (FBG) and hemoglobin A1c (HbA1c), diabetic mellitus (DM) state or long-term outcomes of acute coronary syndrome (ACS) patients. A total of 448 consecutive patients with ACS were enrolled in this study. All patients were followed up for major cardiovascular adverse events (MACEs), and the mean follow-up was 952 days. Linear regression analysis showed that RDW inversely correlated with FBG but not HbA1c or DM. Kaplan-Meier survival curve analysis demonstrated that higher RDW levels were significantly positively associated with MACEs in the whole study population and the ACS patients with high FBG but not the low FBG group. Cox multivariate regression analysis revealed the independent function of RDW on MACEs in all ACS patients and ACS patients with high FBG. The receiver operating characteristic (ROC) curve demonstrated the optimal cutoff value of RDW for MACEs. We first reported that higher RDW was associated with decreased FBG but not HbA1c or DM and an increased risk of MACEs in patients with ACS. This relationship was also found in ACS patients with higher FBG levels but not in ACS patients with lower FBG.

PS-C06-9: THE PREDICTIVE VALUE OF RED CELL DISTRIBUTION WIDTH IN VENOUS THROMBOEMBOLISM AND RELATED MORTALITY

Background: Red cell distribution width (RDW) is a measure of red blood cell size variation and heterogeneity. Recent meta-analyses demonstrated this measurement could be strong predictor in coronary artery disease, chronic heart failure, and atrial fibrillation. Nevertheless, there is no meta-analysis evaluating the role of RDW in prediction of the occurrence of venous thromboembolism (VTE), the third most common cardiovascular disease, and related mortality. Objective: To investigate the association of RDW and VTE including deep vein thrombosis (DVT) and pulmonary embolism (PE) and related mortality. Methods: Major medical databases were systematically searched for observational studies which assessed the association of RDW and VTE published until May 2022. The databases were searched with predefined protocol without language restriction based on PRISMA guidelines. The analysis was performed in RevMan 5.3 to provide pooled measures for Risk Ratio (RR) and weighted mean difference (WMD). Results: Twenty-three studies were enrolled including 6,737 cases and 58,831 controls. Higher RDW was indicated as independent predictors for VTE compared to lower RDW (RR = 1.82, p < 0.00001), comprising DVT and PE (RR = 1.6, p < 0.00001, and RR = 2.05, p = 0.002, respectively). There was also an association between higher RDW and risk of mortality after PE episode with RR = 1.4 (p = 0.0004), involving short- (30-day) and long-term (> 1 year) mortality (RR = 1.79, p = 0.04, and RR = 1.15,p = 0.03, respectively). Compared to lower RDW, higher RDW also revealed a significant association with unprovoked VTE (RR = 2.18, p = 0.02). VTE-group had a higher mean of RDW compared to control with WMD = 1.13% (p < 0.00001), consisting of PE-group (WMD = 0.99%, p < 0.00001) and DVT-group (WMD = 1.07%, p < 0.00001). Among PE patients, The PE-related mortality group also had a higher mean of RDW in comparison to the survivor group with WMD = 1.97%, (p < 0.00001). Conclusions: Higher RDW value was associated with the occurrence of VTE, including DVT, PE, unprovoked VTE, and PE-related mortality. Therefore, the use of the potential role of RDW should be emphasized since it is low-cost and simple to obtain, even in a low-resource setting.

Isolation and characterization of head and neck cancer-derived peritumoral and cancer-associated fibroblasts.

Head and neck squamous cell carcinomas (HNSCC) are characterized by strong cellular and molecular heterogeneity and treatment resistance entailing poor survival. Besides cell-intrinsic properties, carcinoma cells receive important cues from non-malignant cells within the tumor microenvironment (TME). Cancer-associated fibroblasts (CAFs) are a major component of the TME that impact on the molecular make-up of malignant cells and have a decisive function in tumor progression. However, the potential functionality of fibroblasts within tumor-adjacent, macroscopically normal tissue remains poorly explored. Here, we isolated primary peritumoral fibroblasts (PtFs) from macroscopically normal tissue in vicinity of primary human papillomavirus-negative and -positive oropharyngeal HNSCC and compared their phenotype and functionality with matched CAFs (n = 5 pairs) and with human oral fibroblasts (hOFs). Expression patterns of CD90, CD73, CD105, smooth muscle actin, Vimentin, and S100A4 were comparable in PtFs, CAFs, and hOFs. Cell proliferation and doubling times of CAFs and PtFs were heterogeneous across patients (n =2 PtF>CAF; n = 1 CAF>PtF; n = 2 CAF=PtF) and reflected inferior growth than hOFs. Furthermore, PtFs displayed an reduced heterogeneity in cell size compared to matched CAFs, which were characterized by the presence of single large cells. Overall, conditioned supernatants from CAFs had more frequently growth-promoting effects on a panel of carcinoma cell lines of the upper aerodigestive tract carcinoma cell lines (Cal27, Cal33, FaDu, and Kyse30), whereas significant differences in migration-inducing effects demonstrated a higher potential of PtFs. Except for Kyse30, CAFs were significantly superior to hOFs in promoting proliferation, while PtFs induced stronger migration than hOFs in all carcinoma lines tested. Analysis of soluble factors demonstrated significantly increased VEGF-A production in CAFs (except in pat.8), and significantly increased PDGF-BB production in PtFs of two patients. Tube formation assays confirmed a significantly enhanced angiogenic potential of conditioned supernatants from CAFs compared to hOFs on human umbilical vascular endothelial cells (HUVECs) in vitro. Hence, matched CAFs and PtFs present in HNSCC patients are heterogeneous in their proliferation-, migration-, and angiogenesis-promoting capacity. Despite this heterogeneity, CAFs induced stronger carcinoma cell proliferation and HUVEC tube formation overall, whereas PtFs promoted migration of tumor cells more strongly.

High-throughput determination of dry mass of single bacterial cells by ultrathin membrane resonators

How bacteria are able to maintain their size remains an open question. Techniques that can measure the biomass (dry mass) of single cells with high precision and high-throughput are demanded to elucidate this question. Here, we present a technological approach that combines the transport, guiding and focusing of individual bacteria from solution to the surface of an ultrathin silicon nitride membrane resonator in vacuum. The resonance frequencies of the membrane undergo abrupt variations at the instants where single cells land on the membrane surface. The resonator design displays a quasi-symmetric rectangular shape with an extraordinary capture area of 0.14 mm2, while maintaining a high mass resolution of 0.7 fg (1 fg = 10−15 g) to precisely resolve the dry mass of single cells. The small rectangularity of the membrane provides unprecedented frequency density of vibration modes that enables to retrieve the mass of individual cells with high accuracy by specially developed inverse problem theory. We apply this approach for profiling the dry mass distribution in Staphylococcus epidermidis and Escherichia coli cells. The technique allows the determination of the dry mass of single bacterial cells with an accuracy of about 1% at an unparalleled throughput of 20 cells/min. Finally, we revisit Koch & Schaechter model developed during 60 s to assess the intrinsic sources of stochasticity that originate cell size heterogeneity in steady-state populations. The results reveal the importance of mass resolution to correctly describe these mechanisms.

Contact-mediated signaling enables disorder-driven transitions in cellular assemblies.

We show that, when cells communicate by contact-mediated interactions, heterogeneity in cell shapes and sizes leads to qualitatively distinct collective behavior in the tissue. For intercellular coupling that implements lateral inhibition, such disorder-driven transitions can substantially alter the asymptotic pattern of differentiated cells by modulating their fate choice through changes in the neighborhood geometry. In addition, when contact-induced signals influence inherent cellular oscillations, disorder leads to the emergence of functionally relevant partially-ordered dynamical states.

Abstract 770: Multiregional imaging and genomic analysis in high-grade IDH-mutant astrocytoma identifies intratumor subclones with aggressive features

Abstract High-grade gliomas represent the most common type of primary adult malignant brain tumor historically diagnosed and graded from histologic criteria alone. Gliomas harboring isocitrate dehydrogenase (IDH) 1 or 2 mutations, which are present in more than 80% of World Health Organization (WHO) grade 2 or 3 tumors, portend a favorable prognosis as compared to IDH-wildtype tumors. However, recent molecular profiling has identified the presence of additional alterations in these lower grade tumors that transform the biology toward an aggressive high-grade phenotype clinically that are independent of histologic grading and confer a worse clinical prognosis. Of these, the loss of CDKN2A/B is the strongest implicated alteration and is sufficient to define a grade 4 tumor in IDH-mutant astrocytomas regardless of histologic appearance. However, there remain no effective therapies targeted at molecular subgroups in aggressive gliomas to date. Here we have profiled the intratumoral heterogeneity via whole exome and RNA sequencing and copy number alteration analysis within a cohort of de-identified IDH-mutant astrocytoma patient tumors collected through the Mayo Clinic and Barrow Neurological Institute (BNI) biorepositories to determine the spatial evolution of aggressive molecular features. Within this cohort we have collected 42 image-localized intratumoral biopsy specimens representing 13 IDH-mutant patient tumors. We identified 11 biopsies from 3 tumors that harbored loss of CDKN2A/B which included 1 tumor that displayed intratumoral heterogeneity of CDKN2A/B expression across subclones. We performed an unsupervised cluster analysis to profile the transcriptional signatures in IDH-mutant astrocytomas across both differential CDKN2A/B status and tumor geographic location comparing contrast enhancing versus non-enhancing regions, and we analyzed the MRI features in IDH-mutant tumors across CDKN2A/B status. We found that tumors with CDKN2A/B alteration demonstrated lower signal intensity on delayed post-contrast T2*W images, which could represent lower pre-contrast T2*, unique contrast agent kinetics, or dissimilar diffusion properties related to increased cellular density and/or greater heterogeneity of cell size. Interestingly, we uncovered spatial heterogeneity of additional molecular drivers of high-grade glioma including KRAS, MYCN, and PDGFRA. While EGFR alterations are rarely reported in IDH-mutant tumors, we identified one treatment naïve patient tumor that harbored EGFR alteration only in the contrast non-enhancing region. Based on this work we propose there is early spatial evolution of aggressive tumor subclones prior to primary tumor diagnosis, and multiregional molecular profiling may better inform tumor biology and predict therapeutic strategies in IDH-mutant astrocytomas. Citation Format: Shannon P. Fortin Ensign, Fulvio D'Angelo, Francesca P. Caruso, Christopher Sereduk, Kyle Singleton, Jennifer Eschbacher, Gina Mazza, Maciej Mrugala, Alyx Porter, Bernard Bendok, Richard Zimmerman, Kris Smith, Peter Nakaji, Kamala Swanson, Junwen Wang, Nathenael Semmineh, Christopher Quarles, Anna Lasorella, Kristin Swanson, Michele Ceccarelli, Antonio Iavarone, Leland Hu, Nhan L. Tran. Multiregional imaging and genomic analysis in high-grade IDH-mutant astrocytoma identifies intratumor subclones with aggressive features [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 770.

Correlation between red blood cell distribution width, neutrophil to lymphocyte ratio, and neutrophil to platelet ratio with 3-month prognosis of patients with intracerebral hemorrhage: a retrospective study

BackgroundRed cell distribution width (RDW) is a parameter that indsicates the heterogeneity of red blood cell size and could be a prognostic factor in some diseases. Also, intracerebral hemorrhage (ICH) is considered a vascular event with a high mortality rate. We aimed to examine the role of RDW, neutrophil to lymphocyte ratio (NLR), and neutrophil to platelet ratio (NPR) in predicting the prognosis of patients with ICH.MethodsThis is a retrospective cohort study conducted on 140 patients with ICH admitted to the neurology ward and intensive care unit (ICU) in Imam Reza Hospital, Tabriz, Iran. Demographic data, National Institutes of Health Stroke Scale (NIHSS), and complete blood count test parameters were evaluated within 24 h after hospitalization. These variables were collected and re-evaluated three months later.ResultsThe mean age of the study population was 61.14 (± 16) years and 51% were male. The mean NLR (p = 0.05), neutrophil count (p=0.04), platelet count (p = 0.05), and NIHSS (p<0.01) had a significant difference between the deceased patients and those who partially recovered after three months. The ROC curve showed that NIHSS (area under curve (AUC): 0.902), followed by NPR (AUC: 0.682) variables had the highest AUC.ConclusionRDW could be a relevant prognostic factor and predictor in determining 3-months survival in ICH. Nevertheless, further large-scale prospective cohorts might be needed to evaluate the associations.

Towards a Better Characterisation of Leukemic Cells in Chronic Lymphocytic Leukaemia: Cell-Size Heterogeneity Reflects Their Activation Status and Migratory Abilities.

Simple SummaryChronic lymphocytic leukaemia (CLL) is a heterogeneous chronic disease characterised by the clonal expansion of mature CD19+CD23+CD5+ B-cells in blood, bone marrow and lymphoid tissue. Despite the CLL tumour cell population showing considerable heterogeneity in cell size, the functional characteristics of leukemic cells that differ in size have not been explored. The results of our study demonstrate that differences in CLL cell size reflect their activation state, polarisation and migratory capacity, with large CLL cells being more activated, polarised and motile than the small CLL cells present in the CLL cell pool. Our data provide evidence of the importance of cell-size heterogeneity within the CLL cell pool and the dynamics of cell-size changes for disease pathogenesis.Chronic lymphocytic leukaemia (CLL) is a genetically, morphologically and phenotypically heterogeneous chronic disease with clinical variability between patients. Whether the significant heterogeneity of cell size within the CLL population contributes to the heterogeneous features of this disease has not been investigated. The present study aimed to characterise the phenotypic and functional properties of two subpopulations of typical CLL cells that differ in cell size: small (s-CLL) and large (l-CLL) CLL cells delineated by forward scatter cytometry. The s-CLL cells were characterised by the CD5lowCXCR4hi phenotype, while the l-CLL cells were characterised by the CD5hiCXCR4dim phenotype and indicated a higher expression of CXCR3, CD20, CD38 and HLA-DR. The l-CLL cells displayed higher migration activity towards CXCL12, a tendency towards a higher proliferation rate and an increased capacity to produce IgM in the presence of CpG compared with s-CLL cells. When stimulated with CpG and CXCL12, l-CLL cells were characterised by a higher polarisation phenotype and motility than s-CLL cells. Our study revealed that the differences in CLL cell size reflected their activation status, polarisation and migratory abilities. Our data provide evidence of the importance of cell-size heterogeneity within a CLL pool and the dynamics of cell-size changes for disease pathogenesis, thus deserving further investigation.

High-Throughput Characterization of Cell Adhesion Strength Using Long-Channel Constriction-Based Microfluidics.

The adhesion strength of a cancer cell is a valuable biophysical marker of its metastatic potential, tightly associated with various metastatic processes; for example, cancer cells escape from a primary tumor, and circulating tumor cells (CTCs) are anchored to the vessel wall. Although constriction-based microfluidics can realize the high-throughput characterization of single-cell deformability, due to the influence of cell size heterogeneity, accurately evaluating the adhesion strength of a cancer cell at high throughputs in constriction remains difficult. In this paper, we first proposed an approach for the assessment of adhesion strength of BGC-823 and SGC-7901 cell lines at high throughputs based on a friction coefficient using the constant velocity stage of cell transit in a long-channel constriction. Cell size was proven to be independent of adhesion strength by cell detachment assay; however, it has large effects on cell transit velocity in constriction. Therefore, the linear elasticity of a completely deformed cell in constriction is simplified as a compressed spring model, effectively reducing the influence of cell size heterogeneity. Theoretically, our proposed model can well offset the influence of cell size by cell transit velocity, while our experimental results indicate that the friction coefficient has a good linear relationship with the logarithm of the adhesion strength too. Therefore, our proposed approach can realize accurate characterization of cell adhesion strength at high throughputs using long-channel constriction-based microfluidics. Hence, this work might enrich the functions of constriction-based microfluidics and bring new insights into the characterization of mechanical phenotypes.

Combined heterogeneity in cell size and deformability promotes cancer invasiveness.

Phenotypic heterogeneity is increasingly acknowledged to confer several advantages to cancer progression and drug resistance. Here, we probe the collective importance of heterogeneity in cell size and deformability in breast cancer invasion. A computational model of invasion of a heterogeneous cell aggregate predicts that combined heterogeneity in cell size and deformability enhances invasiveness of the whole population, with maximum invasiveness at intermediate cell-cell adhesion. We then show that small cells of varying deformability, a subpopulation predicted to be enriched at the invasive front, exhibit considerable overlap with the biophysical properties of cancer stem cells (CSCs). In MDA-MB-231 cells, these include CD44 hi CD24- mesenchymal CSCs, which are small and soft, and CD44 hi CD24+ hybrid CSCs, which exhibit a wide range of size and deformability. We validate our predictions by tracking the pattern of cell invasion from spheroids implanted in three-dimensional collagen gels, wherein we show temporal enrichment of CD44 hi cells at the invasive front. Collectively, our results illustrate the advantages imparted by biophysical heterogeneity in enhancing cancer invasiveness.This article has an associated First Person interview with the first author of the paper.

First person – Asadullah and Sandeep Kumar

First Person is a series of interviews with the first authors of a selection of papers published in Journal of Cell Science, helping early-career researchers promote themselves alongside their papers. Asadullah and Sandeep Kumar are co-first authors on 'Combined heterogeneity in cell size and deformability promotes cancer invasiveness', published in JCS. Asadullah is a PhD Student in the lab of Prof. Shamik Sen at BSBE, IIT Bombay, Mumbai, India, who is interested in combining computational methods along with biophysics to study disease biology. Sandeep conducted the research described in this article while a PhD Student in Dr Shamik Sen's lab. He is now an Entrepreneur in Residence at TandemLaunch Inc., Montreal, Canada, interested in developing new computational approaches to studying development and cancer growth.

Red Blood Cell Distribution Width/Hemoglobin Ratio Correlate with Severity of Clinical Status in Residents of Long-term Geriatric and Palliative Care

Background: The red blood cell distribution width (RDW) is a simple measure of red blood cell size heterogeneity. A high degree of anisocytosis, expressed by RDW &gt;15%, is observed in certain anemias, but also in other disorders, where RDW &gt;15% is associated with worse prognosis. We questioned whether the RDW/hemoglobin ratio (RDW/Hb) might closer relate to disease severity than RDW. Design: Cross sectional study Patients: 76 residents of two Department of Comprehensive Nursing Care. Method: Physicians classified the patients according to their clinical status in three groups: "stable", “severe-unstable”, and "intermediate". An outside observer extracted from the patients' files the results of RDW (elevated if &gt;15%), hemoglobin, iron, and transferrin. The RDW (%)/Hb (g/dL) ratio and tansferrin saturation (TSAT) were computed. The associations between RDW, RDW/Hb, TSAT, and the patients' clinical status were assessed. Results: In residents of Department A, RDW &gt;15% was found in 33% of 19 stable patients, in 58% of 10 patients with intermediate severity, and in 81% of 10 patients classified severe-unstable. The RDW/Hb ratio &gt;1.4 was found in 24% of stable patients, in 73% with intermediate severity, and in 93% of the severe-unstable patients. In Department B, 36 out of 38 patient's were classified stable: in 61.2 % the RDW was &gt;15% and in 38% the RDW/Hb was &gt;1.4. Conclusions: In a heterogenic population presenting multimorbidity, the RDW and to a higher degree the RDW/Hb (p &lt;0.00001), correlated with the patients' disease severity.

Variable cell division time and asymmetric division site lead to filament-to-rod cell cycle of Lysinibacillus varians.

All well-established cell size homeostasis paradigms are based on the researches of rod bacteria like B. subtilis and E. coli, suggesting a constant division time (timer model), division size (sizer model) or added size (adder model) before division. However, Lysinibacillus varians, a new species with regular filament-to-rod cell cycle, is inconsistent with existing models. In this study, the cell size parameters of the type strain GY32, were investigated by combing multiple microscopy techniques and single-cell approach. Our results showed that the filaments of strain GY32 were unicellular cells with multiple nucleoids. The division time of GY32 cells was variable and their daughter cells produced by asymmetric binary fission had different birth sizes, which were proportional to their elongation rates, resulting in high heterogeneity among the sister cells. Furthermore, the added size from birth to division was significantly shorter than birth size (p<0.01) and decreased along generations. The results above revealed that the asymmetric division site and varied cell size parameters resulted in filament-to-rod cell cycle of L. varians and cell size homeostasis could be a more complex and dynamic process than previously assumed. These findings would be helpful in elucidating the open questions in cell division and cell size heterogeneity.

Continuum descriptions of spatial spreading for heterogeneous cell populations: Theory and experiment

Variability in cell populations is frequently observed in both in vitro and in vivo settings. Intrinsic differences within populations of cells, such as differences in cell sizes or differences in rates of cell motility, can be present even within a population of cells from the same cell line. We refer to this variability as cell heterogeneity. Mathematical models of cell migration, for example, in the context of tumour growth and metastatic invasion, often account for both undirected (random) migration and directed migration that is mediated by cell-to-cell contacts and cell-to-cell adhesion. A key feature of standard models is that they often assume that the population is composed of identical cells with constant properties. This leads to relatively simple single-species homogeneous models that neglect the role of heterogeneity. In this work, we use a continuum modelling approach to explore the role of heterogeneity in spatial spreading of cell populations. We employ a three-species heterogeneous model of cell motility that explicitly incorporates different types of experimentally-motivated heterogeneity in cell sizes: (i) monotonically decreasing; (ii) uniform; (iii) non-monotonic; and (iv) monotonically increasing distributions of cell size. Comparing the density profiles generated by the three-species heterogeneous model with density profiles predicted by a more standard single-species homogeneous model reveals that when we are dealing with monotonically decreasing and uniform distributions a simple and computationally efficient single-species homogeneous model can be remarkably accurate in describing the evolution of a heterogeneous cell population. In contrast, we find that the simpler single-species homogeneous model performs relatively poorly when applied to non-monotonic and monotonically increasing distributions of cell sizes. Additional results for heterogeneity in parameters describing both undirected and directed cell migration are also considered, and we find that similar results apply.