Biophysical Properties Of Cells Research Articles

Mesenchymal morphogenesis of embryonic stem cells dynamically modulates the biophysical microtissue niche

Stem cell fate and function are dynamically modulated by the interdependent relationships between biochemical and biophysical signals constituting the local 3D microenvironment. While approaches to recapitulate the stem cell niche have been explored for directing stem cell differentiation, a quantitative relationship between embryonic stem cell (ESC) morphogenesis and intrinsic biophysical cues within three-dimensional microtissues has not been established. In this study, we demonstrate that mesenchymal embryonic microtissues induced by BMP4 exhibited increased stiffness and viscosity accompanying differentiation, with cytoskeletal tension significantly contributing to multicellular stiffness. Perturbation of the cytoskeleton during ESC differentiation led to modulation of the biomechanical and gene expression profiles, with the resulting cell phenotype and biophysical properties being highly correlated by multivariate analyses. Together, this study elucidates the dynamics of biophysical and biochemical signatures within embryonic microenvironments, with broad implications for monitoring tissue dynamics, modeling pathophysiological and embryonic morphogenesis and directing stem cell patterning and differentiation.

Cell Surface Area and Membrane Folding in Glioblastoma Cell Lines Differing in PTEN and p53 Status

Glioblastoma multiforme (GBM) is characterized by rapid growth, invasion and resistance to chemo−/radiotherapy. The complex cell surface morphology with abundant membrane folds, microvilli, filopodia and other membrane extensions is believed to contribute to the highly invasive behavior and therapy resistance of GBM cells. The present study addresses the mechanisms leading to the excessive cell membrane area in five GBM lines differing in mutational status for PTEN and p53. In addition to scanning electron microscopy (SEM), the membrane area and folding were quantified by dielectric measurements of membrane capacitance using the single-cell electrorotation (ROT) technique. The osmotic stability and volume regulation of GBM cells were analyzed by video microscopy. The expression of PTEN, p53, mTOR and several other marker proteins involved in cell growth and membrane synthesis were examined by Western blotting. The combined SEM, ROT and osmotic data provided independent lines of evidence for a large variability in membrane area and folding among tested GBM lines. Thus, DK-MG cells (wild type p53 and wild type PTEN) exhibited the lowest degree of membrane folding, probed by the area-specific capacitance C m = 1.9 µF/cm2. In contrast, cell lines carrying mutations in both p53 and PTEN (U373-MG and SNB19) showed the highest C m values of 3.7–4.0 µF/cm2, which corroborate well with their heavily villated cell surface revealed by SEM. Since PTEN and p53 are well-known inhibitors of mTOR, the increased membrane area/folding in mutant GBM lines may be related to the enhanced protein and lipid synthesis due to a deregulation of the mTOR-dependent downstream signaling pathway. Given that membrane folds and extensions are implicated in tumor cell motility and metastasis, the dielectric approach presented here provides a rapid and simple tool for screening the biophysical cell properties in studies on targeting chemo- or radiotherapeutically the migration and invasion of GBM and other tumor types.

Changes in cytoskeletal dynamics and nonlinear rheology with metastatic ability in cancer cell lines

Metastatic outcome is impacted by the biophysical state of the primary tumor cell. To determine if changes in cancer cell biophysical properties facilitate metastasis, we quantified cytoskeletal biophysics in well-characterized human skin, bladder, prostate and kidney cell line pairs that differ in metastatic ability. Using magnetic twisting cytometry with optical detection, cytoskeletal dynamics was observed through spontaneous motion of surface bound marker beads and nonlinear rheology was characterized through large amplitude forced oscillations of probe beads. Measurements of cytoskeletal dynamics and nonlinear rheology differed between strongly and weakly metastatic cells. However, no set of biophysical parameters changed systematically with metastatic ability across all cell lines. Compared to their weakly metastatic counterparts, the strongly metastatic kidney cancer cells exhibited both increased cytoskeletal dynamics and stiffness at large deformation which are thought to facilitate the process of vascular invasion.

Margaret Gardel: At the interface of physics and biology

Gardel studies the biophysical properties of cells and cellular force generation.

Regulation of Cell Proliferation and Migration in Glioblastoma: New Therapeutic Approach

Glioblastoma is the most aggressive brain cancer with the poor survival rate. A microRNA, miR-451, and its downstream molecules, CAB39/LKB1/STRAD/AMPK, are known to play a critical role in regulating a biochemical balance between rapid proliferation and invasion in the presence of metabolic stress in microenvironment. We develop a novel multi-scale mathematical model where cell migration and proliferation are controlled through a core intracellular control system (miR-451-AMPK complex) in response to glucose availability and physical constraints in the microenvironment. Tumor cells are modeled individually and proliferation and migration of those cells are regulated by the intracellular dynamics and reaction-diffusion equations of concentrations of glucose, chemoattractant, extracellular matrix, and MMPs. The model predicts that invasion patterns and rapid growth of tumor cells after conventional surgery depend on biophysical properties of cells, dynamics of the core control system, and microenvironment as well as glucose injection methods. We developed a new type of therapeutic approach: effective injection of chemoattractant to bring invasive cells back to the surgical site after initial surgery, followed by glucose injection at the same location. The model suggests that a good combination of chemoattractant and glucose injection at appropriate time frames may lead to an effective therapeutic strategy of eradicating tumor cells.

Optical diagnostics of tumour cells at different stages of pathology development

The differences in optical and biophysical properties between the cells of mammary gland tumour extracted from tumours of different diameter are described. It is shown that the spectral and spectrokinetic properties of fluorescent probes in the cells extracted from the tumours in diameter are essentially different. Thus, the extinction coefficient of rhodamine 6G gradually increases with the pathology development. At the same time the rate of interaction of the triplet states of molecular probes with the oxygen, diluted in the tumour cells cytoplasm, decreases with the growth of the tumour capsule diameter. The observed regularities can be due to the changes in the cell structure, biochemical and biophysical properties. The reported data may be useful for developing optical methods of diagnostics of biotissue pathological conditions.

A hybrid model for cell proliferation and migration in glioblastoma

Glioblastoma is the most aggressive type of brain cancer with themedian survival time of one year. A particular microRNA, miR-451, and its counterpart,AMPK complex are known to play a key role in controlling a balance between rapid proliferationand aggressive invasion in response to metabolic stress in the microenvironment.The present paper develops a hybrid model of glioblastomathat identifiesa key mechanism behind the molecular switches between proliferative phase and migratoryphase in response to metabolic stress and biophysical interaction between cells.We focus on the core miR-451-AMPK control system and show how up- ordown-regulation of components in these pathways affects cell proliferation and migration.The model predicts the larger window of bistable systems when there exists a timedelay in the inhibitory pathway from CAB39/LKB1/STRAD/AMPK to miR-451.Delayed down-regulation of miR-451 along this pathway would let glioma cells staylonger in the proliferative stage despite relatively low glucose levels, making it a possibletherapeutic target.Analysis of the model predicts theexistence of a limit cycle with two time delays. We thenstudy a hybrid model for the biomechanical interaction between invasive and proliferative cells, inwhich all cells are modeled individually, and show how biophysical properties of cells andcore miR-451-AMPK control system affect the growth/invasion patterns of glioma spheroidsin response to various glucose levels in the microenvironment.The model predicts that cell migration not only depends on glucose availabilitybut also on mechanical constraints between cells. The model suggests that adhesion strengthbetween cells plays an important role in cell shedding from the main core and the disruptionof cell-cell adhesion is a pre-requisite for glioma cell invasion.The model also suggests that injection of glucose after surgery will increase visibility of individualmigratory cells and the second surgery may eradicate the remaining cancer cells, preventingregrowth of the invisible migratory glioma cells.

Subcellular spectroscopic markers, topography and nanomechanics of human lung cancer and breast cancer cells examined by combined confocal Raman microspectroscopy and atomic force microscopy

The nanostructures and hydrophobic properties of cancer cell membranes are important for membrane fusion and cell adhesion. They are directly related to cancer cell biophysical properties, including aggressive growth and migration. Additionally, chemical component analysis of the cancer cell membrane could potentially be applied in clinical diagnosis of cancer by identification of specific biomarker receptors expressed on cancer cell surfaces. In the present work, a combined Raman microspectroscopy (RM) and atomic force microscopy (AFM) technique was applied to detect the difference in membrane chemical components and nanomechanics of three cancer cell lines: human lung adenocarcinoma epithelial cells (A549), and human breast cancer cells (MDA-MB-435 with and without BRMS1 metastasis suppressor). Raman spectral analysis indicated similar bands between the A549, 435 and 435/BRMS1 including ~720 cm(-1) (guanine band of DNA), 940 cm(-1) (skeletal mode polysaccharide), 1006 cm(-1) (symmetric ring breathing phenylalanine), and 1451 cm(-1) (CH deformation). The membrane surface adhesion forces for these cancer cells were measured by AFM in culture medium: 0.478 ± 0.091 nN for A549 cells, 0.253 ± 0.070 nN for 435 cells, and 1.114 ± 0.281 nN for 435/BRMS1 cells, and the cell spring constant was measured at 2.62 ± 0.682 mN m(-1) for A549 cells, 2.105 ± 0.691 mN m(-1) for 435 cells, and 5.448 ± 1.081 mN m(-1) for 435/BRMS1 cells.

Biophysical Responses upon the Interaction of Nanomaterials with Cellular Interfaces

The explosion of study of nanomaterials in biological applications (the nano-bio interface) can be ascribed to nanomaterials' growing importance in diagnostics, therapeutics, theranostics (therapeutic diagnostics), and targeted modulation of cellular processes. However, a growing number of critics have raised concerns over the potential risks of nanomaterials to human health and safety. It is essential to understand nanomaterials' potential toxicity before they are tested in humans. These risks are complicated to unravel, however, because of the complexity of cells and their nanoscale macromolecular components, which enable cells to sense and respond to environmental cues, including nanomaterials. In this Account, we explore these risks from the perspective of the biophysical interactions between nanomaterials and cells. Biophysical responses to the uptake of nanomaterials can include conformational changes in biomolecules like DNA and proteins, and changes to the cellular membrane and the cytoskeleton. Changes to the latter two, in particular, can induce changes in cell elasticity, morphology, motility, adhesion, and invasion. This Account reviews what is known about cells' biophysical responses to the uptake of the most widely studied and used nanoparticles, such as carbon-based, metal, metal-oxide, and semiconductor nanomaterials. We postulate that the biophysical structure impairment induced by nanomaterials is one of the key causes of nanotoxicity. The disruption of cellular structures is affected by the size, shape, and chemical composition of nanomaterials, which are also determining factors of nanotoxicity. Currently, popular nanotoxicity characterizations, such as the MTT and lactate dehydrogenase (LDH) assays, only provide end-point results through chemical reactions. Focusing on biophysical structural changes induced by nanomaterials, possibly in real-time, could deepen our understanding of the normal and altered states of subcellular structures and provide useful perspective on the mechanisms of nanotoxicity. We strongly believe that biophysical properties of cells can serve as novel and noninvasive markers to evaluate nanomaterials' effect at the nano-bio interface and their associated toxicity. Better understanding of the effects of nanomaterials on cell structures and functions could help identify the required preconditions for the safe use of nanomaterials in therapeutic applications.

Epigenetic Modulation of the Biophysical Properties of Drug-Resistant Cell Lipids to Restore Drug Transport and Endocytic Functions

In our recent studies exploring the biophysical characteristics of resistant cell lipids, and the role they play in drug transport, we demonstrated the difference of drug-resistant breast cancer cells from drug-sensitive cells in lipid composition and biophysical properties, suggesting that cancer cells acquire a drug-resistant phenotype through the alteration of lipid synthesis to inhibit intracellular drug transport to protect from cytotoxic effect. In cancer cells, epigenetic changes (e.g., DNA hypermethylation) are essential to maintain this drug-resistant phenotype. Thus, altered lipid synthesis may be linked to epigenetic mechanisms of drug resistance. We hypothesize that reversing DNA hypermethylation in resistant cells with an epigenetic drug could alter lipid synthesis, changing the cell membrane's biophysical properties to facilitate drug delivery to overcome drug resistance. Herein we show that treating drug-resistant breast cancer cells (MCF-7/ADR) with the epigenetic drug 5-aza-2'-deoxycytidine (decitabine) significantly alters cell lipid composition and biophysical properties, causing the resistant cells to acquire biophysical characteristics similar to those of sensitive cell (MCF-7) lipids. Following decitabine treatment, resistant cells demonstrated increased sphingomyelinase activity, resulting in a decreased sphingomyelin level that influenced lipid domain structures, increased membrane fluidity, and reduced P-glycoprotein expression. Changes in the biophysical characteristics of resistant cell lipids facilitated doxorubicin transport and restored endocytic function for drug delivery with a lipid-encapsulated form of doxorubicin, enhancing the drug efficacy. In conclusion, we have established a new mechanism for efficacy of an epigenetic drug, mediated through changes in lipid composition and biophysical properties, in reversing cancer drug resistance.

Biophysical Assessment of Single Cell Cytotoxicity: Diesel Exhaust Particle-Treated Human Aortic Endothelial Cells

Exposure to diesel exhaust particles (DEPs), a major source of traffic-related air pollution, has become a serious health concern due to its adverse influences on human health including cardiovascular and respiratory disorders. To elucidate the relationship between biophysical properties (cell topography, cytoskeleton organizations, and cell mechanics) and functions of endothelial cells exposed to DEPs, atomic force microscope (AFM) was applied to analyze the toxic effects of DEPs on a model cell line from human aortic endothelial cells (HAECs). Fluorescence microscopy and flow cytometry were also applied to further explore DEP-induced cytotoxicity in HAECs. Results revealed that DEPs could negatively impair cell viability and alter membrane nanostructures and cytoskeleton components in a dosage- and a time-dependent manner; and analyses suggested that DEPs-induced hyperpolarization in HAECs appeared in a time-dependent manner, implying DEP treatment would lead to vasodilation, which could be supported by down-regulation of cell biophysical properties (e.g., cell elasticity). These findings are consistent with the conclusion that DEP exposure triggers important biochemical and biophysical changes that would negatively impact the pathological development of cardiovascular diseases. For example, DEP intervention would be one cause of vasodilation, which will expand understanding of biophysical aspects associated with DEP cytotoxicity in HAECs.

Hydroxyl radical scavenging mechanism of human erythrocytes by quercetin–germanium (IV) complex

Quercetin is a popular flavonoid in plant foods, herbs, and dietary supplement. Germanium, a kind of trace elements, can enhance the body immunity. This study investigated the hydroxyl-radical-scavenging mechanism of the quercertin–germanium (IV) (Qu–Ge) complex to human erythrocytes, especially the effects on ultrastructure and mechanical properties of cell membrane, plasma membrane potential and intracellular free Ca2+ concentration. Results showed that QuGe2, a kind of the Qu–Ge complex, could reduce the oxidative damage of erythrocytes, change the cell-surface morphology, and partly recover the disruption of plasma membrane potential and intracellular free Ca2+ level. Atomic force microscopy (AFM) was used to characterize the changes of the cell morphology, cell-membrane ultrastructure and biophysical properties at nanoscalar level. QuGe2 has triggered the antioxidative factor to inhibit cellular damage. These results can improve the understanding of hydroxyl-radical-scavenging mechanism of human erythrocytes induced by the Qu–Ge complex, which can be potentially developed as a new antioxidant for treatment of oxidative damage.

Abstract 2374: Performance of a new generation antibody-independent ApoStream™ platform for the enrichment of circulating tumor cells (CTCs)

Abstract Background: Isolation and molecular analysis of Circulating Tumor Cells (CTCs) is technically challenging, but may replace tumor biopsy by providing a minimally-invasive, blood-based, “liquid biopsy” method to monitor metastatic disease progression and guide cancer therapy. The use of currently available EpCAM-based enrichment platforms limits the type of tumor cells that can be recovered. We have developed the first robust CTC isolation device, ApoStream™, which does not rely on specific surface proteins or the size of CTCs. Specific Aims: We demonstrate the performance of a novel antibody-independent dielectrophoretic field-flow fractionation (DEP-FFF) based CTC isolation platform: ApoStream™. Methods: In the ApoStream™ device, DEP-FFF is used to differentially drive cell movement based on different biophysical properties of cells in a continuous flow microfluidic chamber to isolate CTCs from peripheral blood mononuclear cells (PBMCs). To demonstrate performance, pre-labeled SKOV3 ovarian cancer cells (low EpCAM expressing) were spiked into buffer or PBMCs and isolated through ApoStream™ device. Cancer cell recovery, system linearity, and precision were demonstrated. Viability of cancer cells isolated through the ApoStream™ platform, as well as their ability to be propagated in culture, was confirmed. Results: On three different days, the ApoStream™ breadboard device recovered 83.0± 3.0% (mean ± SD), 76.3 ± 2.6%, and 81.5± 1.0% of SKOV3 cells spiked into buffer (average interday recovery of 80.3 ± 3.5%, CV% = 4.3). Additionally, 79.0 ± 1.5%, 81.2 ± 7.8% and 75.2 ± 4.3% of SKOV3 cells spiked into ∼10 million normal PBMCs (average interday recovery 78.5 ± 3.0%, CV% = 3.3) were recovered. When SKOV3 spiking recovery was measured on two different ApoStream™ devices, the CV% was 0.1%. System linearity was evaluated by spiking a range of 4 - 2,600 SKOV3 cells into 10 million PBMCs. The regression analysis for the linearity resulted in a correlation coefficient (R2) of 1.00. Cell viability was not affected by the ApoStream™ device, and was 98.2, 98.8 and 100% of initial viability at 0, 16 and 24 hours after ApoStream™ separation, respectively, as measured by Trypan blue exclusion test conducted on cells propagated in tissue culture. Conclusion: We developed the first antigen-independent and cell size independent device for isolation of CTCs from patient blood with a wide variety of cancers. ApoStream™ technology performance characterization demonstrated high recovery of EpCAM-negative cultured cancer cells over a significant range of spiked cell numbers, as well as excellent linearity performance. Isolation of large numbers of live and unfixed CTCs with the ApoStream™ platform enables downstream molecular characterization of CTCs for the development of personalized treatment options for cancer patients. Acknowledgments: Funded by NCI Contract No. HHSN261200800001E. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2374. doi:1538-7445.AM2012-2374

N-glycosylation of the mammalian dipeptidyl aminopeptidase-like protein 10 (DPP10) regulates trafficking and interaction with Kv4 channels

The dipeptidyl aminopeptidase-like protein 10 (DPP10) is a type II transmembrane protein homologue to the serine protease DPPIV/CD26 but enzymatically inactive. In the mammalian brain, DPP10 forms a complex with voltage-gated potassium channels of the Kv4 family, regulating their cell surface expression and biophysical properties. DPP10 is a glycoprotein containing eight predicted N-glycosylation sites in the extracellular domain. In this study we investigated the role of N-glycosylation on DPP10 trafficking and functional activity. Using site-directed mutagenesis (N to Q) we showed that N-glycosylation occured at six positions. Glycosylation at these specific residues was necessary for DPP10 trafficking to the plasma membrane as observed by flow cytometry. The surface expression levels of the substitutions N90Q, N119Q, N257Q and N342Q were reduced by more than 60%. Hence the interaction with the Kv4.3/KChIP2a channel complex was disrupted preventing the hastening effect of wild type DPP10 on current kinetics. Interestingly, N257 was crucial for this function and its substitution to glutamine completely blocked DPP10 sorting to the cell surface and prevented DPP10 dimerization. In summary, we demonstrated that glycosylation was necessary for both DPP10 trafficking to the cell surface and functional interaction with Kv4 channels.

Optical Tweezers as a New Biomedical Tool to Measure Zeta Potential of Stored Red Blood Cells

During storage, red blood cells (RBCs) for transfusion purposes suffer progressive deterioration. Sialylated glycoproteins of the RBC membrane are responsible for a negatively charged surface which creates a repulsive electrical zeta potential. These charges help prevent the interaction between RBCs and other cells, and especially among each RBCs. Reports in the literature have stated that RBCs sialylated glycoproteins can be sensitive to enzymes released by leukocyte degranulation. Thus, the aim of this study was, by using an optical tweezers as a biomedical tool, to measure the zeta potential in standard RBCs units and in leukocyte reduced RBC units (collected in CPD-SAGM) during storage. Optical tweezers is a sensitive tool that uses light for measuring cell biophysical properties which are important for clinical and research purposes. This is the first study to analyze RBCs membrane charges during storage. In addition, we herein also measured the elasticity of RBCs also collected in CPD-SAGM. In conclusion, the zeta potential decreased 42% and cells were 134% less deformable at the end of storage. The zeta potential from leukodepleted units had a similar profile when compared to units stored without leukoreduction, indicating that leukocyte lyses were not responsible for the zeta potential decay. Flow cytometry measurements of reactive oxygen species suggested that this decay is due to membrane oxidative damages. These results show that measurements of zeta potentials provide new insights about RBCs storage lesion for transfusion purposes.

Bone cell elasticity and morphology changes during the cell cycle

The mechanical properties of cells are reported to be regulated by a range of factors including interactions with the extracellular environment and other cells, differentiation status, the onset of pathological states, as well as the intracellular factors, for example, the cytoskeleton. The cell cycle is considered to be a well-ordered sequence of biochemical events. A number of processes reported to occur during its progression are inherently mechanical and, as such, require mechanical regulation. In spite of this, few attempts have been made to investigate the putative regulatory role of the cell cycle in mechanobiology. In the present study, Atomic Force Microscopy (AFM) was employed to investigate the elastic modulus of synchronised osteoblasts. The data obtained confirm that osteoblast elasticity is regulated by cell cycle phase; specifically, cells in S phase were found to have a modulus approximately 1.7 times that of G1 phase cells. Confocal microscopy studies revealed that aspects of osteoblast morphology, namely F-actin expression, were also modulated by the cell cycle, and tended to increase with phase progression from G0 onwards. The data obtained in this study are likely to have implications for the fields of tissue- and bio-engineering, where prior knowledge of cell mechanobiology is essential for the effective replacement and repair of tissue. Furthermore, studies focused on biomechanics and the biophysical properties of cells are important in the understanding of the onset and progression of disease states, for example cancer at the cellular level. Our study demonstrates the importance of the combined use of traditional and relatively novel microscopy techniques in understanding mechanical regulation by crucial cellular processes, such as the cell cycle.

Biophysical properties of Saccharomyces cerevisiae and their relationship with HOG pathway activation

Parameterized models of biophysical and mechanical cell properties are important for predictive mathematical modeling of cellular processes. The concepts of turgor, cell wall elasticity, osmotically active volume, and intracellular osmolarity have been investigated for decades, but a consistent rigorous parameterization of these concepts is lacking. Here, we subjected several data sets of minimum volume measurements in yeast obtained after hyper-osmotic shock to a thermodynamic modeling framework. We estimated parameters for several relevant biophysical cell properties and tested alternative hypotheses about these concepts using a model discrimination approach. In accordance with previous reports, we estimated an average initial turgor of 0.6 ± 0.2 MPa and found that turgor becomes negligible at a relative volume of 93.3 ± 6.3% corresponding to an osmotic shock of 0.4 ± 0.2 Osm/l. At high stress levels (4 Osm/l), plasmolysis may occur. We found that the volumetric elastic modulus, a measure of cell wall elasticity, is 14.3 ± 10.4 MPa. Our model discrimination analysis suggests that other thermodynamic quantities affecting the intracellular water potential, for example the matrix potential, can be neglected under physiological conditions. The parameterized turgor models showed that activation of the osmosensing high osmolarity glycerol (HOG) signaling pathway correlates with turgor loss in a 1:1 relationship. This finding suggests that mechanical properties of the membrane trigger HOG pathway activation, which can be represented and quantitatively modeled by turgor.Electronic supplementary materialThe online version of this article (doi:10.1007/s00249-010-0612-0) contains supplementary material, which is available to authorized users.

Coordination of plant cell division and expansion in a simple morphogenetic system

Morphogenesis in plants arises from the interplay of genetic and physical interactions within a growing network of cells. The physical aspects of cell proliferation and differentiation are genetically regulated, but constrained by mechanical interactions between the cells. Higher plant tissues consist of an elaborate three-dimensional matrix of active cytoplasm and extracellular matrix, where it is difficult to obtain direct measurements of geometry or cell interactions. To properly understand the workings of plant morphogenesis, it is necessary to have biological systems that allow simple and direct observation of these processes. We have adopted a highly simplified plant system to investigate how cell proliferation and expansion is coordinated during morphogenesis. Coleocheate scutata is a microscopic fresh-water green alga with simple anatomical features that allow for accurate quantification of morphogenetic processes. Image analysis techniques were used to extract precise models for cell geometry and physical parameters for growth. This allowed construction of a deformable finite element model for growth of the whole organism, which incorporated cell biophysical properties, viscous expansion of cell walls, and rules for regulation of cell behavior. The study showed that a simple set of autonomous, cell-based rules are sufficient to account for the morphological and dynamic properties of Coleochaete growth. A variety of morphogenetic behavior emerged from the application of these local rules. Cell shape sensing is sufficient to explain the patterns of cell division during growth. This simplifying principle is likely to have application in modeling and design for engineering of higher plant tissues.

Dielectrophoretic-Field Flow Fractionation Analysis of Dielectric, Density, and Deformability Characteristics of Cells and Particles

Dielectrophoretic field-flow fractionation (DEP-FFF) has been used to discriminate between particles and cells based on their dielectric and density properties. However, hydrodynamic lift forces (HDLF) at flow rates needed for rapid separations were not accounted for in the previous theoretical treatment of the approach. Furthermore, no method was developed to isolate particle or cell physical characteristics directly from DEP-FFF elution data. An extended theory of DEP-FFF is presented that accounts for HDLF. With the use of DS19 erythroleukemia cells as model particles with frequency-dependent dielectric properties, it is shown that the revised theory accounts for DEP-FFF elution behavior over a wide range of conditions and is consistent with sedimentation-FFF when the DEP force is zero. Conducting four elution runs under specified conditions, the theory allows for the derivation of the cell density distribution and provides good estimates of the distributions of the dielectric properties of the cells and their deformability characteristics that affect HDLF. The approach allows for rapid profiling of the biophysical properties of cells, the identification and characterization of subpopulations, and the design of optimal DEP-FFF separation conditions. The extended DEP-FFF theory is widely applicable, and the parameter measurement methods may be adapted easily to other types of particles.

A Novel Approach to High Accuracy of Video-Based Microrheology

Video-based particle tracking monitors the microscopic movement of labeled biomolecules and fluorescent probes within a complex cellular environment. Information gained from this technique enables us to extract the dynamic behavior of biomolecules and the local mechanical properties inside the cell from a tracked particle's mean-square displacement (MSD). However, MSD measurements are highly susceptible to static error introduced by noise in the image acquisition process that leads to an incorrect positioning of the particle. Static error can mask the subtle effects from the local microenvironment on the MSD and potentially generate misleading conclusions about the biophysical properties of cells. An approach that greatly increases the accuracy of MSD measurements is presented herein by combining experimental data with Monte Carlo simulations to eliminate the inherent static error. This practical method of static error correction greatly advances particle-tracking techniques.