Biophysical Signals Research Articles

High Resolution Imaging and Histopathological Characterization of Myocardial Infarction

Magnetic resonance imaging (MRI) can reveal subtle microstructural changes in the heart muscle following infarction, a major cause of sudden cardiac death worldwide. We have developed preclinical models of chronic infarction and used diffusion tensor methods to efficiently probe biophysical MR signals. Here our specific goal was to identify subtle characteristics of the peri-infarct zone, where the mixture of viable cells and collagen fibrils form arrhythmia substrate. We induced myocardial infarction in n=9 Yorkshire swine using an occlusion-reperfusion technique. We occluded the left circumflex artery (LCX) in 5 pigs and the left anterior descendent artery (LAD) in 4 pigs. After 5-6 weeks healing time, high resolution diffusion tensor imaging (∼0.6x0.6x1.2mm spatial resolution) was performed ex vivo in explanted hearts. We then calculated pixel-based fractional anisotropy (FA) in 3 zones: infarct core (dense fibrosis), peri-infarct (arrhythmia substrate) and remote/normal tissue. For analysis, we selected four regions of interests (ROIs) from each tissue zone. A total of 36 ROIs selected from DT images were analyzed and further registered via anatomical markers with corresponding ROIs in histopathology slides. For histology, tissue samples were stained with collagen-sensitive stain to evaluate fibrosis under bright field and polarized light microscopy, as well as CD31 (microvascular integrity) and electrical gap junctions (Cx43). Our results demonstrated a significant decrease in FA (∼25%) in peri-infarct compared to normal myocardium (p<0.05), in agreement with quantitative histopathological and immunohistochemistry findings which showed a reduced CD31 density (∼33%) and Cx43 density (∼56%). These results provide clear evidence of altered microstructure (i.e., fiber disarray due to collagen deposition) and function (i.e., reduced electrical connections and perfusion) in peri-infarct, in large animal models relevant to scar-related human pathophysiology. Future work will focus on using these novel findings to parameterize MRI-based models to predict arrhythmia inducibility.

Rapid translocation of pluripotency-related transcription factors by external uniaxial forces.

Human embryonic stem cells subjected to a one-time uniaxial stretch for as short as 30-min on a flexible substrate coated with Matrigel experienced rapid and irreversible nuclear-to-cytoplasmic translocation of NANOG and OCT4, but not Sox2. Translocations were directed by intracellular transmission of biophysical signals from cell surface integrins to nuclear CRM1 and were independent of exogenous soluble factors. On E-CADHERIN-coated substrates, presumably with minimal integrin engagement, mechanical strain-induced rapid nuclear-to-cytoplasmic translocation of the three transcription factors. These findings might provide fundamental insights into early developmental processes and may facilitate mechanotransduction-mediated bioengineering approaches to influencing stem cell fate determination.

Bio-Integrated Wearable Systems: A Comprehensive Review.

Bio-integrated wearable systems can measure a broad range of biophysical, biochemical, and environmental signals to provide critical insights into overall health status and to quantify human performance. Recent advances in material science, chemical analysis techniques, device designs, and assembly methods form the foundations for a uniquely differentiated type of wearable technology, characterized by noninvasive, intimate integration with the soft, curved, time-dynamic surfaces of the body. This review summarizes the latest advances in this emerging field of "bio-integrated" technologies in a comprehensive manner that connects fundamental developments in chemistry, material science, and engineering with sensing technologies that have the potential for widespread deployment and societal benefit in human health care. An introduction to the chemistries and materials for the active components of these systems contextualizes essential design considerations for sensors and associated platforms that appear in following sections. The subsequent content highlights the most advanced biosensors, classified according to their ability to capture biophysical, biochemical, and environmental information. Additional sections feature schemes for electrically powering these sensors and strategies for achieving fully integrated, wireless systems. The review concludes with an overview of key remaining challenges and a summary of opportunities where advances in materials chemistry will be critically important for continued progress.

Neuron/Glial Antigen 2-Type VI Collagen Interactions During Murine Temporomandibular Joint Osteoarthritis

The degeneration of articular cartilage underscores the clinical pathology of temporomandibular joint osteoarthritis (TMJ-OA) and is promoted through dysfunctional biochemical or biophysical signaling. Transduction of these signals has a multifaceted regulation that includes important cell-matrix derived interactions. The matrix encapsulating the cells of the mandibular condylar cartilage (MCC) is rich in type VI collagen. Neuron/glia antigen 2 (NG2) is a type I transmembrane proteoglycan that binds with type VI collagen. This study defines the temporospatial dynamics of NG2-type VI collagen interactions during the progression of TMJ-OA. Membrane-bound NG2 is found to colocalize with pericellular type VI collagen in superficial layer cells in the MCC perichondrium but is present at high levels in the cytosol of chondroblastic and hypertrophic cells. When TMJ -OA is induced using a surgical instability model, localized disruptions of pericellular type VI collagen are observed on the central and medial MCC and are associated with significantly higher levels of cytosolic NG2. NG2 localized within the cytosol is found to be transported through clathrin and dynamin mediated endocytic pathways. These findings are consistent with NG2 behavior in other injury models and underscore the potential of NG2 as an entirely novel molecular mechanism of chondrocyte function contextually linked with TMJ-OA.

A package for the computational analysis of complex biophysical signals

Electromyograms are biomedical signals which are detected through electrodes. These signals represent measurements of the electric potentials associated to muscles contractions, and they are physically important in view that they provide information on the health of individuals. The computational modeling of electromyograms has been mainly carried out through general-purpose commercial software, which is typically provided with the purchase of computer equipment. In view of the well-known limitations inherent in the commercial software, a lot of research and efforts are devoted continuously to upgrade existing processing methods. However, as expected, these innovations take a considerable amount of time in order to be implemented in commercial software. In order to alleviate this situation, the package has been developed to provide a free and fully open-source package for both applied and theoretical researchers, which includes some fundamental tools to computationally process electromyograms. The purpose of the present paper is to provide an overview of the most important features of this package, and to supply codes and examples to illustrate its use.

Biomaterial Engineering for Controlling Pluripotent Stem Cell Fate.

Pluripotent stem cells (PSCs) represent an exciting cell source for tissue engineering and regenerative medicine due to their self-renewal and differentiation capacities. The majority of current PSC protocols rely on 2D cultures and soluble factors to guide differentiation; however, many other environmental signals are beginning to be explored using biomaterial platforms. Biomaterials offer new opportunities to engineer the stem cell niches and 3D environments for exploring biophysical and immobilized signaling cues to further our control over stem cell fate. Here, we review the biomaterial platforms that have been engineered to control PSC fate. We explore how altering immobilized biochemical cues and biophysical cues such as dimensionality, stiffness, and topography can enhance our control over stem cell fates. Finally, we highlight biomaterial culture systems that assist in the translation of PSC technologies for clinical applications.

Spatio-Temporal Control of Cell Adhesion: Toward Programmable Platforms to Manipulate Cell Functions and Fate.

Biophysical and biochemical signals of material surfaces potently regulate cell functions and fate. In particular, micro- and nano-scale patterns of adhesion signals can finely elicit and affect a plethora of signaling pathways ultimately affecting gene expression, in a process known as mechanotransduction. Our fundamental understanding of cell-material signals interaction and reaction is based on static culturing platforms, i.e., substrates exhibiting signals whose configuration is time-invariant. However, cells in-vivo are exposed to arrays of biophysical and biochemical signals that change in time and space and the way cells integrate these might eventually dictate their behavior. Advancements in fabrication technologies and materials engineering, have recently enabled the development of culturing platforms able to display patterns of biochemical and biophysical signals whose features change in time and space in response to external stimuli and according to selected programmes. These dynamic devices proved to be particularly helpful in shedding light on how cells adapt to a dynamic microenvironment or integrate spatio-temporal variations of signals. In this work, we present the most relevant findings in the context of dynamic platforms for controlling cell functions and fate in vitro. We place emphasis on the technological aspects concerning the fabrication of platforms displaying micro- and nano-scale dynamic signals and on the physical-chemical stimuli necessary to actuate the spatio-temporal changes of the signal patterns. In particular, we illustrate strategies to encode material surfaces with dynamic ligands and patterns thereof, topographic relieves and mechanical properties. Additionally, we present the most effective, yet cytocompatible methods to actuate the spatio-temporal changes of the signals. We focus on cell reaction and response to dynamic changes of signal presentation. Finally, potential applications of this new generation of culturing systems for in vitro and in vivo applications, including regenerative medicine and cell conditioning are presented.

Inorganic semiconductor biointerfaces.

Biological systems respond to and communicate through biophysical cues, such as electrical, thermal, mechanical and topographical signals. However, precise tools for introducing localized physical stimuli and/or for sensing biological responses to biophysical signals with high spatiotemporal resolution are limited. Inorganic semiconductors display many relevant electrical and optical properties, and they can be fabricated into a broad spectrum of electronic and photonic devices. Inorganic semiconductor devices enable the formation of functional interfaces with biological material, ranging from proteins to whole organs. In this Review, we discuss fundamental semiconductor physics and operation principles, with a focus on their behaviour in physiological conditions, and highlight the advantages of inorganic semiconductors for the establishment of biointerfaces. We examine semiconductor device design and synthesis and discuss typical signal transduction mechanisms at bioelectronic and biophotonic interfaces for electronic and optoelectronic sensing, optoelectronic and photothermal stimulation and photoluminescent in vivo imaging of cells and tissues. Finally, we evaluate cytotoxicity and highlight possible new material components and biological targets of inorganic semiconductor devices.

Use of a PTFE Micro-Bioreactor to Promote 3D Cell Rearrangement and Maintain High Plasticity in Epigenetically Erased Fibroblasts.

Phenotype definition is driven by epigenetic mechanisms as well as directly influenced by the cell microenvironment and by biophysical signals deriving from the extracellular matrix. The possibility to interact with the epigenetic signature of an adult mature cell, reversing its differentiated state and inducing a short transient high plasticity window, was previously demonstrated. In parallel, in vitro studies have shown that 3D culture systems, mimicking cell native tissue, exert significant effects on cell behavior and functions. Here we report the production of "PTFE micro-bioreactors" for long-term culture of epigenetically derived high plasticity cells. The system promotes 3D cell rearrangement, global DNA demethylation and elevated transcription of pluripotency markers, that is dependent on WW domain containing transcription regulator 1 (TAZ) nuclear accumulation and SMAD family member 2 (SMAD2) co-shuttling. Our findings demonstrate that the use of 3D culture strategies greatly improves the induction and maintenance of a high plasticity state.

Tactile Chemomechanical Transduction Based on an Elastic Microstructured Array to Enhance the Sensitivity of Portable Biosensors

Tactile sensors capable of perceiving biophysical signals such as force, pressure, or strain have attracted extensive interest for versatile applications in electronic skin, noninvasive healthcare, and biomimetic prostheses. Despite these great achievements, they are still incapable of detecting bio/chemical signals that provide even more meaningful and precise health information due to the lack of efficient transduction principles. Herein, a tactile chemomechanical transduction strategy that enables the tactile sensor to perceive bio/chemical signals is proposed. In this methodology, pyramidal tactile sensors are linked with biomarker-induced gas-producing reactions, which transduce biomarker signals to electrical signals in real time. The method is advantageous as it enhances electrical signals by more than tenfold based on a triple-step signal amplification strategy, as compared to traditional electrical biosensors. It also constitutes a portable and general platform capable of quantifying a wide spectrum of targets including carcinoembryonic antigen, interferon-γ, and adenosine. Such tactile chemomechanical transduction would greatly broaden the application of tactile sensors toward bio/chemical signals perception which can be used in ultrasensitive portable biosensors and chemical-responsive chemomechanical systems.

The Developmental Shift of NMDA Receptor Composition Proceeds Independently of GluN2 Subunit-Specific GluN2 C-Terminal Sequences

SummaryThe GluN2 subtype (2A versus 2B) determines biophysical properties and signaling of forebrain NMDA receptors (NMDARs). During development, GluN2A becomes incorporated into previously GluN2B-dominated NMDARs. This “switch” is proposed to be driven by distinct features of GluN2 cytoplasmic C-terminal domains (CTDs), including a unique CaMKII interaction site in GluN2B that drives removal from the synapse. However, these models remain untested in the context of endogenous NMDARs. We show that, although mutating the endogenous GluN2B CaMKII site has secondary effects on GluN2B CTD phosphorylation, the developmental changes in NMDAR composition occur normally and measures of plasticity and synaptogenesis are unaffected. Moreover, the switch proceeds normally in mice that have the GluN2A CTD replaced by that of GluN2B and commences without an observable decline in GluN2B levels but is impaired by GluN2A haploinsufficiency. Thus, GluN2A expression levels, and not GluN2 subtype-specific CTD-driven events, are the overriding factor in the developmental switch in NMDAR composition.

Toward an integrated watershed zoning framework based on the spatio-temporal variability of land-cover and climate: Application in the Volta river basin

This article details a watershed regionalization approach which uses the concept of entropy in combination with the k-means clusters analysis. The regionalization approach aims to subdivide a watershed into relatively uniform zones based on the spatial variability of the local climate and land-covers. A case study is presented to illustrate the approach and outline the environmental implications of the outcomes. Especially, the study reports an application in the Volta river basin which is a transnational watershed, shared by six different countries in West Africa. Over years, the transboundary status of the Volta watershed seems to have exacerbated its environmental challenges, because the environmental policies in the six countries do not necessarily complement. Subsequently, it is desirable to envision unified scientific tools to support the management platform of the Volta basin. To date, the literature on the Volta has virtually neglected this aspect. Hence, this case study is timely as it intends to create a unified zoning system for the Volta river basin. In the study, formulations of entropy theory and k-means clustering were jointly applied to 16-years gridded time-series of monthly leaf area index, precipitation, and temperature across the Volta basin. Based on a clustering optimization criterion, a total of five zones were identified then the related land-cover and climatic patterns were comparatively analyzed. Significant environmental contrasts were diagnosed then specificities were pinpointed for each zone. A comparison of the new zones with an existing macro-scale ecoregion shows similarities which sustain the capacity of the regionalization approach to capturing meaningful biophysical signals. Hence, the zoning technique may be valued for further applications in environmental management.

An In Vitro Model for Assessing Corneal Keratocyte Spreading and Migration on Aligned Fibrillar Collagen.

Background: Corneal stromal cells (keratocytes) are responsible for developing and maintaining normal corneal structure and transparency, and for repairing the tissue after injury. Corneal keratocytes reside between highly aligned collagen lamellae in vivo. In addition to growth factors and other soluble biochemical factors, feedback from the extracellular matrix (ECM) itself has been shown to modulate corneal keratocyte behavior. Methods: In this study, we fabricate aligned collagen substrates using a microfluidics approach and assess their impact on corneal keratocyte morphology, cytoskeletal organization, and patterning after stimulation with platelet derived growth factor (PDGF) or transforming growth factor beta 1 (TGFβ). We also use time-lapse imaging to visualize the dynamic interactions between cells and fibrillar collagen during wound repopulation following an in vitro freeze injury. Results: Significant co-alignment between keratocytes and aligned collagen fibrils was detected, and the degree of cell/ECM co-alignment further increased in the presence of PDGF or TGFβ. Freeze injury produced an area of cell death without disrupting the collagen. High magnification, time-lapse differential interference contrast (DIC) imaging allowed cell movement and subcellular interactions with the underlying collagen fibrils to be directly visualized. Conclusions: With continued development, this experimental model could be an important tool for accessing how the integration of multiple biophysical and biochemical signals regulate corneal keratocyte differentiation.

Marine birds and mammals as ecosystem sentinels in and near Distributed Biological Observatory regions: An abbreviated review of published accounts and recommendations for integration to ocean observatories

Marine birds and mammals can act as sentinels to shifts in ocean ecosystems, due to their (i) reliance on finding aggregated prey for efficient foraging, (ii) need to respond to biophysical signals in support of successful seasonal migrations, and (iii) capacity to reflect changes in marine food webs. Here we present an abbreviated review of recent published accounts of marine bird and mammal responses to biophysical features of the ocean ecosystem in and near sampling regions of the Distributed Biological Observatory (DBO). The review is meant to provide both context for seabird surveys and marine mammal watches conducted in conjunction with DBO sampling and support for the inclusion of marine bird and mammal research, either by direct sampling or collaboration, in the protocols of other ocean observatories. The review is framed by the ocean processes imbedded in the Arctic Marine Pulses conceptual model, with the overarching goal to promote the routine integration of data on marine birds and mammals during the development of marine ecosystem models for Arctic regions.

MSX2 Initiates and Accelerates Mesenchymal Stem/Stromal Cell Specification of hPSCs by Regulating TWIST1 and PRAME

SummaryThe gap in knowledge of the molecular mechanisms underlying differentiation of human pluripotent stem cells (hPSCs) into the mesenchymal cell lineages hinders the application of hPSCs for cell-based therapy. In this study, we identified a critical role of muscle segment homeobox 2 (MSX2) in initiating and accelerating the molecular program that leads to mesenchymal stem/stromal cell (MSC) differentiation from hPSCs. Genetic deletion of MSX2 impairs hPSC differentiation into MSCs. When aided with a cocktail of soluble molecules, MSX2 ectopic expression induces hPSCs to form nearly homogeneous and fully functional MSCs. Mechanistically, MSX2 induces hPSCs to form neural crest cells, an intermediate cell stage preceding MSCs, and further differentiation by regulating TWIST1 and PRAME. Furthermore, we found that MSX2 is also required for hPSC differentiation into MSCs through mesendoderm and trophoblast. Our findings provide novel mechanistic insights into lineage specification of hPSCs to MSCs and effective strategies for applications of stem cells for regenerative medicine.

Orthosteric and allosteric action of the C5a receptor antagonists.

The C5a receptor (C5aR) is a G-protein-coupled receptor (GPCR) that can induce strong inflammatory response to the anaphylatoxin C5a. Targeting C5aR has emerged as a novel anti-inflammatory therapeutic method. However, developing potent C5aR antagonists as drugs has proven difficult. Here, we report two crystal structures of human C5aR in ternary complexes with the peptide antagonist PMX53 and a non-peptide antagonist, either avacopan or NDT9513727. The structures, together with other biophysical, computational docking and cell-based signaling data, reveal the orthosteric action of PMX53 and its effect of stabilizing the C5aR structure, as well as the allosteric action of chemically diverse non-peptide C5aR antagonists with different binding poses. Structural comparison analysis suggests the presence of similar allosteric sites in other GPCRs. We also discuss critical structural features of C5aR in activation, including a novel conformation of helix 8. On the basis of our results, we suggest novel strategies for developing C5aR-targeting drugs.

Membrane Potential Distinctly Modulates Mobility and Signaling of IL-2 and IL-15 Receptors in T Cells

The high electric field across the plasma membrane might influence the conformation and behavior of transmembrane proteins that have uneven charge distributions in or near their transmembrane regions. Membrane depolarization of T cells occurs in the tumor microenvironment and in inflamed tissues because of K+ release from necrotic cells and hypoxia affecting the expression of K+ channels. However, little attention has been given to the effect of membrane potential (MP) changes on membrane receptor function. Therefore, we studied the influence of membrane de- and hyperpolarization on the biophysical properties and signaling of interleukin-2 (IL-2) and interleukin-15 (IL-15) receptors, which play important roles in T cell function. We investigated the mobility, clustering, and signaling of these receptors and major histocompatibility complex (MHC) I/II glycoproteins forming coclusters in lipid rafts of T cells. Depolarization by high K+ buffer or K+ channel blockers resulted in a decrease in the mobility of IL-2Rα and MHC glycoproteins, as shown by fluorescence correlation spectroscopy, whereas hyperpolarization by the K+ ionophore valinomycin increased their mobility. Contrary to this, the mobility of IL-15Rα decreased upon both de- and hyperpolarization. These changes in protein mobility are not due to an alteration of membrane fluidity, as evidenced by fluorescence anisotropy measurements. Förster resonance energy transfer measurements showed that most homo- or heteroassociations of IL-2R, IL-15R, and MHC I did not change considerably, either. MP changes modulated signaling by the two cytokines in distinct ways: depolarization caused a significant increase in the IL-2-induced phosphorylation of signal transducer and activator of transcription 5, whereas hyperpolarization evoked a decrease only in the IL-15-induced signal. Our data imply that the MP may be an important modulator of interleukin receptor signaling and dynamics. Enhanced IL-2 signaling in depolarized Treg cells highly expressing IL-2R may contribute to suppression of antitumor immune surveillance.

The interacting role of physical stiffness and tumor cells on the macrophages polarization

There is a complex interplay existing between the tumor microenvironment and cellular function, while the influence of tumor biophysical signals on the macrophage function remain unclear. Herein, we constructed a three dimensional hydrogel model to study the symphysial effect of physical signal (stiffness) and biological signal (tumor cells) on macrophage polarization in different stage of tumor progression. In the absence of biological signal, the macrophage response was slightly changed with different stiffness (100 Pa, 300 Pa, and 900 Pa). Once co-incubated with the tumor cell, macrophage induced more significant cancer facilitated cytokines (e.g. IL-10, MMP-9) and chemokines (MCP-1). In contrast to the education role from the co-cultured cancer cells, a stiffness around the normal tissue (100 Pa, CO-L group) provided a physical cue for cancer cell regression. This cue triggered the active role of M1 type macrophages (e.g. IL-12), indicating the interacting role of stiffness in the presence of cancer cell. The stiffness around the premalignant tissue (M, 300 Pa) induced much higher level of cytokines (e.g. IL-6, IL-10, IL-12, CD86, and CD206), while the immune activation cytokines (M1 type) was largely attenuated at high stiffness (900 Pa). This study revealed the pleiotropic tuning of macrophage function under different biophysical signals, which provided important knowledge for cancer progress and may benefit the design of cancer-related therapies.

Characterization of Sensory-Motor Behavior Under Cognitive Load Using a New Statistical Platform for Studies of Embodied Cognition.

The field of enacted/embodied cognition has emerged as a contemporary attempt to connect the mind and body in the study of cognition. However, there has been a paucity of methods that enable a multi-layered approach tapping into different levels of functionality within the nervous systems (e.g., continuously capturing in tandem multi-modal biophysical signals in naturalistic settings). The present study introduces a new theoretical and statistical framework to characterize the influences of cognitive demands on biophysical rhythmic signals harnessed from deliberate, spontaneous and autonomic activities. In this study, nine participants performed a basic pointing task to communicate a decision while they were exposed to different levels of cognitive load. Within these decision-making contexts, we examined the moment-by-moment fluctuations in the peak amplitude and timing of the biophysical time series data (e.g., continuous waveforms extracted from hand kinematics and heart signals). These spike-trains data offered high statistical power for personalized empirical statistical estimation and were well-characterized by a Gamma process. Our approach enabled the identification of different empirically estimated families of probability distributions to facilitate inference regarding the continuous physiological phenomena underlying cognitively driven decision-making. We found that the same pointing task revealed shifts in the probability distribution functions (PDFs) of the hand kinematic signals under study and were accompanied by shifts in the signatures of the heart inter-beat-interval timings. Within the time scale of an experimental session, marked changes in skewness and dispersion of the distributions were tracked on the Gamma parameter plane with 95% confidence. The results suggest that traditional theoretical assumptions of stationarity and normality in biophysical data from the nervous systems are incongruent with the true statistical nature of empirical data. This work offers a unifying platform for personalized statistical inference that goes far beyond those used in conventional studies, often assuming a “one size fits all model” on data drawn from discrete events such as mouse clicks, and observations that leave out continuously co-occurring spontaneous activity taking place largely beneath awareness.

Similar Biophysical Abnormalities in Glomeruli and Podocytes from Two Distinct Models.

Background FSGS is a pattern of podocyte injury that leads to loss of glomerular function. Podocytes support other podocytes and glomerular capillary structure, oppose hemodynamic forces, form the slit diaphragm, and have mechanical properties that permit these functions. However, the biophysical characteristics of glomeruli and podocytes in disease remain unclear.Methods Using microindentation, atomic force microscopy, immunofluorescence microscopy, quantitative RT-PCR, and a three-dimensional collagen gel contraction assay, we studied the biophysical and structural properties of glomeruli and podocytes in chronic (Tg26 mice [HIV protein expression]) and acute (protamine administration [cytoskeletal rearrangement]) models of podocyte injury.Results Compared with wild-type glomeruli, Tg26 glomeruli became progressively more deformable with disease progression, despite increased collagen content. Tg26 podocytes had disordered cytoskeletons, markedly abnormal focal adhesions, and weaker adhesion; they failed to respond to mechanical signals and exerted minimal traction force in three-dimensional collagen gels. Protamine treatment had similar but milder effects on glomeruli and podocytes.Conclusions Reduced structural integrity of Tg26 podocytes causes increased deformability of glomerular capillaries and limits the ability of capillaries to counter hemodynamic force, possibly leading to further podocyte injury. Loss of normal podocyte mechanical integrity could injure neighboring podocytes due to the absence of normal biophysical signals required for podocyte maintenance. The severe defects in podocyte mechanical behavior in the Tg26 model may explain why Tg26 glomeruli soften progressively, despite increased collagen deposition, and may be the basis for the rapid course of glomerular diseases associated with severe podocyte injury. In milder injury (protamine), similar processes occur but over a longer time.