Biology subjects

Gundiah, N.

Publications and source records attributed to Gundiah, N..

11 recordsLinked to original sources

bioRxiv2026

Cell Mechanics Regulate Membrane Tubulation-Driven Trogocytosis in Adherent Cells

Trogocytosis is a widespread and physiologically important process in which one cell ingests fragments of another. It has been observed in diverse biological contexts, such as in Entamoeba histolytica, a human enteric parasite that nibbles host intestinal cells via trogocytosis, thereby invading the tissue and leading to lethal extra-intestinal disease, and in macrophages on interaction with adherent tumor monolayers. Despite its physiological relevance, knowledge of trogocytosis with adherent cells remains elusive. Here, we established a monolayer-based trogocytosis assay and uncovered a distinct membrane-tube-mediated trogocytic mechanism operative in adherent cells. We characterized it into four sequential events: contact, membrane tubulation, stretching, and scission. By targeting the proteic constituents of target cell stiffness, we show that trogocytic output exhibits a non-monotonic dependence on the viscoelastic properties of the target cell, with maximal uptake occurring in an intermediate mechanical regime. Further, we integrated live-cell trogocytosis observations with a viscoelastic model-based theoretical framework to propose a tube-breaking mechanism across distinct mechanical subtypes. Finally, using micropatterning, we demonstrated that target cell shape influences trogocytosis kinetics. This work suggests that trogocytosis is sensitive to the mechanical state of the target cell.

cell biology↗

bioRxiv2025

Substrate viscoelasticity regulates fibroblast adhesion and migration

Mechanical properties of the extracellular matrix (ECM) modulate cell-substrate interactions and influence cellular behaviors such as contractility, migrations, and proliferation. Although the effects of substrate stiffness on mechanobiology have been well studied, the role of ECM viscoelasticity in fibrotic progression remains less understood. To examine how viscoelasticity affects the biophysical properties and regulates the signaling of human mammary fibroblasts, we engineered elastic (E) and viscoelastic (VE) polyacrylamide hydrogels with comparable storage moduli ([~]14.52 {+/-} 1.03 kPa) but distinctly different loss moduli. Fibroblasts cultured on E hydrogels spread extensively (2428.93 {+/-} 864.71 m{superscript 2}), developed prominent stress fibers with higher zyxin intensity, and generated higher traction stresses (2931.57 {+/-} 1732.61 Pa). In contrast, fibroblasts on VE substrates formed smaller focal adhesion areas (54.2% reduction), exhibited lower critical adhesion strengths (51.8%), and generated 21% lower traction stresses (p < 0.001), indicating weaker adhesions. These substrates also promoted migrations and showed enhanced proliferation accompanied by reduced YAP activity, suggesting a mechanotransduction shift that may involve alternative signaling pathways. In contrast, E substrates showed YAP nuclear translocation, consistent with greater cytoskeletal tension and contractility. These findings highlight the importance of energy dissipation mechanisms in regulating fibroblast function on substrates mimicking the fibrotic milieu. Our results demonstrate that tuning the ECM viscoelasticity is a useful strategy to regulate cell behaviors in tissue engineered scaffolds, and develop better disease modeling for regenerative medicine.

bioengineering↗

bioRxiv2025

Mechanotransduction and Biophysical Regulation of Fibroblast Plasticity Under Prolonged Shear Stress

Complex interactions between mechanical stimuli and biochemical signaling shape the evolving fibrotic milieu. Fibroblasts contribute to this process, transitioning into myofibroblasts on stiffer substrates, and under low shear in 3D environments. The time-dependent effects of prolonged physiological shear stress on fibroblasts however remain poorly understood. We demonstrate that sustained shear stress (1.2 Pa) over 12 hours induces a temporal cascade of signaling and biophysical changes, driving fibroblast trans-differentiation into myofibroblasts. Cells adopt a spindle-shaped morphology under shear, actively remodeling through actin and -SMA to form elongated stress fibers. Myofibroblasts exhibit increased vinculin and zyxin expression in mature adhesions that remain elevated post-shear. Traction stresses progressively increased over the shear duration and persisted for 12 hours after the removal of the stimulus, indicating mechanical memory. Post-shear, deadhesion strengths increased significantly, accompanied by shifts in gene expressions governing extracellular matrix homeostasis. Prolonged shear enhanced MSD, directionality, and migration speed, all crucial for effective tissue repair. Finally, we demonstrate temporal changes in the biochemical gene expressions mediating the shear-induced remodeling of the extracellular matrix, cytoskeleton, and focal adhesions during the myofibroblast transition. These findings provide insights into the mechanobiological factors driving fibroblast-to-myofibroblast transitions which may inform the development of mechano-therapeutic strategies.

bioengineering↗

bioRxiv2023

Controlled disturbed flows on the endothelium result in increased inflammation, nuclear lamin disruption and heterochromatin condensation

Early atherosclerotic lesions often develop in areas of disturbed flow within arterial curvatures and bifurcations. Complex flows alter the signaling of inflammatory and other signaling molecules that may exacerbate the disease phenotype. Microfluidic platforms to assess changes in endothelial mechanobiology generally employ laminar unidirectional or oscillatory flows generated in straight channels; such platforms do not however mimic the complex time-varying bi-directional shear patterns on arterial walls. We fabricated an endothelium-on-chip device to generate "controlled" disturbed flows, characterized using a wall shear rosette, such as those reported in aneurysmal vessels and in regions with atherosclerotic plaques. We cultured human aortic endothelial cells (HAEC) in the device and subjected the monolayer to a circular shear rosette which represents bidirectional and oscillatory shear stresses. Immunofluorescence results show large cuboidal cells with significantly higher nuclear areas, changes in the localization of VE-Cadherin, elevated actin and NF-kB expressions in the device as compared to cell monolayers subjected to laminar flow, unidirectional oscillatory flow, and No-flow conditions. The creation of a dysfunctional endothelial monolayer due to bidirectional oscillatory flows also correlated with dramatic changes to the lamin A/C distribution and heterochromatin organization in the nucleus that have not been reported earlier. Such studies are potentially useful to assess novel therapeutics, mitigate effects of vascular disease for personalized medicine, study thrombus creation in the vicinity of inflamed endothelial monolayers, and reduce ourreliance on animal trials. Our device and analytical method represent the first successful demonstration of generating controlled disturbed flows in microfluidic devices

bioengineering↗

bioRxiv2023

Chimeric identities and reduced stiffness characterise the shoot apex of Arabidopsis stem cell mutants

Stem cell homeostasis in the shoot apical meristem involves a core regulatory feedback loop between the signalling peptide CLAVATA3, produced in stem cells, and the transcription factor WUSCHEL, expressed in the underlying organising centre. clavata mutants display massive meristem overgrowth, which is thought to be caused by stem cell overproliferation, although it is unknown how uncontrolled stem cell divisions lead to this altered morphology. Here we first reveal local buckling defects in mutant meristems, and use analytical models to show how mechanical properties and growth rates may contribute to the phenotype. Indeed, clavata meristems are mechanically more heterogeneous than the wild type, and also display regional growth heterogeneities. Furthermore, stereotypical wild-type meristem organisation is lost in mutants, in which cells simultaneously express distinct fate markers. Finally, cells in mutant meristems are auxin responsive, suggesting that they are functionally different from wild-type stem cells. Thus all benchmarks show that clavata meristem cells are different from wild-type stem cells, suggesting that fasciation is caused by the disruption of a more complex regulatory framework that maintains distinct genetic and functional domains at the shoot apex. Summary statementHeterogeneities in cell mechanics, growth, function and identity contribute to buckling in clavata mutant shoot apices.

plant biology↗

bioRxiv2022

Golgi localized Arl15 regulates cargo transport, cell adhesion and motility

Arf-like GTPases (Arls) regulate membrane trafficking and cytoskeletal organization. Genetic studies predicted a role for Arl15 in type-2 diabetes, insulin resistance, adiposity, and rheumatoid arthritis. Recent studies indicate a possible role for Arl15 in multiple physiological processes, including magnesium homeostasis. However, the molecular function of Arl15 is poorly defined. We evaluated the role of Arl15 in vesicular transport using techniques to quantify cargo trafficking, to mechanobiology. Fluorescence microscopy of stably expressing Arl15-GFP HeLa cells showed its localization to the Golgi and cell surface, including filopodia, and a cohort to recycling endosomes. The dissociation of Golgi, using small molecular inhibitors or the expression of Arf1 dominant-negative mutant, completely mislocalized Arl15 to the cytosol. Interestingly, site-directed mutagenesis analysis identified a novel V80A mutation in the GTP-binding domain that turns Arl15 into a dominant-negative form with reduced number of filopodia. Depletion of Arl15 in HeLa cells caused mislocalization of cargo, such as caveolin-2 and STX6, from the Golgi. Arl15 knockdown cells displayed reduced filopodial number, altered focal adhesion kinase organization, and enhanced soluble and receptor-mediated cargo uptake without affecting the TfR recycling. Arl15 knockdown decreased cell migration and enhanced cell spreading and adhesion strength. Traction force microscopy experiments revealed that Arl15 depleted cells exert higher tractions and generate multiple focal adhesion points during the initial phase of cell adhesion as compared to control cells. Collectively, these studies demonstrated a functional role for Arl15 in the Golgi, which includes regulating cargo transport to organize membrane domains at the cell surface. Key pointsO_LIArl15 primarily localizes to Golgi and plasma membrane, including filopodia C_LIO_LIMembrane localization of Arl15 is dependent on Golgi integrity or Arf1 activation C_LIO_LIArl15 knockdown mislocalizes STX6-dependent Golgi localized cargo required for cell surface organization and reduces the filopodial number C_LIO_LIArl15 is involved in cell spreading, adhesion, and migration C_LI

cell biology↗

bioRxiv2022

Cell adhesion strength and tractions are mechano-diagnostic features of cellular invasiveness

The adhesion of cells to substrates occurs via integrin clustering and binding to the actin cytoskeleton. Oncogenes modify anchorage-dependent mechanisms in cells during cancer progression. Fluid shear devices provide a label-free, non-invasive way to characterize cell-substrate interactions and heterogeneities in the cell populations. We quantified the critical adhesion strengths of MCF7, MDAMB-231, A549, HPL1D, HeLa, and NIH3T3 cells using a custom fluid shear device. The detachment response was sigmoidal for each cell type. A549 and MDAMB-231 cells had significantly lower adhesion strengths at {tau}50 than their non-invasive counterparts, HPL1D and MCF7. Detachment dynamics was inversely correlated with cell invasion potentials. A theoretical model, based on {tau}50 values and the distribution of cell areas on substrates, provided good fits to data from de-adhesion experiments. Quantification of cell tractions, using the Reg-FTTC method on 10 kPa polyacrylamide gels, showed highest values for invasive, MDAMB-231 and A549, cells compared to non-invasive cells. Immunofluorescence studies show differences in vinculin distributions: non-invasive cells have distinct vinculin puncta, whereas invasive cells have more dispersed distributions. The cytoskeleton in non-invasive cells was devoid of well-developed stress fibers, and had thicker cortical actin bundles in the boundary. These correlations in adhesion strengths with cell invasiveness, demonstrated here, may be useful in cancer diagnostics and other pathologies featuring misregulation in adhesion.

bioengineering↗

bioRxiv2021

Substrate stiffness modulates integrin α5 expression and ECM-associated gene expression in fibroblasts

Biomaterials, like polydimethylsiloxane (PDMS), are soft, biocompatible, and tuneable, which makes them useful to delineate specific substrate factors that regulate the complex landscape of cell-substrate interactions. We used a commercial formulation of PDMS to fabricate substrates with moduli 40 kPa, 300 kPa, and 1.5 MPa, and cultured HMF3S fibroblasts on them. Gene expression analysis was performed by RT-PCR and Western blotting. Cellular and nuclear morphologies were analyzed using confocal imaging, and MMP-2 and MMP-9 activities were determined with gelatin zymography. Results, comparing mechanotransduction on PDMS substrates with control petridishes, show that substrate stiffness modulates cell morphologies and proliferations. Cell nuclei were rounded on compliant substrates and correlated with increased tubulin expression. Proliferations were higher on stiffer substrates with cell cycle arrest on softer substrates. Integrin 5 expression decreased on lower stiffness substrates, and correlated with inefficient TGF-{beta} activation. Changes to the activated state of the fibroblast on higher stiffness substrates were linked to altered TGF-{beta} secretion. Collagen I, collagen III, and MMP-2 expression levels were lower on compliant PDMS substrates as compared to stiffer ones; there was little MMP-9 activity on substrates. These results demonstrate critical feedback mechanisms between substrate stiffness and ECM regulation by fibroblasts which is highly relevant in diseases like tissue fibrosis.

bioengineering↗

bioRxiv2020

Physical biology of cell-substrate interactions under cyclic stretch

Mechanosensitive focal adhesion complexes mediate the dynamic interactions between cells and substrates, and regulate cellular function. Integrins in adhesion complexes link substrate ligands to stress fibers in the cytoskeleton, and aid in load transfer and traction generation during cell adhesion and migration. A repertoire of signaling molecules, including calcium, facilitate this process. We develop a novel one-dimensional, multi-scale, stochastic finite element model of a fibroblast on a substrate which includes calcium signaling, stress fiber remodeling, and focal adhesion dynamics that describes the formation and clustering of integrins to substrate ligands. We link the stochastic dynamics involving motor-clutches at focal adhesions to continuum level stress fiber contractility at various locations along the cell length. The stochastic module links to a calcium signaling module, via IP3 generation, and adaptor protein dyanamics through feedback. We use the model to quantify changes in cellular responses with substrate stiffness, ligand density, and cyclic stretch. Results show that tractions and integrin recruitments vary along the cell length and depend critically on interactions between the stress fiber and reversibly engaging adaptor proteins. Maximum tractions and integrin recruitments were present at the lamellar regions. Cytosolic calcium increased with substrate stiffness and ligand density. The optimal substrate stiffness, based on maximum tractions exerted by the cell, shifted towards stiffer substrates at high ligand densities. Cyclic stretch increased the cytosolic calcium and tractions at lamellipodial and intermediate cell regions. Tractions and integrin recruitments showed biphasic responses with substrate stiffness that increased with ligand density under stretch. The optimal substrate stiffness under stretch shifted towards compliant substrates at a given ligand density. Cells deadhere under stretch, characterized by near-zero recruitments and tractions, beyond a critical substrate stiffness. The coupling of stress fiber contractility to adhesion dynamics is essential in determining cellular responses under external mechanical perturbations. Statement of SignificanceCells are exquisitely sensitive to substrate ligand density, stiffness, and cyclic stretch. How do cell-substrate interactions change under cyclic stretch? We use a systems biology approach to develop a one-dimensional, multi-scale, stochastic finite element model of cellular adhesions to substrates which includes focal adhesion attachment dynamics, stress fiber activation, and calcium signaling. We quantify tractions along the cell length in response to variations in substrate stiffness, cyclic stretching, and differential ligand densities. Calcium signaling changes the stress fiber contractility and focal adhesion dynamics under stretch and substrate stiffness. Cell tractions and adhesions show a biphasic response with substrate stiffness that increased with higher ligand density and cyclic stretch. Chemomechanical coupling is essential in quantifying mechanosensing responses underlying cell-substrate interactions.

biophysics↗

bioRxiv2019

Stress fiber growth and remodeling determines cellular morphomechanics under uniaxial cyclic stretch

Stress fibers in the cytoskeleton are essential in maintaining cellular shape, and influence their adhesion and migration. Cyclic uniaxial stretching results in cellular reorientation orthogonal to the applied stretch direction via a strain avoidance reaction; the mechanistic cues in cellular mechanosensitivity to this response are currently underexplored. We show stretch induced stress fiber lengthening, their realignment and increased cortical actin in fibroblasts stretched over varied amplitudes and durations. Higher amounts of actin and alignment of stress fibers were accompanied with an increase in the effective elastic modulus of cells. Microtubules did not contribute to the measured stiffness or reorientation response but were essential to the nuclear reorientation. We modeled stress fiber growth and reorientation dynamics using a nonlinear, orthotropic, fiber-reinforced continuum representation of the cell. The model predicts the observed fibroblast morphology and increased cellular stiffness under uniaxial cyclic stretch. These studies are important in exploring the differences underlying mechanotransduction and cellular contractility under stretch.

biophysics↗

bioRxiv2019

Mechanics of wood boring by beetle mandibles

Wood boring is a feature of several insect species and is a major cause of severe and irreparable damage to trees. Adult females typically deposit their eggs on the stem surface under bark scales. The emerging hatchlings live within hard wood during their larval phase, avoiding possible predation, whilst continually boring and tunneling through wood until pupation. A study of wood boring by insects offers unique insights into the bioengineering principles that drive evolutionary adaptations. We show that larval mandibles of the coffee wood stem borer beetle (Xylotrechus quadripes: Cerambycidae) have a highly sharp cusp edge to initiate fractures in Arabica wood and a suitable shape to generate small wood chips that are suitable for digestion. Cuticle hardness at the tip is significantly enhanced through zinc-enrichment. Finite element model of the mandible, based on the differential material properties at the tip, intermediate and base regions of the mandible, showed highest stresses in the tip region; these decreased to significantly lower values at the start of the hollow section. A hollow architecture significantly reduces bending stresses at mandibular base without compromising the structural integrity. A scaling model based on a fracture mechanics framework shows the importance of the mandible shape in generating optimal chip sizes. These findings contain general principles in tool design and put in focus interactions of insects and their woody hosts.

bioengineering↗