Biology subjects

Wolfenson, H.

Publications and source records attributed to Wolfenson, H..

16 recordsLinked to original sources

bioRxiv2026

An AI-assisted platform for quantitative histopathological analysis in interstitial lung disease

Interstitial lung diseases (ILDs) are heterogeneous pulmonary disorders characterized by chronic inflammation and/or fibrosis. 30-40% of ILD patients develop fibrotic disease that is associated with progressive respiratory decline and poor prognosis, particularly in idiopathic pulmonary fibrosis. Current antifibrotic therapies slow disease progression but do not reverse fibrosis, highlighting the need for improved therapeutic strategies. Robust histopathological evaluation in preclinical models is essential for drug development; however, conventional scoring systems are semi-quantitative, labor-intensive, subject to inter-observer variability, and rely on limited field sampling. Here, we introduce FibroSight, a standalone platform for compartment-resolved quantification of lung remodeling in Sirius Red-stained sections. By integrating deep learning- based structural segmentation with color-based feature extraction, FibroSight enables highly automated whole-lobe analysis without requiring complex computational setup. The platform quantifies complementary remodeling parameters, including parenchymal collagen fraction, parenchymal tissue density, nuclear area fraction, parenchymal airspace fraction, and airway- and vascular-associated remodeling. Validated in the bleomycin-induced fibrosis model, FibroSight-derived metrics strongly correlated with expert Ashcroft scoring and showed stronger associations with histological severity than corresponding outputs from a semi-automated ImageJ-based workflow. The platform further distinguished inflammatory from fibrotic remodeling in influenza-induced lung injury and demonstrated translational proof-of-concept applicability in human ILD biopsy specimens. By enabling scalable, reproducible, and multi-compartment histological quantification, FibroSight provides a practical framework for objective assessment of lung remodeling. This approach expands conventional fibrosis evaluation by integrating fibrotic, inflammatory, airway, and vascular-associated readouts, supporting more precise analysis of disease mechanisms and therapeutic responses in preclinical and translational ILD research.

pathology↗

bioRxiv2026

Hypertension-mediated cardiac fibrosis is a mechanical process initiated by smooth muscle cells

Hypertension represents the most prevalent chronic cardiovascular condition, typically culminating in pathological cardiac remodeling characterized by hypertrophy and extensive fibrosis. Although the cellular phenotypes associated with these changes are well-documented, the precise mechanisms by which hypertensive stress is sensed and transduced into a fibrotic program remain poorly defined. To elucidate these mechanisms, we investigated the role of Lysyl oxidase (LOX), an extracellular matrix (ECM)-modifying enzyme that is upregulated during hypertensive stress and associated with cardiovascular diseases. By employing cell-type specific Cre-Lox technology to conditionally delete Lysyl oxidase in either smooth muscle cells (SMCs) or fibroblasts, the primary ECM-secreting cell populations, we demonstrate that fibroblast-specific Lox deletion had no significant impact on the progression of cardiac fibrosis. Conversely, SMC-specific Lox deletion selectively inhibited the fibrotic response without affecting other remodeling parameters, such as cardiac hypertrophy. Notably, in the SMC-specific Lox knockout hearts, fibrosis was restricted to the perivascular niche and failed to propagate into the cardiac interstitium. We find that this transition is a mechanical, ECM-dependent process initiated by SMCs. Our results identify SMCs, rather than fibroblasts, as the primary sensors and initiators of the hypertensive fibrotic response. These findings demonstrate that fibrosis can be uncoupled from other hypertensive manifestations and identify SMC-mediated ECM modification as a potential therapeutic target for treating hypertensive heart disease.

cell biology↗

bioRxiv2026

Analysis of the assembly, stabilization and maturation of the multiphasic TAZ biomolecular condensates

Phase separation is an important mechanism ensuring efficient regulation and function in Hippo signaling. Particularly, phase separation of nuclear TAZ has been demonstrated to be essential for its activity. However, the mechanisms of TAZ condensate assembly and maturation are yet undefined. Here we explored these mechanisms using FRAP with two laser beam sizes complemented by microscopy and cell biology approaches. We show that TAZ condensates are multiphasic, with a more stable core and labile periphery. TAZ initially forms small nascent clusters, likely via self-nucleation through the CC domain. These gradually mature into larger condensates through interaction with additional proteins via the WW domain. The condensates are further stabilized/activated by interaction with transcription factors and complexes including TEAD4 and P-TEFb. Of note, the ability of TAZ to form mature condensates is essential for its activities in cellular morphogenesis and tumorigenesis. Our study presents detailed mechanistic analysis of TAZ phase separation, revealing a highly dynamic nature of TAZ condensate maturation and activation. TeaserTAZ condensates grow from nascent clusters into mature condensates by interactions with transcription factors and complexes.

cell biology↗

bioRxiv2025

Synthetic Mimetics of Exosomal Lipid Profile Enhance Gemcitabine Delivery in Pancreatic Cancer

Pancreatic cancer ranks fourth among cancer-related deaths. Despite decades of research, the cure rate of disease remains disappointingly low. This dismal prognosis is due to late detection and to resistance of tumors to all known systemic therapies. Here, we build upon our previous findings that highlighted the selective uptake of tumor-associated macrophage-derived exosomes by cancer cells. We hypothesize that the prominent lipid contents on the exosome surface play a crucial role in facilitating selective cellular interactions, offering a novel avenue for developing efficient drug delivery platforms for pancreatic cancer. Lipids affinity toward cancer cells was measured, following mass spectral lipidomic analysis for screening the lipidic composition of macrophages and their derived exosomes. Subsequently, optimized and characterized synthetic exosomes underwent detailed intracellular trafficking studies in human pancreatic cancer cells, demonstrating enhanced cellular uptake kinetics. Notably, gemcitabine-loaded synthetic exosomes exhibited superior efficacy in inducing programmed cell death compared to both the free drug and conventional liposome formulations. The biodistribution examination of these synthetic exosomes underscored their potential for tumor specificity. In vivo experiments further demonstrated that the treatment with gemcitabine-incorporated synthetic exosomes inhibited tumor growth by nearly 50% compared to the administration of the free drug, indicating a significantly enhanced treatment efficacy. Our findings indicate that leveraging the lipid membrane composition of exosomes could lead to breakthroughs in drug delivery efficiency. This innovative strategy offers a promising direction in the ongoing battle against pancreatic cancer, highlighting the potential of exosomal lipid profiles for cancer therapy.

cell biology↗

bioRxiv2025

The gut microbiota-derived metabolite indole regulates cytoskeletal functions and virulence in Entamoeba histolytica

Entamoeba histolytica is a pathogenic amoeba that inhabits the human large intestine and causes amoebiasis. E. histolytica interacts with both the intestinal microbiota and the metabolites they produce. These bacterial metabolites play a crucial role in shaping the virulence and stress resistance of E. histolytica. One such metabolite, indole, is synthesized by bacteria from tryptophan and functions as a key signaling molecule. In this study, we investigated the impact of indole on E. histolytica by incubating trophozoites with the metabolite and performing proteomic analyses under various conditions, including trophozoites adapted to indole. Our results show that indole is toxic to E. histolytica, with an inhibitory concentration (IC) of 1.2 mM; however, the parasite can adapt to this concentration. Proteomic analyses reveal that indole-adapted trophozoites display enhanced resistance to oxidative stress (OS), upregulation of cytoskeletal proteins, and increased virulence. These trophozoites exhibit increased F-actin formation, smaller cell size, stronger adhesion to HeLa cells, enhanced migratory capacity, and more effective colonization of the mouse cecum compared to non-adapted trophozoites. Thus, indole exerts a dual effect on E. histolytica physiology. While indole is initially toxic to the parasite, adaptation to indole confers enhanced resistance to OS and promotes a more virulent phenotype. This duality underscores the complex role of microbiota-derived metabolites in modulating parasite behavior and highlights indole as a key microbial signal. Author summaryEntamoeba histolytica is a unicellular parasite living in the human large intestine and can cause a disease called amoebiasis. Inside the intestine, E. histolytica interacts with the microbiota and with the metabolites they produce. One such metabolite is indole, which is produced by gut bacteria from tryptophan and used by human cells and trophozoites. In this study, we explored how indole affects E. histolytica. We found that indole is toxic to the parasite at first, but over time, the amoeba can adapt and survive in its presence. Using protein analysis techniques, we discovered that indole-adapted amoebas become more resistant to stress and more virulent. They produce more F-actin, leading to improved mobility, degrade more HeLa cells, and colonize the gut of mice more efficiently than wild-type amoebas (WT). These results show that, while indole initially harms the parasite, it also triggers changes that make it more resistant to stress and more virulent. Our work highlights how metabolites produced by bacteria in the large intestine can influence the behavior of E. histolytica.

microbiology↗

bioRxiv2025

Hydrogel viscoelasticity modulates cell nascent extracellular matrix deposition

Polymeric hydrogels are valuable platforms for determining how specific mechanical properties of native tissue extracellular matrix (ECM) regulate cell function. Recent research has focused on incorporating viscous and elastic properties into hydrogels to investigate cellular responses to time-dependent mechanical properties of the ECM. However, a critical aspect often overlooked is that cells continuously remodel their microenvironment in hydrogels, such as by the deposition of newly secreted (nascent) ECM. While this nascent ECM has been demonstrated to play a vital role in transmitting mechanical signals across various biological contexts, the mechanisms by which it regulates cellular function in response to time-dependent mechanical properties remain poorly understood. In this study, we developed an interpenetrating polymer network that enables independent control of viscous and elastic hydrogel properties. We show that cells cultured on high-viscosity hydrogels deposit increased nascent ECM which also correlates with enhanced hydrogel remodeling. Interestingly, higher nascent ECM deposition on high-viscosity hydrogels was decoupled from intracellular contractility. These results establish a relationship between hydrogel viscosity and nascent ECM deposition that may extend to diverse cell types and offer new insights into cell-hydrogel interactions.

bioengineering↗

bioRxiv2024

Enteropathogenic Escherichia coli induces Entamoeba histolytica Levy-like movement on fibronectin-rich substrate by reducing traction forces

Amebiasis, caused by Entamoeba histolytica, is a global health concern, affecting millions and causing significant mortality, particularly in areas with poor sanitation. Although recent studies have examined E. histolyticas interaction with human intestinal microbes, the impact of bacterial presence on the parasites motility, mechanical forces, and their potential role in altering invasiveness have not been fully elucidated. In this study, we utilized a micropillar-array system combined with live imaging to investigate the effects of enteropathogenic Escherichia coli on E. histolyticas motility characteristics, F-actin spatial localization, and traction force exerted on fibronectin-coated substrates. Our findings indicate that co-incubation with E. coli significantly enhances the motility of E. histolytica, as evidenced by the enhancement of Levy-like movement patterns, i.e., increased directionality and velocity. This increased motility is accompanied by a reduction in F-actin-dependent traction forces and podosome-like structures on fibronectin-coated substrates, but with increased F-actin localization in the upper part of the cytoplasm. These findings highlight the role of physical interactions and cellular behaviors in modulating the parasites virulence, providing new insights into the mechanistic basis of its pathogenicity. Author SummaryAmebiasis, caused by the protozoan parasite Entamoeba histolytica, is a major global health issue, affecting around 50 million people and resulting in 100,000 deaths annually. The disease is transmitted through contaminated food and water. Upon reaching the intestines, which are teeming with bacteria, E. histolytica begins its invasion by removing the protective mucus layer, followed by adhering to and detaching enterocytes, leading to the disruption and degradation of the epithelial barrier. Afterwards, E. histolytica invades along the fibronectin-rich basement membrane deep into the crypts of Lieberkuhn, eventually penetrating the fibronectin- rich lamina propria. This leads to further tissue destruction and potential dissemination to distant organs, causing severe complications. In our study, we explored how the presence of enteropathogenic bacteria affects the parasites motility and mechanical force generation, both of which are key to its pathogenicity. Using a micropillar-array system and live imaging, we found that exposure to enteropathogenic Escherichia coli significantly increases E. histolyticas motility while reducing its traction force on fibronectin-rich matrices. These changes in behavior highlight the role of bacterial interactions in enhancing the parasites virulence. Our findings provide important insights into the mechanistic basis of E. histolyticas pathogenicity, offering potential avenues for new treatments against amebiasis.

microbiology↗

bioRxiv2024

Spatiotemporal analysis of F-actin polymerization with micropillar arrays reveals synchronization between adhesion sites

We repurposed micropillar-arrays to quantify spatiotemporal inter-adhesion communication. Following the observation that integrin adhesions formed around pillar tops we relied on the precise repetitive spatial control of the pillars to reliably monitor F-actin dynamics in mouse embryonic fibroblasts as a model for spatiotemporal adhesion-related intracellular signaling. Using correlation-based analyses we revealed localized information-flows propagating between adjacent pillars that were integrated over space and time to synchronize the adhesion dynamics within the entire cell. Probing the mechanical regulation, we discovered that stiffer pillars or partial actomyosin contractility inhibition enhances inter-adhesion F-actin synchronization. Our results suggest that adhesions can communicate and highlight the potential of using micropillar arrays as a tool to measure spatiotemporal intracellular signaling.

cell biology↗

bioRxiv2022

Initiation of fibronectin fibrillogenesis is an enzyme-dependent process

Fibronectin fibrillogenesis and mechanosensing both depend on integrin-mediated force transmission to the extracellular-matrix. However, force transmission is in itself dependent on fibrillogenesis, and fibronectin fibrils are found in soft embryos where high forces cannot be applied, suggesting that force cannot be the sole initiator of fibrillogenesis. Here we identify a nucleation step prior to force transmission, driven by fibronectin oxidation mediated by lysyl-oxidase enzyme family members. This oxidation induces fibronectin clustering that promotes early adhesion, alters cellular response to soft matrices, and enhances force transmission to the matrix. In contrast, absence of fibronectin oxidation abrogates fibrillogenesis, perturbs cell-matrix adhesion, and compromises mechanosensation. Moreover, fibronectin oxidation promotes cancer cells colony formation in soft agar as well as collective and single-cell migration. These results reveal a force-independent enzyme-dependent mechanism that initiates fibronectin fibrillogenesis, establishing a critical step in cell adhesion and mechanosensing.

cell biology↗

bioRxiv2022

Characteristic energy scales of active fluctuations in adherent cells

Cell-matrix and cell-cell adhesion play important roles in a wide variety of physiological processes, from the single cell level to the large scale, multicellular organization of tissues. Cells actively apply forces to their environment, either extracellular matrix or neighboring cells, as well as sense its biophysical properties. The fluctuations associated with these active processes occur on an energy scale much larger than that of ordinary thermal equilibrium fluctuations, yet their statistical properties and characteristic scales are not fully understood. Here, we compare measurements of the energy scale of active cellular fluctuations -- an effective cellular temperature -- in four different biophysical settings, involving both single cell and cell aggregates experiments under various control conditions, different cell types and various biophysical observables. The results indicate that a similar energy scale of active fluctuations might characterize the same cell type in different settings, though it may vary among different cell types, being approximately 6 to 8 order of magnitude larger than the ordinary thermal energy at room temperature. These findings call for extracting the energy scale of active fluctuations over a broader range of cell types, experimental settings and biophysical observables, and for understanding the biophysical origin and significance of such cellular energy scales.

biophysics↗

bioRxiv2022

Perturbed actin cap and nuclear morphology in primary fibroblasts of Huntington's disease patients as a new phenotypic marker for personalized drug evaluation

Human primary skin fibroblast cells from patients skin biopsies were used previously as a model to study different neurodegenerative diseases, including Huntingtons Disease (HD). These cells are directly isolated from the patients tissue without any alteration in the genome, retaining in culture conditions their endogenous cellular characteristics and biochemical properties, as well as their cellular proliferation capacity for several passages. The aim of this study was to identify a distinctive cellular phenotype in primary skin fibroblasts from various HD patients, using image-based high content analysis, which could be used in the future for personalized drug screening treatment evaluation. We show that HD fibroblasts have a distinctive nuclear morphology associated with a nuclear actin cap deficiency, which in turn affects cell motility in a similar manner to primary skin fibroblasts from Hutchinson-Gilford progeria syndrome (HGPS) patients used as known actin cap deficient cells. Moreover, treatment of the HD cells with either Latrunculin B, used to disrupt actin cap formation, or the antioxidant agent Mitoquinone, used to improve mitochondrial activity, show opposite effects on actin cap associated morphological features and cell motility. The former exacerbates the HD phenotype while the latter improves it. Deep data analysis of the HD nuclear and actin cap features using custom developed image analysis algorithms allow strong cluster classification distinct from HGPS and healthy matching controls, supporting the finding of a novel HD cellular phenotypic marker that could be modulated by pharmacological agents in this patient-based disease model.

cell biology↗

bioRxiv2021

Soft limbal niche maintains stem cell compartmentalization and function through YAP

Stem cells (SCs) decision to self-renew or differentiate largely depends on the external control of their niche. However, the complex mechanisms that underlie this crosstalk are poorly understood. To address this question, we focused on the corneal epithelial SC model in which the SC niche, known as the limbus, is spatially segregated from the differentiation compartment. We report that the unique biomechanical property of the limbus supports the nuclear localization and function of Yes-associated protein (YAP), a putative mediator of the mechanotransduction pathway. Perturbation of tissue stiffness or YAP activity affects SC function as well as tissue integrity under homeostasis and significantly inhibited the regeneration of the SC population following SC depletion. In vitro experiments revealed that substrates with the rigidity of the corneal differentiation compartment inhibit YAP localization and induce differentiation, a mechanism that is mediated by the TGF{beta}-SMAD2/3 pathway. Taken together, these results indicate that SC sense biomechanical niche signals and that manipulation of mechano-sensory machinery or its downstream biochemical output may bear fruits in SC expansion for regenerative therapy. HighlightsO_LIYAP is essential for limbal SC function, regeneration, and dedifferentiation C_LIO_LILox over-expression stiffens the limbal niche, affects SC phenotype and corneal integrity C_LIO_LICorneal rigidity represses YAP and stemness in a SMAD2/3-dependent manner C_LIO_LIManipulation of mechanosensory or TGF-{beta} pathway influences limbal SC expansion in vitro C_LI

cell biology↗

bioRxiv2021

Alpha-catenin regulates mechanosensing and rigidity-dependent growth by linking integrin adhesions to F-actin

Both cell-cell and cell-matrix adhesions are regulated by mechanical signals, but the mechanobiological processes that mediate the crosstalk between these structures are poorly understood. Here we show that - catenin, a mechanosensitive protein that is classically associated with cadherin-based adhesions, directly interacts with and regulates integrin adhesions. -catenin is recruited to the edges of mesenchymal cells, where it interacts with F-actin. This is followed by mutual retrograde flow of -catenin and F-actin from the cell edge, during which -catenin interacts with vinculin within integrin adhesions. This interaction affects adhesion maturation, stress-fiber assembly, and force transmission to the matrix. In epithelial cells, -catenin is present in cell-cell adhesions and absent from cell-matrix adhesions. However, when these cells undergo epithelial-to-mesenchymal transition, -catenin transitions to cell-matrix adhesions, where it facilitates proper mechanosensing. This is highlighted by the ability of -catenin-depleted cells to grow on soft matrices. These results suggest a dual role of -catenin in mechanosensing, through both cell-cell and cell-matrix adhesions.

cell biology↗

bioRxiv2019

Cellular contractile forces emerge from time and F-actin dependent non-mechanosensitive displacements

Cells ability to apply contractile forces to their environment and to sense its mechanical properties (e.g. rigidity) are among their most fundamental features. Yet, the interrelations between contractility and mechanosensing, in particular whether contractile force generation depends on mechanosensing, are not understood. We use theory and extensive experiments to study the time evolution of cellular contractile forces and show that they are generated by time-dependent actomyosin contractile displacements that are independent of the environments rigidity. Consequently, contractile forces are non-mechanosensitive. We further show that the force-generating displacements are directly related to the evolution of the actomyosin network, most notably to the time-dependent concentration of F-actin. The emerging picture of force generation and mechanosensitivity offers a unified framework for understanding contractility.\n\nOne Sentence SummaryCellular contractile forces are generated by rigidity-independent displacements that are determined by the time evolution of F-actin assembly.

cell biology↗

bioRxiv2018

Tumor suppressor DAPK1 catalyzes adhesion assembly on rigid but anoikis on soft matrices

Cancer cells will normally grow on soft surfaces, but if rigidity sensing modules are restored in cancer cells, they will undergo apoptosis on soft surfaces (anoikis) like most normal cells. DAPK1 is a major tumor suppressor that activates cell death, but it is unclear how DAPK1 could activate anoikis through rigidity sensing. Here we find that when rigidity sensing is decreased through inhibition of DAPK1 activity, cells are transformed for growth on soft matrices. Further, DAPK1 catalyzes matrix adhesion assembly and is part of adhesions on rigid surfaces. Additional factors include DAPK1 phosphorylation of tropomyosin2.1, talin1 head domain and tyrosine phosphorylation of DAPK1 by Src. On soft surfaces, DAPK1 rapidly dissociates from the adhesion complexes and activates apoptosis that requires PTPN12 activity and talin1 head. Thus, DAPK1 is important for adhesion assembly on rigid surfaces and the activation of anoikis on soft surfaces through its binding to rigidity-sensing modules.

cell biology↗

bioRxiv2017

Stopping Transformed Growth with Cytoskeletal Proteins: Turning a Devil into an Angel

The major hallmark of cancer cells is uncontrollable growth on soft matrices (transformed growth), which indicates that they have lost the ability to properly sense the rigidity of their surroundings. Recent studies of fibroblasts show that local contractions by cytoskeletal rigidity sensor units block growth on soft surfaces and their depletion causes transformed growth. The contractile system involves many cytoskeletal proteins that must be correctly assembled for proper rigidity sensing. We tested the hypothesis that cancer cells lack rigidity sensing due to their inability to assemble contractile units because of altered cytoskeletal protein levels. In four widely different cancers, there were over ten-fold fewer rigidity-sensing contractions compared with normal fibroblasts. Restoring normal levels of cytoskeletal proteins restored rigidity sensing and rigidity-dependent growth in transformed cells. Most commonly, this involved restoring balanced levels of the tropomyosins 2.1 (often depleted by miR-21) and 3 (often overexpressed). Restored cells could be transformed again by depleting other cytoskeletal proteins including myosin IIA. Thus, the depletion of rigidity sensing modules enables growth on soft surfaces and many different perturbations of cytoskeletal proteins can disrupt rigidity sensing thereby causing transformed growth of cancer cells.

cell biology↗