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#cell

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📰 "Beating the gold standard: A review of Mycobacterium tuberculosis lysis using bead beating and the need for standardization."
biorxiv.org/content/10.1101/20 #Mechanical #Cell

bioRxiv · Beating the gold standard: A review of Mycobacterium tuberculosis lysis using bead beating and the need for standardization.Bead beating is widely used for mechanical lysis of Mycobacterium tuberculosis, a bacterium with a highly resistant, lipid-rich cell wall. Despite its status as a de facto gold standard for mycobacterial lysis, there is no standardized protocol for bead beating, resulting in significant variability across studies. We conducted a literature review of 73 studies, identifying 38 with explicit mycobacterial bead beating protocols. Our analysis revealed heterogeneity in bead types, sizes, device models, and operational parameters, with 37% of studies failing to report critical details such as lysis speed. We experimentally assessed the impact of key variables - tube type, bead quantity, and device settings - on lysis efficiency using qPCR of M. tuberculosis DNA. Results showed that even minor changes, such as tube shape or bead volume, can significantly affect DNA yield. These findings underscore the need for standardized bead-beating protocols to improve reproducibility and comparability. Future efforts should prioritize developing consensus methods tailored to sample type and analytical application. ### Competing Interest Statement The authors have declared no competing interest.

📰 "Mechanical Coordination of Intestinal Cell Extrusion by Supracellular 3D Force Patterns"
biorxiv.org/content/10.1101/20 #Mechanical #Forces #Cell

bioRxiv · Mechanical Coordination of Intestinal Cell Extrusion by Supracellular 3D Force PatternsEvery day, the mammalian intestinal epithelium extrudes millions of cells to sustain tissue self-renewal. Despite its fundamental role in intestinal homeostasis, the mechanisms that trigger, compartmentalize, and execute intestinal cell extrusion remain largely unknown. Here, using intestinal organoids, we map the three-dimensional forces and cytoskeletal dynamics that drive intestinal cell extrusion. We show that, unlike in other epithelia, extrusion is initiated by the sudden dissolution of a contractile myosin 2A meshwork triggered by a calcium influx. Following meshwork dissolution, the extruding cell and its neighbors generate an upwards traction force that requires myosin contractility but is generated by lamellipodial protrusions in neighboring cells. Importantly, these lamellipodia not only act as force generators but also determine whether extrusion occurs apically or basally, serving as symmetry breakers of the process. Finally, we show that compartmentalization of cell extrusion to the outside of the intestinal crypt does not require curvature and instead depends on myosin 2A. Our findings reveal that the intestinal epithelium exhibits a distinctive mode of extrusion, in which tension differentials - rather than compressive stresses from crowding - trigger and compartmentalize cell removal. ### Competing Interest Statement The authors have declared no competing interest. Spanish Ministry for Science and Innovation, FJC2018-037440-I, PID2022-142672NB-I00, PID2021-128635NB-I00 MCIN/AEI/ 10.13039/501100011033, ERDF-EU A way of making Europe EMBO, ALTF-1169 European Research Council, Adv-101097753, Adv-883739 Generalitat de Catalunya, 2017-SGR-1602, AGAUR SGR-2017-01602 ICREA Academia Fundació la Marató de TV3, 201903-30-31-32 European Commission, H2020-FETPROACT-01-2016-731957 La Caixa Foundation, LCF/PR/HR24/00326 Human Frontiers Science Program, HFSPRGP022/2024

📰 "From Development to Regeneration: Insights into Flight Muscle Adaptations from Bat Muscle Cell Lines"
biorxiv.org/content/10.1101/20 #Mechanical #Cell

bioRxiv · From Development to Regeneration: Insights into Flight Muscle Adaptations from Bat Muscle Cell LinesSkeletal muscle regeneration depends on muscle stem cells, which give rise to myoblasts that drive muscle growth, repair, and maintenance. In bats, the only mammals capable of powered flight, these processes must also sustain contractile performance under extreme mechanical and metabolic stress. However, the cellular and molecular mechanisms underlying bat muscle physiology remain largely unknown. To enable mechanistic investigation of these traits (Graphical Abstract), we established the first myoblast cell lines from the pectoralis muscle of Pteronotus mesoamericanus , a highly maneuverable aerial insectivore. Using both spontaneous immortalization and exogenous hTERT/CDK4 overexpression, we generated two stable cell lines that retain proliferative capacity and differentiate into contractile myotubes. These cells exhibit frequent spontaneous contractions, suggesting robust functional integrity at the neuromuscular junction. In parallel, we performed transcriptomic and metabolic profiling of native pectoralis tissue to define molecular programs supporting muscle specialization. Gene expression analyses revealed enriched pathways for muscle metabolism, development, and regeneration, highlighting the supporting roles in tissue maintenance and repair. Consistent with this profile, the flight muscle is triglyceride-rich, which serves as an important fuel source for energetically demanding processes, including muscle contraction and cellular recovery. Integration of transcriptomic and metabolic data identified three key metabolic modules—glucose utilization, lipid handling, and nutrient signaling—that likely coordinate ATP production and support metabolic flexibility. Together, these complementary tools and datasets provide the first in vitro platform for investigating bat muscle research, enabling direct exploration of muscle regeneration, metabolic resilience, and evolutionary physiology. ### Competing Interest Statement The authors have declared no competing interest. National Science Foundation, https://ror.org/021nxhr62, 2109717 National Institutes of Health, 1DP2AG071466 Burroughs Wellcome Fund, 1022339 Howard Hughes Medical Institute, 1599

📰 "Multiparametric Correlative Topographical and Volumetric Fluorescence Microscopy"
biorxiv.org/content/10.1101/20 #Mechanical #Cell

bioRxiv · Multiparametric Correlative Topographical and Volumetric Fluorescence MicroscopyLive-cell imaging of cell surface topography and intracellular architecture is essential for understanding cellular function. However, conventional approaches often involve trade-offs between resolution, invasiveness, and volumetric coverage. Here, we present an integrated Scanning Ion Conductance Microscope and single-objective Oblique Plane Microscope (SICM-OPM) system that enables simultaneous non-contact topographical imaging and volumetric fluorescence imaging within the same live cell. Beyond correlative live imaging, the platform supports nanomechanical mapping with tens-of-nanometres resolution, fluorescence-guided localised molecular delivery via the SICM, and benefits from reduced photobleaching due to light-sheet excitation. We demonstrate this platform's capabilities by visualising imipramine-induced T-tubule remodelling in live cardiomyocytes, revealing subsurface detubulation while surface morphology remains preserved. Additionally, we show precision delivery of fluorescent cargos—including dextrans and α-synuclein—into diatom and mammalian cells, alongside localised stiffness mapping to evaluate mechanical responses. We believe this technique opens new avenues for correlative structural, functional, and biophysical studies in live cells, with broad relevance to cell biology, neurodegeneration, and mechanobiology. ### Competing Interest Statement AS is a shareholder in ICAPPIC, Ltd., a company commercialising nanopipette-based instrumentation. EPSRC, EP/W012219/1, EP/W015005/1, EP/X034968/1 Imperial College London and the China Scholarship Council (CSC) Scholarship, File No. 202309370006 Early Career Pathway Award of the CRUK International Alliance for Cancer Early Detection, EDDAPA-2024/100006 British Heart Foundation, RG/F/22/110081 Alzheimer's Society, Dementia Research Leader Fellowship AS-DRL-24-012 UKRI Future Leaders Fellowship, MR/S033947/1, MR/Y003616/1 Alzheimer's Research UK, Major Project Grant ARUK-PG2019B-020 UK Natural Environment Research Council, NE/V01451X/2

📰 "Microtubule curling as an efficient readout to uncover fundamental concepts of axonal cell biology"
biorxiv.org/content/10.1101/20 #Microtubule #Mechanical #Cell

bioRxiv · Microtubule curling as an efficient readout to uncover fundamental concepts of axonal cell biologyIn neurodegeneration, axons tend to be prime lesion sites. Axons are the slender, up-to-meter-long processes of nerve cells that essentially wire nervous systems. These delicate structures must survive for an organism's lifetime. Their long-term maintenance requires complex cell biology to be locally present in axons, and this depends on the steady supply with organelles and materials. These are delivered by motor protein-driven axonal transport, which uses continuous bundles of MTs as highways that run interrupted from the cell body to the axon tip. By studying the regulation of these bundles in our past work, we identified many conditions in which microtubules become severely disorganised, a phenotype expected to be detrimental for axons. Further work on these conditions led us to propose the Dependency Cycle of Local Axon Homeostasis model explaining how aberration of different cell biological processes can lead to one common outcome - as is the case in neurodegeneration. Here we put this model to the test. Through live imaging we trace the origins and time course of microtubule disorganisation in normal and mutant neurons, and our findings confirm key mechanisms proposed in our model. Combining published data with the re-analysis and new investigation of 105 gene deficiencies from a wide range of cell biological contexts, we find that ~40% cause the shared microtubule disorganisation phenotype, thus clearly corroborating a key idea of the cycle. Through probing the phenotypes of 18 genes for ROS involvement, we find them to correctly classify into a mechanical ROS-independent versus a physiological ROS-dependent group, providing strong support for a central aspect of the dependency model. ### Competing Interest Statement The authors have declared no competing interest. Biotechnology and Biological Sciences Research Council, https://ror.org/00cwqg982, BB/P020151/1, BB/L000717/1, BB/M007553/1, BB/I002448/1, BB/C515998/1