Biology subjects

Zhang, K. S.

Publications and source records attributed to Zhang, K. S..

4 recordsLinked to original sources

bioRxiv2025

Characterization of cellular wound resistance in the giant ciliate Stentor coeruleus

Resistance to mechanical stress is essential for cells to prevent wounding and maintain structural integrity. This capability is especially critical for free-living single-celled organisms, which routinely encounter mechanical stress from their natural habitats. We investigated Stentor coeruleus, a single-celled ciliate known for its remarkable wound repair capacity, as a model for studying mechanical wound resistance. While previous work focused on wound repair in Stentor, the structures that enable it to resist wounding remain poorly understood. We characterized how Stentor resisted mechanical stress during transit through a microfluidic constriction. Using high- speed imaging, we tracked the transit dynamics of the cells and linked them to wounding outcomes. Larger cells experienced longer transit times in the constriction and were more prone to rupture, often failing to recover shape due to membrane rupture and loss of cytoplasm. To elucidate the role of the Stentor cytoskeleton, we performed drug-mediated disruption of KM fibers, which are microtubule bundles in the Stentor cytoskeleton. Drug-treated cells exhibited an increased likelihood of membrane rupture at the constriction, implicating KM fibers in wound resistance. To investigate the resistance of Stentor cells to hydrodynamic stress, we injected the cells at increasing flow rates through the constriction. Interestingly, cells were more resistant to larger hydrodynamic stresses up to a threshold, potentially due to shear-thinning of the cytoplasm. Together, these results suggest that Stentor relies on both cytoskeletal architecture and cytoplasmic rheology to withstand mechanical stress, offering insights into cellular strategies for wound resistance in the absence of rigid extracellular structures.

cell biology↗

bioRxiv2024

SMORES: A Simple Microfluidic Operating Room for the Examination and Surgery of Stentor coeruleus

Ciliates are powerful unicellular model organisms that have been used to elucidate fundamental biological processes. However, the high motility of ciliates presents a major challenge in studies using live-cell microscopy and microsurgery. While various immobilization methods have been developed, they are physiologically disruptive to the cell and incompatible with microscopy and/or microsurgery. Here, we describe a Simple Microfluidic Operating Room for the Examination and Surgery of Stentor coeruleus (SMORES). SMORES uses Quake valve-based microfluidics to trap, compress, and perform surgery on Stentor as our model ciliate. Compared with previous methods, immobilization by physical compression in SMORES is more effective and uniform. The mean velocity of compressed cells is 24 times less than that of uncompressed cells. The compression is minimally disruptive to the cell and is easily applied or removed using a 3D-printed pressure rig. We demonstrate cell immobilization for up to 2 hours without sacrificing cell viability. SMORES is compatible with confocal microscopy and is capable of media exchange for pharmacokinetic studies. Finally, the modular design of SMORES allows laser ablation or mechanical dissection of a cell into many cell fragments at once. These capabilities are expected to enable biological studies previously impossible in ciliates and other motile species.

bioengineering↗

bioRxiv2022

Hydrodynamic dissection of single cells in a microfluidic system

Stentor coeruleus, a single-cell ciliated protozoan, is a model organism for wound healing and regeneration studies. Despite Stentors large size (up to 2 mm in extended state), microdissection of Stentor remains challenging. In this work, we describe a hydrodynamic cell splitter, consisting of a cross junction, capable of splitting Stentor cells in a non-contact manner at a high throughput of [~]500 cells/min under continuous operation. Introduction of asymmetry in the flow field at the cross junction leads to asymmetric splitting of the cells to generate cell fragments as small as [~]8.5 times the original cell size. Characterization of cell fragment viability shows reduced 5-day survival as fragment size decreases and as the extent of hydrodynamic stress imposed on the fragments increases. Our results suggest that cell fragment size and composition, as well as mechanical stress, play important roles in the long-term repair of Stentor cells and warrant further investigations. Nevertheless, the hydrodynamic splitter can be useful for studying phenomena immediately after cell splitting, such as the closure of wounds in the plasma membrane which occurs on the order of 100 - 1000 seconds in Stentor.

biophysics↗

bioRxiv2020

Microfluidic guillotine reveals multiple timescales and mechanical modes of wound response in Stentor coeruleus

BackgroundWound healing is one of the defining features of life and is seen not only in tissues but also within individual cells. Understanding wound response at the single-cell level is critical for determining fundamental cellular functions needed for cell repair and survival. This understanding could also enable the engineering of single-cell wound repair strategies in emerging synthetic cell research. One approach is to examine and adapt self-repair mechanisms from a living system that already demonstrates robust capacity to heal from large wounds. Towards this end, Stentor coeruleus, a single-celled free-living ciliate protozoan, is a unique model because of its robust wound healing capacity. This capacity allows one to perturb the wounding conditions and measure their effect on the repair process without immediately causing cell death, thereby providing a robust platform for probing the self-repair mechanism. ResultsHere we used a microfluidic guillotine and a fluorescence-based assay to probe the timescales and mechanisms of wound repair in Stentor. We found that Stentor requires [~]100 - 1000 s to close bisection wounds, depending on the severity of the wound. This corresponds to a healing rate of [~]8 - 80 m2/s, faster than most other single cells reported in the literature. Further, we observed and characterized three distinct mechanical modes of wound repair in Stentor: contraction, cytoplasm retrieval, and twisting/pulling. Using chemical perturbations, active cilia were found to be important for only the twisting/pulling mode. Contraction of myonemes, a major contractile fiber in Stentor, was surprisingly not important for the contraction mode and was of low importance for the others. ConclusionsWhile events local to the wound site have been the focus of many single-cell wound repair studies, our results suggest that large-scale mechanical behaviors may be of greater importance to single-cell wound repair than previously thought. The work here advances our understanding of the wound response in Stentor, and will lay the foundation for further investigations into the underlying components and molecular mechanisms involved.

cell biology↗