Biology subjects

Nishikawa, M.

Publications and source records attributed to Nishikawa, M..

9 recordsLinked to original sources

bioRxiv2026

Super-resolution imaging flow cytometry

Organelle architecture is a key determinant of cellular state. However, its heterogeneity remains difficult to quantify at population scale because conventional imaging flow cytometry is diffraction-limited. Here we present super-resolution imaging flow cytometry (SR-IFC), a high-throughput structured-illumination platform combining stroboscopic excitation, microfluidic hydrodynamic focusing and AI-assisted reconstruction. From three phase-shifted frames acquired along the flow axis, SR-IFC reconstructs images with approximately 118-nm isotropic in-plane resolution, about two-fold beyond the diffraction limit, at up to approximately 720 cells per second. In microbiology, SR-IFC resolves individual lipid droplets in nitrogen-starved Euglena gracilis and reveals spatial reorganization during lipid biogenesis missed by diffraction-limited imaging. In immunology, it maps activation-associated mitochondrial remodeling across primary human CD8+ T-cell differentiation, including a transient networked state. SR-IFC thus provides a broadly applicable platform for population-scale super-resolution phenotyping across diverse areas of biology.

bioengineering↗

bioRxiv2026

Computational analysis reveals the requirement for cell-cell interaction in liver repopulation by transplanted hepatocytes

Hepatocyte transplantation is a promising alternative to liver transplantation; however, it currently serves only as a temporary treatment until a donor organ becomes available. In contrast, animal studies have demonstrated "liver repopulation", a phenomenon in which transplanted hepatocytes progressively replace host hepatocytes. Despite extensive documentation, the mechanisms driving this process remain poorly understood. More fundamentally, it remains unclear whether liver repopulation is driven by active cell-cell interactions between host and transplanted hepatocytes that induce host cell death, or whether it can be explained solely by intrinsic differences in proliferation and survival between these populations. To address this, we performed computational simulations using an agent-based model constrained by experimental data from repopulation in uninjured rat livers. Our analysis reveals that host hepatocyte death rate is the dominant determinant of repopulation kinetics, whereas variations in proliferation rate have only a limited impact. Notably, experimentally observed repopulation dynamics were only reproduced when cell- cell interactions were incorporated, or alternatively when host hepatocyte lifespan was set to unrealistically short values (approximately 25 days). These findings support a model in which cell- cell interactions play a critical role in efficient liver repopulation. More broadly, this study establishes a conceptual and computational framework for evaluating the requirement for cell-cell interactions in tissue replacement, even in the absence of a defined molecular mechanism.

systems biology↗

bioRxiv2026

Mechanochemical coupling tunes robustness of PAR polarity across developmental contexts in the C. elegans embryo

Proper establishment of cell polarity is essential for reliable asymmetric divisions during embryonic development. In the Caenorhabditis elegans zygote, polarity is established through mechanochemical coupling between advective transport driven by cortical flows and mutually antagonistic PAR protein interactions. How these mechanisms are integrated across developmental contexts to establish cell polarity in the early embryo remains unexplored. Here we show that P1 polarization relies primarily on sustained antagonistic activity of anterior PAR proteins mediated by CDC-42 and PKC-3. Cortical contractility is regulated by CDC-42 through MRCK-1 to drive cortical flow, and is largely independent of the RHO-1 - LET-502 pathway that is crucial in zygotic polarity establishment. We find that these flows contribute only weakly to P1 polarity and primarily during the late phase. P1 polarization is sensitive to CDC-42 dosage, as indicated by reduced cytoplasmic asymmetry of MEX-5 and loss of division asynchrony between daughter cells. In contrast, zygotic polarization remains robust to comparable perturbations in CDC-42 expression when cortical flow is intact, but becomes sensitized when cortical flow is suppressed. Together, these findings show that cortical flow acts as a transport-mediated reinforcement that buffers PAR polarity against perturbations in CDC-42 expression, and that weakening this reinforcement sensitizes polarity establishment across developmental contexts.

biophysics↗

bioRxiv2025

Active geometrodynamics predicts the emergence of cytokinesis

Cell division accomplishes the segregation of genetic material and involves remarkable changes in the cellular geometry culminating in cytokinesis: the cleavage of a mother cell giving rise to two daughter cells. Cytokinesis in animal cells is driven by flows resulting from cortical tension gradients in the actomyosin cortex. Here, we combine a theory for the active geometrodynamics of the cortical surface and quantitative measurements in the C. elegans zygote to reveal the physical principles of cytokinesis. At high activity, we observe the spontaneous emergence of ring-like patterns of myosin concentration and cell shape in the theory. The constriction dynamics of this self-organized pattern agrees with the ingression of the cytokinetic furrow and concomitant myosin accumulation during the first division in the C. elegans embryo. Through RNAi perturbations, we quantitatively test our theoretical predictions of myosin accumulation rates linearly varying with the ingression rate, and the emergence of asymmetric ingression. Our work suggests that the self-organised geometrodynamics of active fluid surfaces underlies cytokinesis.

biophysics↗

bioRxiv2025

A novel sorting method reveals superior metabolic activity of mononucleated over binucleated tetraploid hepatocytes

Hepatocytes display notable ploidy diversity, varying both in the number of genomic copies and in the number of nuclei. Most mouse hepatocytes are tetraploid (4n), which can exist as either mononucleated (1x4n) or binucleated (2x2n) cells. Despite this distinction, these two cell types have traditionally been grouped and studied as a single population. One likely reason for this is that conventional ploidy-sorting methods classify hepatocytes based solely on total DNA content, without distinguishing between mononucleated and binucleated states. In this study, we developed a novel flow cytometry strategy to distinguish and isolate 1x4n and 2x2n hepatocytes. Our approach leverages Hoechst-Area to assess total ploidy and Hoechst-Height to differentiate mononucleated and binucleated hepatocytes. Transcriptome analysis comparing these two populations revealed that 1x4n hepatocytes exhibit a broader and more metabolically active gene expression profile. Importantly, this enhanced metabolic activity was independent of liver zonation, a well-known driver of metabolic heterogeneity in hepatocytes. Our findings uncover a previously underappreciated layer of functional diversity in the liver and provide a new framework for studying the physiological and pathological roles of nuclear configuration in hepatocytes.

cell biology↗

bioRxiv2025

Attenuative effect of Grifola frondosa (maitake mushroom) on severe DSS-induced colitis in vitamin D-deficient mice

ScopeVitamin D deficiency, which has been a global health issue for decades, is involved in gut health and diseases. We examined the health benefits of Grifola frondosa (maitake mushroom), a potent dietary source of fungal vitamin D (D2), on DSS-induced colitis in vitamin D-deficient mice. Methods and resultsC57BL/6J mice were fed a control diet, vitamin D3-deficient diet (DD), maitake-fortified DD, or vitamin D2-fortified DD for 6 weeks. LC-MS/MS analysis demonstrated that maitake-fed mice showed an increased 25(OH)D2 alternative to 25(OH)D3 in plasma, as well as the mice fed an equivalent dose of vitamin D2. The mRNA expression profiles of vitamin D-responsive genes, including renal Cyp24a1 and Cyp27b1, were normalized in the maitake-fed mice. Severe DSS-induced colitis observed in DD mice was attenuated in maitake-fed mice; the accumulation of immune cells in the colonic mucosa and protein expression of colonic claudin-2, a target gene of the vitamin D receptor, were comparable to that in control mice. ConclusionDietary intake of maitake was effective in improving vitamin D status and biological function, demonstrating a potential attenuative effect on severe DSS-induced colitis in vitamin D3 deficient mice, as well as equivalent doses of vitamin D2.

biochemistry↗

bioRxiv2024

Modeling of vascular interactions in endochondral ossification using human embryonic stem cells-derived organoid on a microfluidic chip

Vascular interactions play a crucial role in embryogenesis, including skeletal development. During endochondral ossification, vascular networks are formed as mesenchymal cells condense and later invade skeletal elements to form the bone marrow. We and other groups developed a model of endochondral ossification by implanting human embryonic stem cell (hESC)-derived sclerotome into immunodeficient mice. However, in vitro models of endochondral ossification, particularly vascular interaction with mesenchymal cells at its initial stage, are yet to be established. Therefore, we developed a method to model the initial stage of endochondral ossification using a microfluidic chip-based platform, with a particular focus on the vascular interaction. On the chip, we found that the fibrin gel helped align mCherry-expressing human umbilical vein endothelial cells (HUVECs) better than the collagen-I gel, suggesting that the fibrin gel is more suitable for the formation of a vascular-like network. The perfusability of the vascular-like networks was partially confirmed using fluorescein isothiocyanate (FITC)-dextran and fluorescent microbeads. We then mixed hESC-derived sclerotome with enhanced green fluorescent protein (EGFP)-expressing HUVECs and applied this mixture on the chip. We named this mixture of cells SH organoids. The SH organoids showed superior abilities to maintain the vascular-like network, which was formed by the mCherry-expressing HUVECs, compared with the sclerotome spheroids on the chip. The EGFP-expressing HUVECs migrated from the SH organoid, formed a vascular-like networks, and partially interacted with the mCherry-expressing vascular-like networks on the chip. Histological analysis showed that SRY-box transcription factor 9 (SOX9) and type I collagen were expressed mutually exclusively in the condensed mesenchymal cells and perichondrial-like cells, respectively. This study demonstrates that our SH organoid-on-a-chip method reproduces vascular networks that are formed at the initial stage of endochondral ossification. This model may provide insights into human endochondral ossification and has potential applications in bone disease modeling and drug screening.

bioengineering↗

bioRxiv2024

Super homotypic targeting by exosome surface engineering

Homotypic targeting is the inherent ability of cells for preferential interaction with cells of similar or identical types, a phenomenon commonly seen in cell adhesion, tissue formation, and immune responses. Unfortunately, its full potential remains largely untapped. Here we introduce an approach to drastically boost the homotypic targeting capabilities of cells via exosomes (nanoscale extracellular vesicles secreted by cells). By engineering exosome surfaces with lanthanides, we amplify specific cell-exosome interactions by more than 25-fold, significantly accelerating the selective capture of exosomes by cells of the same lineage. This substantial enhancement in cellular homophilicity opens up an entirely new class of applications, two of which we showcase here with unprecedented performance: using cells to detect specific exosomes and using exosomes to detect specific cells. The concept of "super homotypic targeting" offers enormous potential to transform cancer diagnostics, immunotherapy, targeted drug delivery, tissue engineering, and vaccine development.

cell biology↗

bioRxiv2022

Porcine placenta extract upregulates ceramide synthase 3 expression via the PPARδ/ILK/Akt/mTOR/STAT3 pathway

Porcine placenta extract (PPE) is widely accepted as an ingredient in complementary and alternative medicine and previous studies have reported its availability, however, its underlying action mechanism remains unclear. In this study, we investigated the underlying mechanism of porcine placenta extract-induced ceramide synthase 3 upregulation. PPE enhanced the expression of ceramide synthase 3 at both the mRNA and protein levels in HaCaT cells. Moreover, porcine placenta extract-induced ceramide synthase3 upregulation was suppressed by the Akt inhibitor, suggesting the involvement of Akt in the underlying mechanism. As the PI3K inhibitor did not affect porcine placenta extract-induced ceramide synthase 3 upregulation, the factors upstream of Akt were estimated. Inhibition and small interfering RNA experiments demonstrated that the peroxisome proliferator-activated receptors {delta} (PPAR {delta}) and integrin-linked kinase (ILK) are involved in the phosphorylation of Akt. Next, we explored the factors downstream of Akt and found that porcine placenta extract induced phosphorylation of the STAT3 while porcine placenta extract-induced upregulation of ceramide synthase 3 was significantly suppressed by the inhibitor of the mTOR, suggesting that the mTOR/STAT3 pathway is involved in the downstream of Akt. These results demonstrate that porcine placenta extract upregulates CerS3 expression via the PPAR{delta}/ILK/Akt/mTOR/STAT3 pathway.

biochemistry↗