Biology subjects

Inui, M.

Publications and source records attributed to Inui, M..

6 recordsLinked to original sources

bioRxiv2026

A Col22a1-EGFP reporter reveals dynamic three-dimensional organization of the myotendinous junction

The myotendinous junction (MTJ) is a specialized extracellular matrix-rich interface that transmits contractile force generated by skeletal muscle to tendon, but its three-dimensional organization and dynamics remain poorly characterized. Here, we generate a Col22a1-EGFP knock-in mouse that enables direct visualization and quantitative analysis of MTJs from embryonic development to adulthood. Tissue-clearing and three-dimensional imaging reveal extensive variation in MTJ morphology and spatial distribution among skeletal muscles and between myofiber types. During embryonic MTJ formation, COL22A1-EGFP undergoes a striking transition from intracellular puncta within developing myofibers to extracellular accumulation at myofiber tips, associated with polarized Golgi organization. We further identify complex tendon protrusions that emerge during postnatal growth and connect tendon bodies with subsets of MTJs. These interfaces contain morphologically distinct Scx-lineage cells that express Col22a1, identifying a specialized tendon-associated population at the MTJ. Finally, muscle unloading and overload differentially alter COL22A1-EGFP signal intensity and area, respectively, demonstrating that the reporter provides a quantitative readout of MTJ responses to altered mechanical conditions. Thus, Col22a1-EGFP reveals previously unrecognized spatial, developmental, and adaptive features of the muscle-tendon interface and provides an experimental platform for investigating MTJ biology in vivo.

developmental biology↗

bioRxiv2026

Enterococcus faecalis is involved in the progression of the early stages of latent chronic pancreatitis surrounding pancreatic cancer tissue

(Objective) In our previous research, we identified latent chronic pancreatitis in the normal tissue surrounding pancreatic cancer. We also discovered the presence of Enterococcus faecalis (E. faecalis), a type of intestinal bacterium, in the pancreatic fluid and tissue of pancreatic cancer patients, suggesting it may be one of the factors contributing to the development of latent chronic pancreatitis. In this study, we performed pathological analyses to investigate its characteristics and investigate a possibility of E. faecalis infection. (Methods) Pathological analyses were performed, using 16 cases of pancreatic cancer and intraductal papillary mucinous neoplasia (IPMN) involving lesions in the pancreas tail. The involvement of E. faecalis was investigated with immunohistochemical analysis and serological methods. (Results) All cases exhibited inflammatory changes in pancreatic tissue without a clinical diagnosis of chronic pancreatitis, along with macrophage infiltration. These changes did not significantly differ according to preoperative treatment. DNA encoding E. faecalis 16s ribosomal RNA was detected in many cases, however, a positive immunostaining to E. faecalis was observed in only a few cases. Serum capsular polysaccharide (CPS) antibody levels exceeding the mean values were observed in patients with established chronic pancreatitis, while the level was not correlated with E. faecalis immunostaining. (Conclusion) These results suggest the E. faecalis infection is involved in the early stage of the progression of chronic latent pancreatitis and the diagnostic technology incorporating novel multi-biomarkers may be useful for identifying high-risk individuals for future pancreatic cancer development.

cancer biology↗

bioRxiv2026

Cardiac-Detargeted MyoAAV Enables Systemic Nrl-Mediated Fast Myofiber Remodeling and Hypertrophy Across Multiple Skeletal Muscles

Inducing fast myofiber programs offers therapeutic potential for skeletal muscle disorders such as sarcopenia, where fast myofibers are preferentially lost. Engineered muscle-specific AAV (MyoAAV) vectors enable efficient transduction of skeletal muscles after systemic administration; however, cardiac transgene expression limits applications requiring skeletal muscle-selective delivery. We generated modified MyoAAV vectors by incorporating cardiac-specific miR-208a target sequences into the transgene 3'UTR. This design markedly suppressed cardiac expression while preserving skeletal muscle output, with target-site variation enabling tunable trade-offs between cardiac detargeting and skeletal muscle expression levels. We validated this platform using neural retina leucine zipper (Nrl), a large Maf transcription factor regulating type IIb myofiber identity. Systemic delivery of conventional MyoAAV-Nrl caused severe cardiac hypertrophy and uniform lethality within one month. Conversely, incorporating miR-208a target sequences prevented detectable hypertrophy and eliminated mortality during the experimental observation period. This modification significantly reduced cardiac Nrl expression while maintaining skeletal muscle levels, successfully promoting type IIb myofiber formation and hypertrophy across multiple skeletal muscles. These findings demonstrate that miR-208a-mediated cardiac detargeting combined with MyoAAV-Nrl enables safe systemic induction of fast myofiber remodeling and hypertrophy, establishing a platform for gene therapies targeting skeletal muscle disorders associated with fast myofiber loss.

cell biology↗

bioRxiv2024

Automated Xeno-Free Chondrogenic Differentiation from Human Embryonic Stem Cells: Enhancing Efficiency and Ensuring High-Quality Mass Production

IntroductionRepairing damaged cartilage poses significant challenges, particularly in cases of congenital cartilage defects such as microtia or congenital tracheal stenosis, or as a consequence of traumatic injury, as the regenerative potential of cartilage is inherently limited. Stem cell therapy and tissue engineering offer promising approaches to overcome these limitations in cartilage healing. However, the challenge lies in the size of cartilage-containing organs, which necessitates a large quantity of cells to fill the damaged areas. Therefore, pluripotent stem cells that can proliferate indefinitely are highly desirable as a cell source. This study aims to delineate the differentiation conditions for cartilage derived from human embryonic stem cells (ESCs) and to develop an automated cell culture system to facilitate mass production for therapeutic applications. MethodsCartilage cell sheets were derived from human ESCs (SEES2, clinical trial-compatible line) by forming embryoid bodies (EBs) with either conventional manual culture or a benchtop multi-pipetter and an automated medium exchange integrated cell incubator, using xeno-free media. Cell sheets were implanted into the subcutaneous tissue of immunodeficient NOG mice to obtain cartilage tissue. The properties of cartilage tissues were examined by histological staining and quantitative PCR analysis. ResultsWe have optimized an efficient xeno-free system for cartilage production with the conventional culture method and successfully transitioned to an automated system. Differentiated cartilage was histologically uniform with cartilage-specific elasticity and strength. The cartilage tissues were stained by alcian blue, safranin O, and toluidine blue, and quantitative PCR showed an increase in differentiation markers such as ACAN, COL2A1, and Vimentin. Automation significantly enhanced the efficiency of human ESC-derived chondrocyte differentiation. The number of constituent cells within EBs and the seeding density of EBs were identified as key factors influencing chondrogenic differentiation efficiency. By automating the process of chondrogenic differentiation, we achieved scalable production of chondrocytes. ConclusionsBy integrating the differentiation protocol with an automated cell culture system, there is potential to produce cartilage of sufficient size for clinical applications in humans. The resulting cartilage tissue holds promise for clinical use in repairing organs such as the trachea, joints, ears, and nose.

cell biology↗

bioRxiv2021

Scx-positive tendon cells are required for correct muscle patterning

The elaborate movement of the vertebrate body is supported by the precise connection of muscle, tendon and bone. Each of the >600 distinct skeletal muscles in the human body has unique attachment sites; however, the mechanism through which muscles are reproducibly attached to designated partner tendons during embryonic development is incompletely understood. We herein show that Screlaxis-positive tendon cells have an essential role in correct muscle attachment in mouse embryos. Specific ablation of Screlaxis-positive cells resulted in dislocation of muscle attachment sites and abnormal muscle bundle morphology. Step-by-step observation of myogenic cell lineage revealed that post-fusion myofibers, but not migrating myoblasts, require tendon cells for their morphology. Furthermore, muscles could change their attachment site, even after the formation of the insertion. Our study demonstrated an essential role of tendon cells in the reproducibility and plasticity of skeletal muscle patterning, in turn revealing a novel tissue-tissue interaction in musculoskeletal morphogenesis. Graphical abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

developmental biology↗

bioRxiv2019

Lin28a/let-7 Pathway Modulates the Hox Code via Polycomb Regulation during Axial Patterning in Vertebrates

The body plan along the anteroposterior axis and regional identities are specified by the spatiotemporal expression of Hox genes. Multistep controls are required for their unique expression patterns; however, the molecular mechanisms behind the tight control of Hox genes are not fully understood. In this study, we demonstrated that the Lin28a/let-7 reciprocal regulatory pathway is critical for vertebral specification. Lin28a-/- mice exhibited homeotic transformations of vertebrae which were caused by the global dysregulation of posterior Hox genes. The accumulation of let-7-family microRNAs in Lin28a-/- mice resulted in the reduction of PRC1 occupancy at the Hox cluster loci by targeting Cbx2. Consistently, Lin28a loss in embryonic stem-like cells led to aberrant induction of posterior Hox genes, which was rescued by the knockdown of let-7-family microRNAs. These results suggest that Lin28/let-7 pathway is possibly involved in the modulation of the "Hox code" via Polycomb regulation during axial patterning in vertebrates.

developmental biology↗