Biology subjects

Yagasaki, R.

Publications and source records attributed to Yagasaki, R..

3 recordsLinked to original sources

bioRxiv2024

Ex vivo SIM-AFM measurements reveal the spatial correlation of stiffness and molecular distributions in 3D living tissue

Living tissues each exhibit a distinct stiffness, which provides cells with key environmental cues that regulate their behaviors. Despite this significance, our understanding of the spatiotemporal dynamics and the biological roles of stiffness in three-dimensional tissues is currently limited due to a lack of appropriate measurement techniques. To address this issue, we propose a new method combining upright structured illumination microscopy (USIM) and atomic force microscopy (AFM) to obtain precisely coordinated stiffness maps and biomolecular fluorescence images of thick living tissue slices. Using mouse embryonic skin as a representative tissue with mechanically heterogeneous structures inside, we validate the measurement principle of USIM-AFM. Live measurement of tissue stiffness distributions revealed the highly heterogeneous mechanical nature of embryonic skin as well as the role of collagens in maintaining its integrity. Furthermore, quantitative comparisons of stiffness distributions of preserved tissue samples unveiled the distinct impacts of conventional tissue preservation techniques on the tissue stiffness pattern. This series of experiments highlights the importance of live mechanical testing of tissue-scale samples. Our USIM-AFM technique provides a new methodology to reveal the dynamic nature of tissue stiffness and its correlation with biomolecular distributions in live tissues and thus could serve as a technical basis for exploring tissue-scale mechanobiology. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=89 SRC="FIGDIR/small/595975v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@8a32f5org.highwire.dtl.DTLVardef@117a350org.highwire.dtl.DTLVardef@e2fb99org.highwire.dtl.DTLVardef@1f41d24_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2024

The gut contractile organoid: a novel model for studying the gut motility regulated by coordinating signals between interstitial cells of Cajal and smooth muscles

The gut undergoes peristaltic movements regulated by intricate cellular interactions. How these interactions emerge in the developing gut remains poorly explored due to a lack of model system. We here developed a novel contractile organoid that is derived from the muscle layer of chicken embryonic hindgut. The organoid contained smooth muscle cells (SMCs) and interstitial cells of Cajal (ICCs; pacemaker) with few enteric neurons, and underwent periodic contractions. The organoid was formed by self-organization with morphological arrangements of ICCs (internal) and SMCs (peripheral), allowing identification of these cells in live. GCaMP-Ca2+ imaging analyses revealed that Ca2+ transients between ICC-ICC, SMC-SMC or SMC-ICC were markedly coordinated. Pharmacological studies further suggested a role of gap junctions in ICC-to-SMC signaling, and also possible mechanical feedback from SMCs contraction to ICCs pace-making activities. In addition, two organoids with different rhythm became synchronized when mediated by SMCs, unveiling a novel contribution of SMCs to ICCs pace-making. The gut contractile organoid developed in this study offers a useful model to understand how the rhythm coordination between/among ICCs and SMCs is regulated and maintained during gut development.

developmental biology↗

bioRxiv2022

Newly raised anti-c-Kit antibody detects interstitial cells of Cajal in the gut of chicken embryos

The gut peristaltic movement, a wave-like propagation of a local contraction, is important for the transportation and digestion of ingested materials. Among three types of cells, the enteric nervous system (ENS), smooth muscle cells, and interstitial cells of Cajal (ICCs), the ICCs have been thought to act as a pacemaker, and therefore it is important to decipher the cellular functions of ICCs for the understanding of gut peristalsis. c-Kit, a tyrosine kinase receptor, has widely been used as a marker for ICCs. Most studies with ICCs have been conducted in mammals using commercially available anti-c-Kit antibody. Recently, the chicken embryonic gut has emerged as a powerful model to study the gut peristalsis. However, since the anti-c-Kit antibody for mammals does not work for chickens, cellular mechanisms by which ICCs are regulated have largely been unexplored. Here, we report a newly raised polyclonal antibody against the chicken c-Kit protein. The specificity of the antibody was validated by both Western blotting analyses and immunocytochemistry. Co-immunostaining with the new antibody and anti- smooth muscle actin (SMA) antibody successfully visualized ICCs in the chicken developing hindgut in the circular muscle- and longitudinal muscle layers: as previously shown in mice, common progenitors of ICCs and smooth muscle cells at early stages were double positive for SMA and c-Kit, and at later stages, differentiated ICCs and smooth muscle cells exhibited only c-Kit and SMA, respectively. A novel ICC population was also found that radially extended from the submucosal layer to circular muscle layer. Furthermore, the new antibody delineated individual ICCs in a cleared hindgut. The antibody newly developed in this study will facilitate the study of peristaltic movement in chicken embryos.

developmental biology↗