Biology subjects

Ishihara, S.

Publications and source records attributed to Ishihara, S..

27 records · Page 2Linked to original sources

bioRxiv2022

Scale space calibrates present and subsequent spatial learning in Barnes maze in mice

Animals including humans are capable of representing different scale spaces from smaller to larger ones. However, most laboratory animals live their life in a narrow range of scale spaces like home-cages and experimental setups, making it hard to extrapolate the spatial representation and learning process in large scale spaces from those in conventional scale spaces. Here, we developed a 3-meter diameter Barnes maze (BM3), then explored whether spatial learning in Barnes maze (BM) is calibrated by scale spaces. In the BM3, mice exhibited lower learning rate compared to a conventional 1-meter diameter Barnes maze (BM1), suggesting that the BM3 requires more trial-and-error and larger computational resources to solve the task than the BM1. Analyzing network structures of moving trajectories, betweenness centrality would contrast spatial learning in a larger scale space with that in a smaller one, as it diverges between the BM1 and the BM3 along with the learning progression. We then explored whether prior learning in either BM scale calibrates subsequent spatial learning in the other BM scale, and found asymmetric facilitation such that the prior learning in the BM3 facilitated the subsequent learning in the BM1, but not vice versa. Network structures of trajectories in the subsequent BM scale were changed by both prior and subsequent BM scale. These results suggest that scale space calibrates both the present and subsequent BM learning. This is the first study to explore and demonstrate scale-dependent spatial learning in Barnes maze in mice. Significance StatementAnimals are capable of representing different scale spaces. However, whether scale space calibrates goal-directed spatial learning remains unclear. The Barnes maze is a well-established experimental paradigm to evaluate spatial learning in rodents. Here, we developed a larger scale 3-meter diameter Barnes maze (BM3) then compared various navigation features in mice between the BM3 and a conventional 1-meter diameter Barnes maze (BM1). We demonstrated that learning on the BM3 required more computational resources than in the BM1, prompting mice to exploit unique navigation patterns. Such learning experiences in the BM3 facilitated subsequent spatial learning in the BM1, but not vice versa. These results suggest that scale space calibrates immediate and subsequent spatial learning.

animal behavior and cognition↗

bioRxiv2022

Attachment/detachment of cortical myosin regulates cell junction exchange during cell rearrangement

Epithelial cells remodel cell adhesion and change their neighbors to shape a tissue. This cell rearrangement proceeds in three steps: the shrinkage of a junction, exchange of junctions, and elongation of the newly generated junction. Herein, by combining live imaging and physical modeling, we showed that the formation of myosin-II (myo-II) cables around the cell vertices underlies the exchange of junctions. The local and transient detachment of myo-II from the cell cortex is coupled with the junction shrinkage and elongation via an interplay between the LIM domain-containing protein Jub and the tricellular junction protein M6. Furthermore, we developed a mechanical model based on the wetting theory and clarified the way by which the physical properties of myo-II cables are integrated with the junction geometry to induce the transition between the attached and detached states and support the unidirectionality of cell rearrangement. Collectively, the present study elucidates the orchestration of geometry, mechanics and signaling for exchanging junctions.

cell biology↗

bioRxiv2022

Age-associated changes in knee osteoarthritis, pain-related behaviors, and dorsal root ganglia immunophenotyping of male and female mice

ObjectiveOsteoarthritis (OA) is a leading cause of chronic pain, yet OA pain management remains poor. Age is the strongest predictor of OA development, and mechanisms driving OA pain are unclear. While injury-induced OA models are useful, only a subset of OA is linked to traumatic injury. Here, we aimed to characterize age-associated joint damage, mechanical sensitization, and dorsal root ganglia (DRG) immune phenotypes in mice of both sexes. MethodsMale or female mice aged 6- or 20-months old were evaluated for histopathologic knee OA, pain-related behaviors, and L3-L5 dorsal root ganglia (DRG) immune characterization via flow cytometry. DRG gene expression in aged mice and humans was also examined. ResultsTwenty-month old male mice had worse cartilage degeneration than 6-month old mice. Older female knees showed increased cartilage degeneration, but to a lesser degree than males. Older mice of both sexes had worse mechanical allodynia, knee hyperalgesia, and grip strength compared to younger mice. For both sexes, DRGs from older mice showed decreased CD45+ cells, and a significant increase in F4/80+ macrophages and CD11c+ dendritic cells. Older male DRGs showed increased expression of Ccl2 and Ccl5 and older female DRGs showed increased Cxcr4 and Ccl3 compared to 6-month DRGs, among other differentially expresssed genes. Human DRG analysis from six individuals >80 years old revealed elevated CCL2 in male DRGs compared to females, whereas CCL3 was higher in female DRGs. ConclusionsHere we show that aging in male and female mice is accompanied by mild knee OA, mechanical sensitization, and changes to immune cell populations in the DRG, suggesting novel avenues for development of analgesic therapies.

cell biology↗

bioRxiv2022

Piezo2 expressing nociceptors mediate mechanical sensitization in experimental osteoarthritis

Osteoarthritis is a very common painful joint disease, for which few treatment options exist. New non-opioid targets are needed for addressing osteoarthritis pain, which is mechanical in nature and associated with daily activities such as walking and climbing stairs. Piezo2 has been implicated in development of mechanical pain, but the mechanisms by which this occurs remain poorly understood. We observed that in two different murine models of osteoarthritis (destabilization of the medial meniscus and natural aging), nociceptor-specific Piezo2 conditional knock-out mice developed osteoarthritic joint damage, but were protected from associated mechanical sensitization. Since nerve growth factor (NGF) is known to mediate nociceptor sensitization, and antibodies that neutralize NGF are effective as a treatment for osteoarthritis pain, we explored the effects of intra-articularly injected NGF on the development of mechanical joint pain. Wild-type mice developed knee swelling and mechanical pain in response to intra-articular NGF, while nociceptor-specific Piezo2 conditional knock-out mice were protected from these effects. Single cell RNA sequencing and in situ hybridization of mouse and human lumbar dorsal root ganglia (DRG) revealed that a subset of nociceptors co-express Piezo2 and Ntrk1 (the gene that encodes the NGF receptor TrkA). These results indicate that Piezo2 plays a key role in nociceptor sensitization processes in the osteoarthritic joint, and targeting Piezo2 may represent a novel therapy for osteoarthritis pain control. One Sentence SummaryNociceptor sensitization to mechanical stimuli is dependent on Piezo2 in mouse models of osteoarthritis.

physiology↗

bioRxiv2021

Conversion of cancer-associated fibroblasts from pro- to antitumor improves the sensitivity of pancreatic cancer to chemotherapeutics

Previous therapeutic attempts to deplete cancer-associated fibroblasts (CAFs) or inhibit their proliferation in pancreatic ductal adenocarcinoma (PDAC) were not successful in mice or patients. Thus, CAFs may be tumor suppressive or heterogeneous, with distinct cancer-restraining and -promoting CAFs (rCAFs and pCAFs, respectively). Here, we show that induced expression of the glycosylphosphatidylinositol-anchored protein Meflin, a rCAF-specific marker, in CAFs by genetic and pharmacological approaches improved the chemosensitivity of mouse PDAC. A chemical library screen identified Am80, a synthetic, non-natural retinoid, as a reagent that effectively induced Meflin expression in CAFs. Am80 administration improved the sensitivity of PDAC to chemotherapeutics, accompanied by increases in tumor vessel area and intratumoral drug delivery. Mechanistically, Meflin was involved in the suppression of tissue stiffening by interacting with lysyl oxidase to inhibit its collagen crosslinking activity. These data suggested that modulation of CAF heterogeneity may represent a strategy for PDAC treatment.

cancer biology↗

bioRxiv2020

Comparative mapping of crawling-cell morphodynamics in deep learning-based feature space

Navigation of fast migrating cells such as amoeba Dictyostelium and immune cells are tightly associated with their morphologies that range from steady polarized forms that support high directionality to those more complex and variable when making frequent turns. Model simulations are essential for quantitative understanding of these features and their origins, however systematic comparisons with real data are underdeveloped. Here, by employing deep-learning-based feature extraction combined with phase-field modeling framework, we show that a low dimensional feature space for 2D migrating cell morphologies obtained from the shape stereotype of keratocytes, Dictyostelium and neutrophils can be fully mapped by interlinked signaling network of cell-polarization and protrusion dynamics. Our analysis links the data-driven shape analysis to the underlying causalities by identifying key parameters critical for migratory morphologies both normal and aberrant under genetic and pharmacological perturbations. The results underscore the importance of deciphering self-organizing states and their interplay when characterizing morphological phenotypes.

cell biology↗

bioRxiv2020

Macrophages drive the inflammatory phase in experimental osteoarthritis

Macrophages fulfill critical functions in maintaining tissue homeostasis in steady-state, as well as in inflammation and immune response. Inflammation is not considered a major driver of osteoarthritis (OA), but macrophages have been implicated in its pathogenesis. Here, we use two mouse models of experiment OA - collagenase-induced osteoarthritis (CIOA) and destabilization of the medial meniscus (DMM) - to quantify the immune cell infiltration into the knee joint during the early stages of disease. We find that the peak of inflammation occurs at day 3 in CIOA and is characterized by a transitory increase in neutrophils and monocytes and a longer-lived expansion of synovial macrophages. Macrophage sub-populations are disproportionally expanded with CX3CR1+ cells accounting for a larger proportion of the macrophage compartment. Transcriptional profiling demonstrates that synovial macrophages up-regulate inflammatory genes coinciding with peak inflammation and down-regulate genes associated with homeostasis and tissue-residence. Female mice exhibit a similar expansion of macrophages post-CIOA indicating that the inflammatory phase is not sex-specific. Finally, we find day 7 post-DMM is also characterized by increases in neutrophil, monocyte, and macrophage sub-population numbers. These results support a role for macrophages in early stages of OA through driving the inflammatory phase. Further investigation may elucidate potential targets for the prevention or attenuation of OA-associated cartilage damage.

immunology↗

bioRxiv2019

The local state of chromatin compaction at transcription start sites controls transcription levels

A defined amount of transcript is produced from transcription start sites (TSSs) of each gene, suggesting that the binding frequency of RNA polymerase varies among genes. However, what structure in chromatin controls this frequency remains elusive. We established a method to fractionate chromatin according to its degree of three-dimensional compaction. Nucleosomes were evenly detected through all of the fractions, but histone H1 was more highly enriched in the more compact chromatin fractions. Similarly, HP1 and MBD2b were more abundant in more compact chromatin, while the levels of tri-methylated H3 (Lys9) and 5-methyl cytosine subtly increased. Via genome-wide analyses, nearly the entire genome was found to exist in compact chromatin without variations between repeat and non-repeat sequences; however, active TSSs were rarely found in compact chromatin. Based on a correlation between weak compaction and RNA polymerase binding at TSSs, it appears that local states of chromatin compaction determine transcription levels.

molecular biology↗

bioRxiv2017

An aggrecan fragment drives osteoarthritis pain through Toll-like receptor 2

Pain is the predominant symptom of osteoarthritis, but the connection between joint damage and the genesis of pain is not well understood. Loss of articular cartilage is a hallmark of osteoarthritis, and it occurs through enzymatic degradation of aggrecan by ADAMTS-4/5-mediated cleavage in the interglobular domain (E373-374 A). Further cleavage by MMPs (N341-342 F) releases a 32-amino-acid aggrecan fragment (32-mer). We investigated the role of this 32-mer in driving joint pain. We demonstrated that the 32-mer excites dorsal root ganglion (DRG) nociceptive neurons, both in culture and in intact explants. Treatment of cultured sensory neurons with the 32-mer induced them to express the pro-algesic chemokine, MCP-1/CCL2. These effects were mediated through Toll-like receptor (TLR)2, which we demonstrated was expressed by nociceptive neurons. In addition, intra-articular injection of the 32-mer provoked knee hyperalgesia in wild-type but not Tlr2 null mice. Blocking the production or action of the 32-mer in transgenic mice prevented the development of knee hyperalgesia in a murine model of osteoarthritis. These findings suggest that the aggrecan 32-mer fragment directly activates TLR2 on joint nociceptors and is an important mediator of the development of osteoarthritis-associated joint pain.

neuroscience↗