Imaging the bone-immune cell interaction in bone destruction

Bone is a highly dynamic organ that is continuously being remodeled by the reciprocal interactions between bone and immune cells. We have originally established an advanced imaging system for visualizing the in vivo behavior of osteoclasts and their precursors in the bone marrow cavity using two-photon microscopy. Using this system, we found that the blood-enriched lipid mediator, sphingosine-1-phosphate, controlled the migratory behavior of osteoclast precursors. We also developed pH-sensing chemical fluorescent probes to detect localized acidification by bone-resorbing osteoclasts on the bone surface in vivo, and identified two distinct functional states of differentiated osteoclasts, “bone-resorptive” and “non-resorptive.” Here, we summarize our studies on the dynamics and functions of bone and immune cells within the bone marrow. We further discuss how our intravital imaging techniques can be applied to evaluate the mechanisms of action of biological agents in inflammatory bone destruction. Our intravital imaging techniques would be beneficial for studying the cellular dynamics in arthritic inflammation and bone destruction in vivo and would also be useful for evaluating novel therapies in animal models of bone-destroying diseases.Hasegawa T., Kikuta J., Ishii M.. Imaging the bone-immune cell interaction in bone destruction. Frontiers in Immunology 10, 596 (2019); https://doi.org/10.3389/fimmu.2019.00596

網膜静脈分枝閉塞症に伴う黄斑浮腫眼おける光干渉断層計での中心窩視細胞内節外節ラインの隆起 (foveal bulge) の評価

PURPOSE: To determine whether a significant correlation exists between the presence of a bulge in the photoreceptor inner segment/outer segment (IS/OS) line and the best-corrected visual acuity (BCVA) in eyes with resolved macular edema associated with branch retinal vein occlusion (BRVO). DESIGN: Retrospective, observational case series. METHODS: We retrospectively reviewed the medical records of patients who had a complete resolution of macular edema and had an intact IS/OS line in the central fovea in the spectral-domain optical coherence tomographic (SDOCT) images. Thirty-one eyes with macular edema associated with BRVO (BRVO group) and 31 unaffected fellow eyes (control group) of 31 patients were evaluated. In normal eyes, the intact IS/OS line determined by SDOCT has a bulge at the central fovea, called the foveal bulge. The eyes in the BRVO group were classified by the presence or absence of foveal bulge, and the characteristics of the 2 groups were compared. RESULTS: A foveal bulge was present in 7 of 31 eyes in the BRVO group. The incidence of a foveal bulge was significantly lower in the BRVO group (22.6%) than in the control group (100%; P < .0001). All 7 eyes with foveal bulge had a decimal BCVA of ≥1.0 at the final visit. The incidence of a foveal bulge was significantly higher in eyes with BCVA of ≥1.0 (77.8%) than in the eyes with BCVA of <1.0 (0%; P < .0001). CONCLUSIONS: The foveal bulge is a good marker of the functional properties of the fovea in eyes with resolved macular edema associated with BRVO.博士（医学）・乙第1416号・平成30年3月15日Copyright © 2014 Elsevier Inc. All rights reserved

Imaging the Bone-Immune Cell Interaction in Bone Destruction

Bone is a highly dynamic organ that is continuously being remodeled by the reciprocal interactions between bone and immune cells. We have originally established an advanced imaging system for visualizing the in vivo behavior of osteoclasts and their precursors in the bone marrow cavity using two-photon microscopy. Using this system, we found that the blood-enriched lipid mediator, sphingosine-1-phosphate, controlled the migratory behavior of osteoclast precursors. We also developed pH-sensing chemical fluorescent probes to detect localized acidification by bone-resorbing osteoclasts on the bone surface in vivo, and identified two distinct functional states of differentiated osteoclasts, “bone-resorptive” and “non-resorptive.” Here, we summarize our studies on the dynamics and functions of bone and immune cells within the bone marrow. We further discuss how our intravital imaging techniques can be applied to evaluate the mechanisms of action of biological agents in inflammatory bone destruction. Our intravital imaging techniques would be beneficial for studying the cellular dynamics in arthritic inflammation and bone destruction in vivo and would also be useful for evaluating novel therapies in animal models of bone-destroying diseases

Establishment and evaluation of the suspension culture system for umbilical cord- derived mesenchymal stromal cells

Mesenchymal stromal cells (MSCs) derived from various tissues including bone marrow, adipose and umbilical cord tissues have been shown to modulate aberrantly activated immune system. With the features, MSC-based therapies targeting graft-versus-host disease (GvHD) by the administration of bone marrow-derived MSCs (BM-MSCs) have been available in some countries including Japan, and the expectations for the stable and cost effective supply system are getting higher and higher recently. However, the conventional culture systems which usually use plastic flask or multi-chamber equipment require space and manpower, thus the maximal expansion of MSCs at one production is likely to be limited. To compensate the limitation, repetitive productions have been unavoidable, and higher the production cost. Here, we introduced a new suspension-culture system, using micro-carriers and single-use-bioreactors, for the preparation of MSCs in anticipation of establishment of mass production system. Since the umbilical cord (UC) tissues can be collected through noninvasive procedure, and UC-derived MSCs (UC-MSCs) are shown to present higher proliferation rate and lower immunogenicity in comparison with BM-MSCs, we evaluated the potential and the versatility of UC-MSCs for the treatment of several diseases including GvHD. Results from several in vitro assays demonstrated that our new culture system maintains major key characteristics of MSCs, such as adhesiveness to cell culture surface, the expression of cell surface markers, differentiation capacities toward osteoblasts, chondroblasts, and adipocytes, and immunosuppressive effects on activated T cells. We are currently investigating cellular profiles and characteristics which are specific to the cells prepared in our suspension-culture system through meta-analysis. The established suspension-culture system is presumed to attain the mass production of UC-MSCs, contributing to lower the cost and also providing possible applications for MSCs from other origins

Development of an intravital imaging system for the synovial tissue reveals the dynamics of CTLA-4 Ig in vivo

There have been many attempts to visualize the inflamed joints using multiphoton microscopy. However, due to the hypervascular and multilayered structure of the inflamed synovium, intravital imaging of the deep synovial tissue has been difficult. Here, we established original intravital imaging systems to visualize synovial tissue and pathological osteoclasts at the pannus–bone interface using multiphoton microscopy. Combined with fluorescence-labeling of CTLA-4 Ig, a biological agent used for the treatment of rheumatoid arthritis, we identified that CTLA-4 Ig was distributed predominantly within the inflamed synovium and bound to CX3CR1+ macrophages and CD140a+ fibroblasts 6 h after injection, but not to mature osteoclasts. Intravital imaging of blood and lymphatic vessels in the inflamed synovium further showed that extravasated CTLA-4 Ig was immediately drained through lymphatic vessels under acute arthritic conditions, but the drainage activity was retarded under chronic conditions. These results indicate that this intravital synovial imaging system can serve as a platform for exploring the dynamics of immune cells, osteoclasts, and biological agents within the synovial microenvironment in vivo.Hasegawa T., Kikuta J., Sudo T., et al. Development of an intravital imaging system for the synovial tissue reveals the dynamics of CTLA-4 Ig in vivo. Scientific Reports 10, 13480 (2020); https://doi.org/10.1038/s41598-020-70488-y