Deep imaging of LepR+ stromal cells in optically cleared murine bone hemisections
Abstract Tissue clearing combined with high-resolution confocal imaging is a cutting-edge approach for dissecting the three-dimensional (3D) architecture of tissues and deciphering cellular spatial interactions under physiological and pathological conditions. Deciphering the spatial interaction of l...
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| Format: | Article |
| Language: | English |
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Nature Publishing Group
2025-01-01
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| Series: | Bone Research |
| Online Access: | https://doi.org/10.1038/s41413-024-00387-9 |
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| author | Yuehan Ni Jiamiao Wu Fengqi Liu Yating Yi Xiangjiao Meng Xiang Gao Luyi Xiao Weiwei Zhou Zexi Chen Peng Chu Dan Xing Ye Yuan Donghui Ding Ge Shen Min Yang Ronjie Wu Ling Wang Luiza Martins Nascentes Melo Sien Lin Xiaoguang Cheng Gang Li Alpaslan Tasdogan Jessalyn M. Ubellacker Hu Zhao Shentong Fang Bo Shen |
| author_facet | Yuehan Ni Jiamiao Wu Fengqi Liu Yating Yi Xiangjiao Meng Xiang Gao Luyi Xiao Weiwei Zhou Zexi Chen Peng Chu Dan Xing Ye Yuan Donghui Ding Ge Shen Min Yang Ronjie Wu Ling Wang Luiza Martins Nascentes Melo Sien Lin Xiaoguang Cheng Gang Li Alpaslan Tasdogan Jessalyn M. Ubellacker Hu Zhao Shentong Fang Bo Shen |
| author_sort | Yuehan Ni |
| collection | DOAJ |
| description | Abstract Tissue clearing combined with high-resolution confocal imaging is a cutting-edge approach for dissecting the three-dimensional (3D) architecture of tissues and deciphering cellular spatial interactions under physiological and pathological conditions. Deciphering the spatial interaction of leptin receptor-expressing (LepR+) stromal cells with other compartments in the bone marrow is crucial for a deeper understanding of the stem cell niche and the skeletal tissue. In this study, we introduce an optimized protocol for the 3D analysis of skeletal tissues, enabling the visualization of hematopoietic and stromal cells, especially LepR+ stromal cells, within optically cleared bone hemisections. Our method preserves the 3D tissue architecture and is extendable to other hematopoietic sites such as calvaria and vertebrae. The protocol entails tissue fixation, decalcification, and cryosectioning to reveal the marrow cavity. Completed within approximately 12 days, this process yields highly transparent tissues that maintain genetically encoded or antibody-stained fluorescent signals. The bone hemisections are compatible with diverse antibody labeling strategies. Confocal microscopy of these transparent samples allows for qualitative and quantitative image analysis using Aivia or Bitplane Imaris software, assessing a spectrum of parameters. With proper storage, the fluorescent signal in the stained and cleared bone hemisections remains intact for at least 2–3 months. This protocol is robust, straightforward to implement, and highly reproducible, offering a valuable tool for tissue architecture and cellular interaction studies. |
| format | Article |
| id | doaj-art-7bfe9982835d45019259dde4bc9473d1 |
| institution | Kabale University |
| issn | 2095-6231 |
| language | English |
| publishDate | 2025-01-01 |
| publisher | Nature Publishing Group |
| record_format | Article |
| series | Bone Research |
| spelling | doaj-art-7bfe9982835d45019259dde4bc9473d12025-01-19T12:13:41ZengNature Publishing GroupBone Research2095-62312025-01-0113111710.1038/s41413-024-00387-9Deep imaging of LepR+ stromal cells in optically cleared murine bone hemisectionsYuehan Ni0Jiamiao Wu1Fengqi Liu2Yating Yi3Xiangjiao Meng4Xiang Gao5Luyi Xiao6Weiwei Zhou7Zexi Chen8Peng Chu9Dan Xing10Ye Yuan11Donghui Ding12Ge Shen13Min Yang14Ronjie Wu15Ling Wang16Luiza Martins Nascentes Melo17Sien Lin18Xiaoguang Cheng19Gang Li20Alpaslan Tasdogan21Jessalyn M. Ubellacker22Hu Zhao23Shentong Fang24Bo Shen25College of Life Sciences, Beijing Normal UniversityNational Institute of Biological Sciences, Beijing (NIBS)School of Biopharmacy, China Pharmaceutical UniversityChinese Institute for Brain Research, Beijing (CIBR)National Institute of Biological Sciences, Beijing (NIBS)National Institute of Biological Sciences, Beijing (NIBS)National Institute of Biological Sciences, Beijing (NIBS)National Institute of Biological Sciences, Beijing (NIBS)Chinese Institute for Brain Research, Beijing (CIBR)National Institute of Biological Sciences, Beijing (NIBS)Arthritis Clinic and Research Center, Peking University People’s Hospital, Peking UniversityArthritis Clinic and Research Center, Peking University People’s Hospital, Peking UniversitySchool of Biopharmacy, China Pharmaceutical UniversityNational Institute of Biological Sciences, Beijing (NIBS)College of Life Sciences, Beijing Normal UniversityMusculoskeletal Research Laboratory, Department of Orthopaedics & Traumatology & Li Ka Shing Institute of Health Sciences, Faculty of Medicine, The Chinese University of Hong KongDepartment of Radiology, Beijing Jishuitan Hospital, Capital Medical University, National Center for OrthopaedicsDepartment of Dermatology, University Hospital Essen & German Cancer Consortium, Partner SiteMusculoskeletal Research Laboratory, Department of Orthopaedics & Traumatology & Li Ka Shing Institute of Health Sciences, Faculty of Medicine, The Chinese University of Hong KongDepartment of Radiology, Beijing Jishuitan Hospital, Capital Medical University, National Center for OrthopaedicsMusculoskeletal Research Laboratory, Department of Orthopaedics & Traumatology & Li Ka Shing Institute of Health Sciences, Faculty of Medicine, The Chinese University of Hong KongDepartment of Dermatology, University Hospital Essen & German Cancer Consortium, Partner SiteDepartment of Molecular Metabolism, Harvard T.H. Chan School of Public HealthChinese Institute for Brain Research, Beijing (CIBR)School of Biopharmacy, China Pharmaceutical UniversityNational Institute of Biological Sciences, Beijing (NIBS)Abstract Tissue clearing combined with high-resolution confocal imaging is a cutting-edge approach for dissecting the three-dimensional (3D) architecture of tissues and deciphering cellular spatial interactions under physiological and pathological conditions. Deciphering the spatial interaction of leptin receptor-expressing (LepR+) stromal cells with other compartments in the bone marrow is crucial for a deeper understanding of the stem cell niche and the skeletal tissue. In this study, we introduce an optimized protocol for the 3D analysis of skeletal tissues, enabling the visualization of hematopoietic and stromal cells, especially LepR+ stromal cells, within optically cleared bone hemisections. Our method preserves the 3D tissue architecture and is extendable to other hematopoietic sites such as calvaria and vertebrae. The protocol entails tissue fixation, decalcification, and cryosectioning to reveal the marrow cavity. Completed within approximately 12 days, this process yields highly transparent tissues that maintain genetically encoded or antibody-stained fluorescent signals. The bone hemisections are compatible with diverse antibody labeling strategies. Confocal microscopy of these transparent samples allows for qualitative and quantitative image analysis using Aivia or Bitplane Imaris software, assessing a spectrum of parameters. With proper storage, the fluorescent signal in the stained and cleared bone hemisections remains intact for at least 2–3 months. This protocol is robust, straightforward to implement, and highly reproducible, offering a valuable tool for tissue architecture and cellular interaction studies.https://doi.org/10.1038/s41413-024-00387-9 |
| spellingShingle | Yuehan Ni Jiamiao Wu Fengqi Liu Yating Yi Xiangjiao Meng Xiang Gao Luyi Xiao Weiwei Zhou Zexi Chen Peng Chu Dan Xing Ye Yuan Donghui Ding Ge Shen Min Yang Ronjie Wu Ling Wang Luiza Martins Nascentes Melo Sien Lin Xiaoguang Cheng Gang Li Alpaslan Tasdogan Jessalyn M. Ubellacker Hu Zhao Shentong Fang Bo Shen Deep imaging of LepR+ stromal cells in optically cleared murine bone hemisections Bone Research |
| title | Deep imaging of LepR+ stromal cells in optically cleared murine bone hemisections |
| title_full | Deep imaging of LepR+ stromal cells in optically cleared murine bone hemisections |
| title_fullStr | Deep imaging of LepR+ stromal cells in optically cleared murine bone hemisections |
| title_full_unstemmed | Deep imaging of LepR+ stromal cells in optically cleared murine bone hemisections |
| title_short | Deep imaging of LepR+ stromal cells in optically cleared murine bone hemisections |
| title_sort | deep imaging of lepr stromal cells in optically cleared murine bone hemisections |
| url | https://doi.org/10.1038/s41413-024-00387-9 |
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