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Topic: Innovative concepts, new technologies, case reports
Geurts, Jeroen(1)*;Wen, Chunyi(2);Briki, Myriam(1);Moradpour, Elke(1);Hügle, Thomas(1);
(1)Lausanne University Hospital, Rheumatology, Lausanne, Switzerland;(2)Hong Kong Polytechnic University, Biomedical Engineering, Hong Kong, Hong Kong;
Abstract Text
Background/Introduction: Osteoarthritis (OA) is characterized by progressive degeneration of articular cartilage and remodelling of subchondral bone and marrow adipose tissue contained therein. It is commonly believed that OA progresses by abnormal bone turnover (hypomineralization) as well as increased bone angiogenesis and vascular invasion of articular cartilage. Present knowledge of these pathological features stems from 2D histological studies only.
Purpose: To evaluate feasibility of tissue clearing with advanced 3D microscopy for ex vivo imaging of vascularization and mineralization of human osteochondral tissues.Methods: Tissue specimens (2x2cm2) were obtained from regions with mild and severe degeneration of human knee OA tibial plateaus (n=8). The calcification front was identified by en bloc 0.01% Alizarin red staining (Aaron et al 1984). For vascular imaging, decalcified were stained with AF647-labeled CD34 antibody and propidium iodine. Sagittal 1-2mm thick sections were cleared using an optimized ethyl cinnamate (ECi) method (Masselink et al 2019). 3D imaging was performed using stereofluorescence microscopy and light sheet fluorescence microscopy (LSFM).
Results: Stereomicroscopy revealed a sharp calcification demarcation line between calcified and non-calcified cartilage and discreet staining of trabecular bone. Subchondral vasculature in undecalcified bone could be visualized by tissue autofluorescence (Top image). The cartilage calcification front was disrupted in severely degenerative tissue and subjacent sclerotic bone showed reduced calcification, but increased vascular density. LSFM ex vivo vascular imaging displayed blood vessels originating from bone marrow terminating in calcified cartilage (Lower image). Vascular invasion into non-decalcified cartilage was not observed in mild or severe OA grades. Vascular density and vessel size appeared both increased in severe compared with mild OA grade.
Conclusion: We provided proof of concept for ECi-based optical 3D ex vivo imaging of human OA osteochondral tissue. This method opens up new avenues for spatial analysis of cells/structures in bone and joint research. 
Topic: Innovative concepts, new technologies, case reports
Geurts, Jeroen(1)*;Wen, Chunyi(2);Briki, Myriam(1);Moradpour, Elke(1);Hügle, Thomas(1);
(1)Lausanne University Hospital, Rheumatology, Lausanne, Switzerland;(2)Hong Kong Polytechnic University, Biomedical Engineering, Hong Kong, Hong Kong;
Abstract Text
Background/Introduction: Osteoarthritis (OA) is characterized by progressive degeneration of articular cartilage and remodelling of subchondral bone and marrow adipose tissue contained therein. It is commonly believed that OA progresses by abnormal bone turnover (hypomineralization) as well as increased bone angiogenesis and vascular invasion of articular cartilage. Present knowledge of these pathological features stems from 2D histological studies only.
Purpose: To evaluate feasibility of tissue clearing with advanced 3D microscopy for ex vivo imaging of vascularization and mineralization of human osteochondral tissues.Methods: Tissue specimens (2x2cm2) were obtained from regions with mild and severe degeneration of human knee OA tibial plateaus (n=8). The calcification front was identified by en bloc 0.01% Alizarin red staining (Aaron et al 1984). For vascular imaging, decalcified were stained with AF647-labeled CD34 antibody and propidium iodine. Sagittal 1-2mm thick sections were cleared using an optimized ethyl cinnamate (ECi) method (Masselink et al 2019). 3D imaging was performed using stereofluorescence microscopy and light sheet fluorescence microscopy (LSFM).
Results: Stereomicroscopy revealed a sharp calcification demarcation line between calcified and non-calcified cartilage and discreet staining of trabecular bone. Subchondral vasculature in undecalcified bone could be visualized by tissue autofluorescence (Top image). The cartilage calcification front was disrupted in severely degenerative tissue and subjacent sclerotic bone showed reduced calcification, but increased vascular density. LSFM ex vivo vascular imaging displayed blood vessels originating from bone marrow terminating in calcified cartilage (Lower image). Vascular invasion into non-decalcified cartilage was not observed in mild or severe OA grades. Vascular density and vessel size appeared both increased in severe compared with mild OA grade.
Conclusion: We provided proof of concept for ECi-based optical 3D ex vivo imaging of human OA osteochondral tissue. This method opens up new avenues for spatial analysis of cells/structures in bone and joint research.
Topic: Innovative concepts, new technologies, case reports
Geurts, Jeroen(1)*;Wen, Chunyi(2);Briki, Myriam(1);Moradpour, Elke(1);Hügle, Thomas(1);
(1)Lausanne University Hospital, Rheumatology, Lausanne, Switzerland;(2)Hong Kong Polytechnic University, Biomedical Engineering, Hong Kong, Hong Kong;
Abstract Text
Background/Introduction: Osteoarthritis (OA) is characterized by progressive degeneration of articular cartilage and remodelling of subchondral bone and marrow adipose tissue contained therein. It is commonly believed that OA progresses by abnormal bone turnover (hypomineralization) as well as increased bone angiogenesis and vascular invasion of articular cartilage. Present knowledge of these pathological features stems from 2D histological studies only.
Purpose: To evaluate feasibility of tissue clearing with advanced 3D microscopy for ex vivo imaging of vascularization and mineralization of human osteochondral tissues.Methods: Tissue specimens (2x2cm2) were obtained from regions with mild and severe degeneration of human knee OA tibial plateaus (n=8). The calcification front was identified by en bloc 0.01% Alizarin red staining (Aaron et al 1984). For vascular imaging, decalcified were stained with AF647-labeled CD34 antibody and propidium iodine. Sagittal 1-2mm thick sections were cleared using an optimized ethyl cinnamate (ECi) method (Masselink et al 2019). 3D imaging was performed using stereofluorescence microscopy and light sheet fluorescence microscopy (LSFM).
Results: Stereomicroscopy revealed a sharp calcification demarcation line between calcified and non-calcified cartilage and discreet staining of trabecular bone. Subchondral vasculature in undecalcified bone could be visualized by tissue autofluorescence (Top image). The cartilage calcification front was disrupted in severely degenerative tissue and subjacent sclerotic bone showed reduced calcification, but increased vascular density. LSFM ex vivo vascular imaging displayed blood vessels originating from bone marrow terminating in calcified cartilage (Lower image). Vascular invasion into non-decalcified cartilage was not observed in mild or severe OA grades. Vascular density and vessel size appeared both increased in severe compared with mild OA grade.
Conclusion: We provided proof of concept for ECi-based optical 3D ex vivo imaging of human OA osteochondral tissue. This method opens up new avenues for spatial analysis of cells/structures in bone and joint research.
Topic: Innovative concepts, new technologies, case reports
Geurts, Jeroen(1)*;Wen, Chunyi(2);Briki, Myriam(1);Moradpour, Elke(1);Hügle, Thomas(1);
(1)Lausanne University Hospital, Rheumatology, Lausanne, Switzerland;(2)Hong Kong Polytechnic University, Biomedical Engineering, Hong Kong, Hong Kong;
Abstract Text
Background/Introduction: Osteoarthritis (OA) is characterized by progressive degeneration of articular cartilage and remodelling of subchondral bone and marrow adipose tissue contained therein. It is commonly believed that OA progresses by abnormal bone turnover (hypomineralization) as well as increased bone angiogenesis and vascular invasion of articular cartilage. Present knowledge of these pathological features stems from 2D histological studies only.
Purpose: To evaluate feasibility of tissue clearing with advanced 3D microscopy for ex vivo imaging of vascularization and mineralization of human osteochondral tissues.Methods: Tissue specimens (2x2cm2) were obtained from regions with mild and severe degeneration of human knee OA tibial plateaus (n=8). The calcification front was identified by en bloc 0.01% Alizarin red staining (Aaron et al 1984). For vascular imaging, decalcified were stained with AF647-labeled CD34 antibody and propidium iodine. Sagittal 1-2mm thick sections were cleared using an optimized ethyl cinnamate (ECi) method (Masselink et al 2019). 3D imaging was performed using stereofluorescence microscopy and light sheet fluorescence microscopy (LSFM).
Results: Stereomicroscopy revealed a sharp calcification demarcation line between calcified and non-calcified cartilage and discreet staining of trabecular bone. Subchondral vasculature in undecalcified bone could be visualized by tissue autofluorescence (Top image). The cartilage calcification front was disrupted in severely degenerative tissue and subjacent sclerotic bone showed reduced calcification, but increased vascular density. LSFM ex vivo vascular imaging displayed blood vessels originating from bone marrow terminating in calcified cartilage (Lower image). Vascular invasion into non-decalcified cartilage was not observed in mild or severe OA grades. Vascular density and vessel size appeared both increased in severe compared with mild OA grade.
Conclusion: We provided proof of concept for ECi-based optical 3D ex vivo imaging of human OA osteochondral tissue. This method opens up new avenues for spatial analysis of cells/structures in bone and joint research.