Periodic Reporting for period 4 - PROVEC (Promoting Osteogenesis through Vascular Endothelial Cells)
Periodo di rendicontazione: 2023-08-01 al 2024-01-31
Reduced bone mineral density is an aspect of normal aging but can lead to osteoporosis, a disease characterized by bone weakness, increased fracture risk, loss of mobility, and chronic pain. Accordingly, osteoporosis and other disease conditions involving bone loss represent major health burdens in the European Union. While it widely appreciated that bone development, lifelong homeostasis and regeneration rely on the balanced activity of bone-forming osteoblasts and bone-resorbing osteoclasts, the role of blood vessels of the skeletal system had remained little understood. This was, in part, due to technical limitations associated with the processing, imaging and analysis of the matrix-rich and heavily calcified bone tissue.
Our research in PROVEC has overcome major technical obstacles and has combined high-resolution microscopy, genetic and pharmacological approaches in mice, single cell RNA-sequencing and other powerful methods to gain fundamental insights into the organization and function of the bone vasculature. In particular, PROVEC has uncovered critical aspects of organ-specific and functional specialization of blood vessels in bone. Whereas certain endothelial cells, which form the innermost lining of the vessel wall, regulate the abundance of osteoprogenitor cells and thereby osteogenesis, other parts of the vasculature are critical for hematopoiesis in the bone marrow. Our findings also improve our understanding of the changes affecting the aging skeletal system. This process involves the gradual disappearance of vessels promoting bone formation together with loss of the vessel-associated osteoprogenitor cells and trabecular bone. We find that genetic and pharmacological strategies aiming at blood vessels and osteoprogenitors can overcome some of the bone loss in aging mice, which suggests that such strategies might have potential relevance for the treatment of human diseases affecting the skeletal system. Finally, we have also gained interesting insights into the processes mediating fracture repair and the relevance of different cell populations in this process.
Taken together, PROVEC has systemically identified and characterized endothelial cell subpopulations, their gene expression and functional properties in the healthy, aging, diseased and regenerating skeletal system. We have established that blood vessels are an essential component of the skeletal system and contribute to the regulation of bone growth, homeostasis and aging through molecular interactions and feedback loops with perivascular bone-forming and bone-degrading cells. These findings might be relevant for the development of treatments targeting osteoporosis and other diseases affecting the skeletal system.
Our research in PROVEC has overcome major technical obstacles and has combined high-resolution microscopy, genetic and pharmacological approaches in mice, single cell RNA-sequencing and other powerful methods to gain fundamental insights into the organization and function of the bone vasculature. In particular, PROVEC has uncovered critical aspects of organ-specific and functional specialization of blood vessels in bone. Whereas certain endothelial cells, which form the innermost lining of the vessel wall, regulate the abundance of osteoprogenitor cells and thereby osteogenesis, other parts of the vasculature are critical for hematopoiesis in the bone marrow. Our findings also improve our understanding of the changes affecting the aging skeletal system. This process involves the gradual disappearance of vessels promoting bone formation together with loss of the vessel-associated osteoprogenitor cells and trabecular bone. We find that genetic and pharmacological strategies aiming at blood vessels and osteoprogenitors can overcome some of the bone loss in aging mice, which suggests that such strategies might have potential relevance for the treatment of human diseases affecting the skeletal system. Finally, we have also gained interesting insights into the processes mediating fracture repair and the relevance of different cell populations in this process.
Taken together, PROVEC has systemically identified and characterized endothelial cell subpopulations, their gene expression and functional properties in the healthy, aging, diseased and regenerating skeletal system. We have established that blood vessels are an essential component of the skeletal system and contribute to the regulation of bone growth, homeostasis and aging through molecular interactions and feedback loops with perivascular bone-forming and bone-degrading cells. These findings might be relevant for the development of treatments targeting osteoporosis and other diseases affecting the skeletal system.
In the course of the project, we have generated substantial insights into the heterogeneity, molecular properties and functional specialization of bone endothelial cells and other cell populations in the skeletal system.
Specifically, we have been able to show that the Hippo pathway is an important regulator of angiogenic blood vessel growth in bone and trabecular bone formation by regulating hypoxia-inducible factor signaling (Sivaraj et al., eLife 2020). We also showed that Notch signaling plays important roles in bone angiogenesis and osteogenesis, which can be therapeutically utilized in mice as a preclinical model (Xu, Dinh et al, eLife 2022; Remark et al., Bone Res. 2023). Furthermore, we discovered that vessel-associated bone mesenchymal stromal cells are regionally specialized and that a subset of these cells mediates chondrocyte resorption during developmental bone growth and fracture healing (Sivaraj et al., Cell Reports 2021; Sivaraj et al., Nature Commun. 2022). While blood vessels are coupled to the progenitors of bone-forming cells during fracture healing in long bone, the same is not the case in skull (Bixel et al., Nature Commun., accepted for publication). Skull bone marrow is also functionally different from long bone and is subjected to lifelong expansion (Koh et al., under revision). All these papers utilized single cell RNA-sequencing to analyze the properties of different cell populations in bone, which is also the topic of another earlier publication (Tikhonova et al., Nature 2019). More recently, we have used single cell RNA-sequencing and mouse genetics for the identification of another blood vessel subtype associated with remodeling bone (Mohanakrishnan et al., final revision). Furthermore, we demonstrated that certain blood vessels protect bone and marrow against fibrosis (Sivaraj, Majev et al., Nature CVR, accepted for publication).
In the context of the functional specialization of bone endothelial cells, we showed that a subset of Apelin-expressing cells promotes vessel and bone marrow regeneration after irradiation (Chen et al., Cell Stem Cell 2019). The function of bone marrow also relies of vessel-associated nerve fibers and the neurotransmitter dopamine (Liu et al., Blood 2021; Deng et al., FASEB Journal 2022). Furthermore, we showed that the development of fetal bone marrow relies on signals from arterial endothelial cells (Liu et al., Nature Commun. 2022).
Taken together, research in PROVEC has discovered many fundamental properties of the bone vasculature and its interactions with other cell types with relevance for fracture healing, regeneration, fibrosis and age-related bone loss. These results will form the basis for future preclinical and translational research addressing the function of blood vessels in health and disease.
Specifically, we have been able to show that the Hippo pathway is an important regulator of angiogenic blood vessel growth in bone and trabecular bone formation by regulating hypoxia-inducible factor signaling (Sivaraj et al., eLife 2020). We also showed that Notch signaling plays important roles in bone angiogenesis and osteogenesis, which can be therapeutically utilized in mice as a preclinical model (Xu, Dinh et al, eLife 2022; Remark et al., Bone Res. 2023). Furthermore, we discovered that vessel-associated bone mesenchymal stromal cells are regionally specialized and that a subset of these cells mediates chondrocyte resorption during developmental bone growth and fracture healing (Sivaraj et al., Cell Reports 2021; Sivaraj et al., Nature Commun. 2022). While blood vessels are coupled to the progenitors of bone-forming cells during fracture healing in long bone, the same is not the case in skull (Bixel et al., Nature Commun., accepted for publication). Skull bone marrow is also functionally different from long bone and is subjected to lifelong expansion (Koh et al., under revision). All these papers utilized single cell RNA-sequencing to analyze the properties of different cell populations in bone, which is also the topic of another earlier publication (Tikhonova et al., Nature 2019). More recently, we have used single cell RNA-sequencing and mouse genetics for the identification of another blood vessel subtype associated with remodeling bone (Mohanakrishnan et al., final revision). Furthermore, we demonstrated that certain blood vessels protect bone and marrow against fibrosis (Sivaraj, Majev et al., Nature CVR, accepted for publication).
In the context of the functional specialization of bone endothelial cells, we showed that a subset of Apelin-expressing cells promotes vessel and bone marrow regeneration after irradiation (Chen et al., Cell Stem Cell 2019). The function of bone marrow also relies of vessel-associated nerve fibers and the neurotransmitter dopamine (Liu et al., Blood 2021; Deng et al., FASEB Journal 2022). Furthermore, we showed that the development of fetal bone marrow relies on signals from arterial endothelial cells (Liu et al., Nature Commun. 2022).
Taken together, research in PROVEC has discovered many fundamental properties of the bone vasculature and its interactions with other cell types with relevance for fracture healing, regeneration, fibrosis and age-related bone loss. These results will form the basis for future preclinical and translational research addressing the function of blood vessels in health and disease.
PROVEC will uncover fundamental aspects of the crosstalk between ECs and other cell types in bone. It will explore in preclinical models whether ECs can be used to promote bone formation or halt the loss of bone mineral density. In addition, the project will establish whether responses in bone ECs are required for therapeutic approaches aiming at osteoblasts or osteoclasts. Results obtained in the course of PROVEC could be used in future translational research approaches to investigate the status of bone ECs in human patients. Most importantly, future research could lead to the development of completely new treatment strategies that utilize bone ECs for improved fracture healing or to stabilize/increase bone mineral density.