RESUMO
BACKGROUND: Hematopoietic stem cells (HSCs) reside in the bone marrow (BM) niche, which includes bone-forming and bone-resorbing cells, i.e., osteoblasts (OBs) and osteoclasts (OCs). OBs originate from mesenchymal progenitors, while OCs are derived from HSCs. Self-renewal, proliferation and differentiation of HSCs are under the control of regulatory signals generated by OBs and OCs within the BM niche. Consequently, OBs and OCs control both bone physiology and hematopoiesis. Since the human developmental and bone marrow failure genetic syndrome fanconi anemia (FA) presents with skeletal abnormalities, osteoporosis and HSC impairment, we wanted to test the hypothesis that the main pathological abnormalities of FA could be related to a defect in OC physiology and/or in bone homeostasis. RESULTS: We revealed here that the intrinsic differentiation of OCs from a Fanca-/- mouse is impaired in vitro due to overactivation of the p53-p21 axis and defects in NF-kB signaling. The OC differentiation abnormalities observed in vitro were rescued by treating Fanca-/- cells with the p53 inhibitor pifithrin-α, by treatment with the proinflammatory cytokine TNFα or by coculturing them with Fanca-proficient or Fanca-deficient osteoblastic cells. CONCLUSIONS: Overall, our results highlight an unappreciated role of Fanca in OC differentiation that is potentially circumvented in vivo by the presence of OBs and TNFα in the BM niche.
RESUMO
BACKGROUND: The zebrafish is widely used in research due in part to its readily manipulable genome. Zebrafish models of spinal deformities including scoliosis were developed recently. However, the methods used to assess the spine in these models vary across studies. The primary objective of this study was to investigate the feasibility and modalities of local and regional spine structure evaluation by micro-CT in the normal zebrafish. The secondary objectives were to assess the feasibility of spinal angle measurements in normal zebrafish subjected to external stresses designed to mimic spinal deformities, to determine normal angle values in the coronal and sagittal planes, and to detail the micro-CT features of the zebrafish spine. HYPOTHESIS: Micro-CT is an effective and reproducible tool for determining orthopaedic parameters to characterise the zebrafish spine. MATERIAL AND METHODS: Two observers conducted preliminary analyses on 15 zebrafish including 12 adults (aged 18 months) and 3 juveniles (aged 12 weeks). For the analyses, 6 of the animals were placed in an artificial position to mimic a scoliosis spinal deformity. Micro-CT (Quantum FX Caliper™) was used with 59µm resolution and a 30-mm field of view. Image processing was with RadiAnt DICOM Viewer™ software. RESULTS: We defined several assessment planes on the 3D micro-CT reconstructions to measure orthopaedic parameters in the sagittal plane (thoracic and maximal kyphotic curves with their apices, length of the various spinal segments, and sagittal index) and coronal plane (Cobb angles, apices, end-vertebrae, coronal alignment, and side of the convexity). Mean thoracic kyphosis was 20.5°±5.0° in the adults and 8.7° in the juveniles. No curvature was apparent in the coronal plane in the zebrafish left in the neutral position. In the zebrafish with artificially induced curves, micro-CT was effective in determining the Cobb angles and apical vertebrae. DISCUSSION: This work defines a standardised micro-CT method for assessing the zebrafish spine. In addition, spinal parameter values that can be considered normal were determined, namely, less than 30° of thoracic kyphosis in the sagittal plane and less than 10° in the coronal plane. Our method was effective in assessing induced spinal deformities on micro-CT reconstructions. We hope it will prove of value in future studies of the zebrafish model. LEVEL OF EVIDENCE: IV.
Assuntos
Escoliose/diagnóstico , Coluna Vertebral/diagnóstico por imagem , Microtomografia por Raio-X/métodos , Animais , Modelos Animais de Doenças , Reprodutibilidade dos Testes , Peixe-ZebraRESUMO
Idiopathic pulmonary fibrosis (IPF) is a devastating lung disease characterized by fibroblast proliferation, extracellular matrix deposition, destruction of pulmonary alveolar architecture and vascular remodeling. Apart pirfenidone or nintendanib that only slow down the fibrotic process, there is no curative treatment other than lung transplantation. Because cell therapy approaches have been proposed in IPF, we hypothesized that injection of endothelial colony-forming cells (ECFCs), the vasculogenic subtype of endothelial progenitor cells, could modulate fibrosis in a Nude mouse model of bleomycin induced-pulmonary fibrosis. Mice were injected with ECFCs isolated from cord blood and from peripheral blood of adult IPF patients at two time-points: during the development of the fibrosis or once the fibrosis was constituted. We assessed morbidity, weight variation, collagen deposition, lung imaging by microCT, Fulton score and microvascular density. Neither ECFCs isolated from cord blood nor from IPF patients were able to modulate fibrosis or vascular density during fibrogenesis or when fibrosis was constituted. These findings indicate that human ECFCs do not promote an adaptive regenerative response in the lung upon fibrosis or angiogenic process in the setting of bleomycin-induced pulmonary fibrosis in Nude mice.