Use of Hes1-GFP reporter mice to assess activity of the Hes1 promoter in bone cells under chronic inflammation.
Notch signaling plays a critical role in maintaining bone homeostasis partially by controlling the formation of osteoblasts from mesenchymal stem cells (MSCs). We reported that TNF activates Notch signaling in MSCs which inhibits osteoblast differentiation in TNF transgenic (TNF-Tg) mice, a mouse model of chronic inflammatory arthritis. In the current study, we used Hes1-GFP and Hes1-GFP/TNF-Tg mice to study the distribution and dynamic change of Notch active cells in normal and inflammatory bone loss and mechanisms mediating their enhanced proliferation. We found that Hes1-GFP+ cells are composed of cells expressing mesenchymal, hematopoietic and endothelial surface markers. CD45-/Hes1-GFP+ cells express high levels of mesenchymal markers and form CFU-F and CFU-ALP colonies. Expansion of CFU-F colonies is associated with a rapid increase in Hes1-GFP+ cell numbers and their GFP intensity. The GFP signal is lost when a CFU-F colony differentiates into an ALP+ osteoblast colony. TNF increases the numbers of CD45-/Hes1-GFP+ cells, which are stained negatively for osteoblast marker osteocalcin and localized adjacent to endosteal and trabecular bone surfaces. CD45-/Hes1-GFP+ cells in Hes1-GFP/TNF-Tg mice have increased BrdU incorporation and PDGFRβ levels. TNF increases the number of proliferating Hes1-GFP+ cells, which is prevented by a specific PDGFRβ inhibitor. Notch inhibition blocks TNF-mediated PDGFRβ expression and cell proliferation. Thus, TNF-induced MSC proliferation is mediated by PDGFRβ signal, which works at downstream of Notch. Hes1-GFP mice can be used to assess the activation status of Notch in bone cells.
Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, Rochester, NY 14642, USA.
Hengwei
H
Zhang
Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, Rochester, NY 14642, USA; The Research Center for Bone and Stem Cells, Department of Anatomy, Histology and Embryology, Nanjing Medical University, Nanjing 210029, People's Republic of China.
Wen
W
Sun
Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, Rochester, NY 14642, USA.
Xing
X
Li
Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, Rochester, NY 14642, USA; Department of Medicine, Minzu University of China, Beijing 100081, People's Republic of China.
Mengmeng
M
Wang
Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, Rochester, NY 14642, USA; Center for Musculoskeletal Research, University of Rochester Medical Center, Rochester, NY 14642, USA.
Brendan F
BF
Boyce
Department of Orthopedic Surgery, Duke University School of Medicine, Durham, NC 27710, USA.
Matthew J
MJ
Hilton
Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, Rochester, NY 14642, USA; Center for Musculoskeletal Research, University of Rochester Medical Center, Rochester, NY 14642, USA. Electronic address: Lianping_xing@urmc.rochester.edu.
Lianping
L
Xing
D000818:Animals; D001853:Bone Marrow / Q000378:metabolism; D002452:Cell Count; D002454:Cell Differentiation; D049109:Cell Proliferation / Q000187:drug effects; D002908:Chronic Disease; D003114:Colony-Forming Units Assay; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000378:metabolism*; D006801:Humans; D007249:Inflammation / Q000235:genetics* / Q000473:pathology*; D017493:Leukocyte Common Antigens / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000378:metabolism; D008822:Mice, Transgenic; D010006:Osteoblasts / Q000473:pathology; D011401:Promoter Regions, Genetic; D020797:Receptor, Platelet-Derived Growth Factor beta / Q000378:metabolism; D051880:Receptors, Notch / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D051880:Receptors, Notch; D014409:Tumor Necrosis Factor-alpha; D049452:Green Fluorescent Proteins; D020797:Receptor, Platelet-Derived Growth Factor beta; D017493:Leukocyte Common Antigens
Inflammation; Mesenchymal stem cells; Notch; Osteoporosis; PDGFRβ; Proliferation; TNF