NOTCH inhibits osteoblast formation in inflammatory arthritis via noncanonical NF-κB
Hengwei Zhang, … , Brendan F. Boyce, Lianping Xing
Hengwei Zhang, … , Brendan F. Boyce, Lianping Xing
Published June 2, 2014
Citation Information: J Clin Invest. 2014;124(7):3200-3214. https://doi.org/10.1172/JCI68901.
View: Text | PDF
Research Article Bone biology

NOTCH inhibits osteoblast formation in inflammatory arthritis via noncanonical NF-κB

Abstract

NOTCH-dependent signaling pathways are critical for normal bone remodeling; however, it is unclear if dysfunctional NOTCH activation contributes to inflammation-mediated bone loss, as observed in rheumatoid arthritis (RA) patients. We performed RNA sequencing and pathway analyses in mesenchymal stem cells (MSCs) isolated from transgenic TNF-expressing mice, a model of RA, to identify pathways responsible for decreased osteoblast differentiation. 53 pathways were dysregulated in MSCs from RA mice, among which expression of genes encoding NOTCH pathway members and members of the noncanonical NF-κB pathway were markedly elevated. Administration of NOTCH inhibitors to RA mice prevented bone loss and osteoblast inhibition, and CFU-fibroblasts from RA mice treated with NOTCH inhibitors formed more new bone in recipient mice with tibial defects. Overexpression of the noncanonical NF-κB subunit p52 and RELB in a murine pluripotent stem cell line increased NOTCH intracellular domain–dependent (NICD-dependent) activation of an RBPjκ reporter and levels of the transcription factor HES1. TNF promoted p52/RELB binding to NICD, which enhanced binding at the RBPjκ site within the Hes1 promoter. Furthermore, MSC-enriched cells from RA patients exhibited elevated levels of HES1, p52, and RELB. Together, these data indicate that persistent NOTCH activation in MSCs contributes to decreased osteoblast differentiation associated with RA and suggest that NOTCH inhibitors could prevent inflammation-mediated bone loss.

Authors

Hengwei Zhang, Matthew J. Hilton, Jennifer H. Anolik, Stephen L. Welle, Chen Zhao, Zhenqiang Yao, Xing Li, Zhiyu Wang, Brendan F. Boyce, Lianping Xing

×

Figure 2

Short-term DAPT treatment revised decreased osteoblast differentiation of MSCs in TNF-Tg mice.

Options: View larger image (or click on image) Download as PowerPoint
Short-term DAPT treatment revised decreased osteoblast differentiation o...
(A–C) TNF-Tg mice and WT littermates were gavaged with DAPT (5 mg/kg each time) or vehicle daily for 4 days. The inhibitory effect of short-term DAPT treatment on NOTCH activation (Hes1 mRNA) was confirmed in the popliteal lymph nodes (A; positive control) and in CD45– MSC-enriched cells (B) by qPCR. (C) Representative images and number of CFU-ALP+ colonies in BM stromal cells. (D and E) CFU colony cells from vehicle- or DAPT-treated TNF-Tg mice were implanted to bone matrix in tibial cortical defects of SCID mice. Mice were sacrificed 6 weeks after surgery, and volume of new bone formed in the defects, relative to total defect volume (BV/TV), was measured by μCT (D), followed by histomorphometric analysis of the area of newly formed trabecular bone observed in decalcified H&E-stained bone sections (E). n = 8 per group. Scale bars: 1 mm (broken); 100 mm (solid). (F) CFU cells generated from BM stromal cells of Rosa26-LacZ mice were implanted to tibial cortical defects for 6 weeks as in D. Frozen sections were stained for LacZ enzymatic activity (blue) and counterstained with nuclear fast red (pink). Percent bone surface area covered by donor cells (LacZ+; red arrows) over total bone surface within the area of bone defect (green line) was assessed. n = 5 per group. Scale bars: 1 μm (broken); 100 μm (solid). *P < 0.05 vs. WT; #P < 0.05 vs. vehicle.