Inactivation of Patched1 in Murine Chondrocytes Causes Spinal Fusion Without Inflammation

Abstract

Objective. During development of the vertebrate skeleton, chondrocytes form a cartilage template that is gradually replaced by bone. Hormones of the Hedgehog (HH) family have been implicated in the ossification process, but their exact relationship to normal or pathogenic bone formation is unclear. This study was undertaken to establish a genetic tool that allows the discrete inactivation of genes in spinal chondrocytes, and to investigate in vivo how chondrocyte-specific ablation of the inhibitory HH receptor Patched 1 (Ptch1) affects skeleton integrity.Methods. A Cre-deleter mouse strain, mb1-Cre, for selective gene recombination in spinal chondrocytes was identified by in situ hybridization and histologic analysis. The mb1-Cre ؉/؊ animals were crossed with mice that harbor a loxP-flanked Ptch1 gene (Ptch1 flox/flox ) to abrogate the inhibition of the HH signaling pathway in chondrocytes. The skeletal integrity of F1 mice was characterized by high-resolution flat-panel-based volume computed tomography and histologic staining procedures.Results. During the first weeks after birth, all mb1-Cre ؉/؊ /Ptch1 flox/flox mice developed progressive spinal fusion with malformation of the vertebrae. This phenotype was caused by aberrant chondrocyte proliferation in the intervertebral discs that blocked endochondral ossification. Importantly, the disease pattern occurred in an inflammation-independent manner. Conclusion.Our findings indicate that chronic activation of the HH signal pathway in spinal chondrocytes can trigger an ankylosing spine morphology without immune cell contributions. Hence, the destruction of cartilage and loss of axial joint integrity can result from chondrocyte-intrinsic defects of monogenic origin.Development of the axial and appendicular skeleton of vertebrates is initiated by local condensation of mesenchymal cells and their differentiation into chondrocytes, which form cartilage anlagen (1,2). Proper ossification of the anlagen is associated with a complex chondrocyte maturation program (3). Small, highly proliferating chondrocytes produce extracellular matrix proteins that comprise types II, IX, and XI collagen as well as the proteoglycan aggrecan.Indian hedgehog (IHH), a member of the hedgehog (HH) family of secreted morphogens, and parathyroid hormone-related protein (PTHrP) have been reported to maintain chondrocytes in a proliferative state (4-7). Other growth factors, such as thyroid hormone or bone morphogenic proteins, antagonize the actions of IHH and PTHrP, thereby promoting cell cycle exit. This is followed by terminal differentiation of chondrocytes into hypertrophic cells, which secrete type X collagen (ColX) and ultimately die by apoptosis. Following its calcification, the cartilage matrix provides a template for the balanced action of hematopoietic osteoclasts and mesenchyme-derived osteoblasts to finalize bone formation. The long bones and the vertebrae of the spinal column are connected by synovial joints and intervertebral discs, respectively.Chondrogenesis and endochondral oss...

DKK-1 Is Underexpressed in Mesenchymal Stem Cells from Patients with Ankylosing Spondylitis and Further Downregulated by IL-17

“…Mechanisms of new bone formation in the context of AS are not well understood. Nonetheless, a considerable amount of experimental evidence indicates that developmental pathways, such as the Wnt, hedgehog and bone morphogenetic protein pathways are crucially involved [ 2 , 3 , 4 , 5 , 6 , 7 ]. Most data implicate the Wnt pathway as a key driver of osteoblastogenesis/new bone formation [ 8 , 9 ].…”

Anti-TNFα treatment decreases the previously increased serum Indian Hedgehog levels in patients with ankylosing spondylitis and affects the expression of functional Hedgehog pathway target genes

“…(10) Moreover, a mouse model that was genetically manipulated in order to exhibit enhanced HH signaling in spinal chondrocytes showed spinal fusion indicating that chronic activation of the HH pathway can trigger an ankylosing phenotype. (11) There are 3 ligands able to stimulate the HH pathway called Indian, Sonic and Desert Hedgehog but the main ligand in the skeleton is Indian Hedghog (Ihh). Ihh is mainly produced by hypertrophic chondrocytes and favors the process of endochondral ossification (12); on the other hand parathyroid hormone related peptide (PTHrP) is an inhibitor of this process.…”

Development of the Synarcual in the Elephant Sharks (Holocephali; Chondrichthyes): Implications for Vertebral Formation and Fusion

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