Osteoblast CFTR Inactivation Reduces Differentiation and Osteoprotegerin Expression in a Mouse Model of Cystic Fibrosis-Related Bone Disease

Abstract

Low bone mass and increased fracture risk are recognized complications of cystic fibrosis (CF). CF-related bone disease (CFBD) is characterized by uncoupled bone turnover—impaired osteoblastic bone formation and enhanced osteoclastic bone resorption. Intestinal malabsorption, vitamin D deficiency and inflammatory cytokines contribute to CFBD. However, epidemiological investigations and animal models also support a direct causal link between inactivation of skeletal cystic fibrosis transmembrane regulator (CFTR), the gene that when mutated causes CF, and CFBD. The objective of this study was to examine the direct actions of CFTR on bone. Expression analyses revealed that CFTR mRNA and protein were expressed in murine osteoblasts, but not in osteoclasts. Functional studies were then performed to investigate the direct actions of CFTR on osteoblasts using a CFTR knockout (Cftr−/−) mouse model. In the murine calvarial organ culture assay, Cftr−/− calvariae displayed significantly less bone formation and osteoblast numbers than calvariae harvested from wildtype (Cftr+/+) littermates. CFTR inactivation also reduced alkaline phosphatase expression in cultured murine calvarial osteoblasts. Although CFTR was not expressed in murine osteoclasts, significantly more osteoclasts formed in Cftr−/− compared to Cftr+/+ bone marrow cultures. Indirect regulation of osteoclastogenesis by the osteoblast through RANK/RANKL/OPG signaling was next examined. Although no difference in receptor activator of NF-κB ligand (Rankl) mRNA was detected, significantly less osteoprotegerin (Opg) was expressed in Cftr−/− compared to Cftr+/+ osteoblasts. Together, the Rankl:Opg ratio was significantly higher in Cftr−/− murine calvarial osteoblasts contributing to a higher osteoclastogenesis potential. The combined findings of reduced osteoblast differentiation and lower Opg expression suggested a possible defect in canonical Wnt signaling. In fact, Wnt3a and PTH-stimulated canonical Wnt signaling was defective in Cftr−/− murine calvarial osteoblasts. These results support that genetic inactivation of CFTR in osteoblasts contributes to low bone mass and that targeting osteoblasts may represent an effective strategy to treat CFBD.

Anoctamin-6 Controls Bone Mineralization by Activating the Calcium Transporter NCX1

“…Cystic fibrosis patients carrying the common F508del mutation in the CFTR gene and mice carrying an inactivated CFTR allele suffer from bone mineralization defects leading to osteoporosis and osteopenia (37)(38)(39)(40). The mechanisms of how CFTR operates in osteoblasts are only poorly understood.…”

Epidermal CFTR Suppresses MAPK/NF-κB to Promote Cutaneous Wound Healing

“…Of the identified 1,900 mutations of CFTR in humans, deletion of the phenylalanine at position 508 (DF508) is the most frequent mutation found in CF patients, which results in a folding defect and endoplasmic reticulum associated degradation of CFTR [17]. It is being recently recognized that apart from transporting anions, CFTR is involved in other cellular functions, including inflammation [6,18], proliferation [19][20][21], and differentiation [22][23][24] in bronchial epithelia, granulosa cells, enterocytes, and osteoblasts. Several signaling molecules associated with CFTR have been reported, including MAPK and NF-κB [6,18,[25][26][27][28].…”

Rare causes of osteoporosis

“…Mutation of the CFTR gene itself may have a direct role in the pathogenesis of CF-related bone disease (17). Some studies have indicated that the loss of CFTR activity in osteoblasts decreases the secretion of osteoprotegerin (OPG) resulting in accentuated inflammationdriven bone resorption (21). The evidence available about the use of bisphosphonates for osteoporosis in individuals with CF is limited.…”