Defective alternative pathway of NF-κB causes mild osteopetrosis in mice

Alymphoplasia (aly/aly) mice have been shown to carry a spontaneous recessive point mutation in the gene NF-κB inducing Kinase (NIK). NIK is necessary for processing of NF-κB2 (p100) in NF-κB alternative pathway. Objective: To clarify the importance of alternative pathway in osteoclast biology we began by characterizing the skeletal phenotype of aly/aly mice. Methods: Eight-week old male wild-type (wt), heterozygous (aly/+) and homozygous (aly/aly) mice were used. Radiographical analysis of femora, tibiae and lumbar vertebrae were carried out. Bone mineral density (BMD) was analyzed by peripheral quantitative computed tomography (pQCT). Histomorphometry was performed to measure bone volume parameters. Co-culture of wt osteoblasts with wt, aly/+ and aly/aly bone marrow cells were done in the presence of 1, 25-dihydroxyvitamin D3 and prostaglandin E2 to check the osteoclastogenesis and resorption assay on dentine slices. Results: Wt, aly/aly and aly/+ were normal in appearance and in body weight. Radiographical analysis of aly/aly showed normal shape and size of bones with an increase on bone mass compared to aly/+ mice. The pQCT data revealed a significantly high trabecular and total BMD (p<0.01) in aly/aly mice compared to that of wt and aly/+. Bone-histomorphometric analysis demonstrated significantly higher trabecular volume and trabecular number with reduced trabecular separation. Abundant cartilage remnants in cancellous region were detected in aly/aly mice. Osteoclast surface and number to bone surface were reduced in aly/aly compared to that of aly/+ (p<0.001). Osteoblast surface to bone surface and mineral apposition rate were similar in all groups. A profound defect in osteoclastogenesis and resorption activity was observed in vitro in aly/aly cells compared to wt and aly/+. Conclusion: The aly/aly mice show mild osteopetrosis due to defective osteoclastogenesis and function. These data indicate that alternative pathway is also important in regulation of osteoclast formation and function.