Trabecular Bone Deterioration in col9a1+/− Mice Associated With Enlarged Osteoclasts Adhered to Collagen IX–Deficient Bone

Abstract

Short collagen IX, the exclusive isoform expressed by osteoblasts, is synthesized through alternative transcription of the col9a1 gene. The function of short collagen IX in bone was characterized in col9a1-null mutant mice. Trabecular bone morphometry of lumbar bones and tibias was evaluated by muCT and nondecalcified histology. Osteoblastic and osteoclastic activities were evaluated by PCR- and microarray-based gene expression assays and TRACP-5b and C-terminal telopeptide (CTX) assays, as well as in vitro using bone marrow stromal cells and splenocytes. The effect of col9a1(+/-) mutation on osteoclast morphology was evaluated using RAW264.7-derived osteoclastic cells cultured on the mutant or wildtype calvarial bone substrates. Col9a1 knockout mutation caused little effects on the skeletal development; however, young adult female col9a1(-/-) and col9a1(+/-) mice exhibited significant loss of trabecular bone. The trabecular bone architecture was progressively deteriorated in both male and female heterozygous col9a1(+/-) mice while aging. The aged mutant mice also exhibited signs of thoracic kyphosis and weight loss, resembling the clinical signs of osteoporosis. The col9a1(+/-) osteoblasts synthesized short col9a1 transcripts at decreased rates. Whereas bone formation activities in vitro and in vivo were not affected, the mutant osteoblast expressed the elevated ratio of RANKL/osteoprotegerin. Increased serum TRACP-5b and CTX levels were found in col9a1(+/-) mice, whose bone surface was associated with osteoclastic cells that were abnormally flattened and enlarged. The mutant and wildtype splenocytes underwent similar osteoclastogenesis in vitro; however, RAW264.7-derived osteoclastic cells, when cultured on the col9a1(+/-) calvaria, widely spread over the bone surface and formed large resorption pits. The surface of col9a1(+/-) calvaria was found to lack the typical nanotopography. The mineralized bone matrix deficient of short collagen IX may become susceptible to osteoclastic bone resorption, possibly through a novel non-cell-autonomous mechanism. The data suggest the involvement of bone collagen IX in the pathogenesis of osteoporosis.