Histochemical assessment of accelerated bone remodeling and reduced mineralization in Il-6 deficient mice

Interleukin-6 (IL-6) contributes to the regulation of functions in various tissues and organs. Even though IL-6 has been reported to modulate bone metabolism in previous studies, this finding is controversial. This study aims to evaluate the possible involvement of IL-6 in bone metabolism by examining the histological activity of osteoblasts and osteoclasts in the femora of Il-6 deficient (Il-6−/−) mice. Eight-week-old male Il-6−/− mice and their wild-type littermates were fixed with a paraformaldehyde solution, and their femora were extracted for micro-CT analysis, immunohistochemistry, and real-time PCR analysis. Il-6−/− femora showed an increased bone volume/tissue volume (TV) but a reduced bone mineral density compared with the wild-type. Furthermore, the tissue-nonspecific alkaline phosphatase positive area/TV ratio, the expression of Runx2, Osterix, and Rankl, and the number of tartrate-resistant acid phosphatase-positive osteoclasts were all increased in the Il-6−/− mice. A considerable number of unmineralized areas within the bone matrix and abundant sclerostin-reactive osteocytes were observed in Il-6−/− femoral metaphyses but not in the wild-type. Interestingly, the gene expression of Cd206 was elevated in Il-6−/− femora, and many F4/80-positive macrophages/monocytes and CD206-immunoreactive macrophages in the primary trabeculae had migrated closer to the growth plate, where intense RANKL immunoreactivity was detected. These results suggest that, in an IL-6-deficient state, CD206-positive macrophages may differentiate into osteoclasts when in contact with RANKL-reactive osteoblastic cells. In a state of IL-6 deficiency, the population and cell activities of osteoblast, osteoclasts, and macrophages seemed to be facilitated, except for the reduced mineralization in bone.