Resting and injury‐induced inflamed periosteum contain multiple macrophage subsets that are located at sites of bone growth and regeneration

Kylie Anne Alexander, Liza‐Jane Raggatt, Susan Millard, Lena Batoon, Andy Chiu‐Ku Wu, Ming‐Kang Chang, David Arthur Hume, Allison Robyn Pettit

Better understanding of bone growth and regeneration mechanisms within periosteal tissues will improve understanding of bone physiology and pathology. Macrophage contributions to bone biology and repair have been established but specific investigation of periosteal macrophages has not been undertaken. We used an immunohistochemistry approach to characterise macrophages in growing murine bone and within activated periosteum induced in a mouse model of bone injury. Osteal tissue macrophages (osteomacs) and resident macrophages were distributed throughout resting periosteum. Tissues were collected from 4 week old mice and osteomacs were observed intimately associated with sites of periosteal diaphyseal and metaphyseal bone dynamics associated with normal growth. This included F4/80(+)Mac-2(-/low) osteomac association with extended tracks of bone formation (modeling) on diphyseal periosteal surfaces. While this recapitulated endosteal osteomac characteristics, there was subtle variance in the morphology and spatial organization of modelling-associated osteomacs, which likely reflects the greater structural complexity of periosteum. We also demonstrated that osteomacs, resident macrophages and inflammatory macrophages (F4/80(+)Mac-2(hi)) were associated with the complex bone dynamics occurring within the periosteum at the metaphyseal corticalization zone. These 3 macrophage subsets were also present within activated native periosteum after bone injury across a 9 day time course that spanned the inflammatory through remodeling bone healing phases. This included osteomac association with foci of endochondral ossification within the activated native periosteum. These observations confirm that osteomacs are key components of both osteal tissues, in spite of salient differences between endosteal and periosteal structure and that multiple macrophage subsets are involved in periosteal bone dynamics.Immunology and Cell Biology accepted article preview online, 24 August 2016. doi:10.1038/icb.2016.74.