The distribution of macrophage subtypes and their relationship to bone morphogenetic protein 2 in calcified aortic valve stenosis

Activation of the osteogenic signaling cascade (OSC) is thought to be involved in aortic valve stenosis. The aim of this study was to clarify the distribution of macrophage (M) subtypes in the calcified aortic valve and to clarify the relationship between osteoblast-like cells (OLC) and OSC activation. Thirty-six cases of calcified aortic valve were set as the calcification group, and six autopsy cases of aortic valve without pathological calcification comprised the noncalcification group. Aortic valve tissues were used in histological studies including single and double immunostaining to identify M subtypes, bone morphogenetic protein 2 (BMP2) and osteopontin, reverse transcription polymerase chain reaction (RT-PCR) for CD206, heme oxygenase-1 (HO-1), and BMP2 mRNAs and in situ RT-PCR for BMP2 mRNA. Ms positive for CD68, CD163, CD206, and HO-1 were significantly higher in the calcification group than in the noncalcification group (P < 0.01). Comparison of the positive cells in each section of the calcification group showed that cells of all M subtypes were found around calcifications. Osteopontin⁺ cells were also observed around calcifications. CD163⁺/CD206⁺ M2 and CD163⁺/HO-1⁺ Mox were significantly higher in the sponge layer in both groups. In double immunofluorescence, CD206⁺ and a portion of HO-1⁺ Ms expressed BMP2, and in RT-PCR, CD206 or HO-1 mRNA was expressed in cases in which BMP2 was expressed. In in situ RT-PCR, expression of BMP2 mRNA was observed around calcifications. This work clarifies the distribution of M subtypes in calcified aortic valves. In addition, the results suggest that CD206⁺ M2 and HO-1⁺ Mox, which express BMP2 in calcified aortic valves, are OLC candidates.