AIM: Early calcification of atherosclerotic plaques are colocalized with macrophage and high mobility group box 1( HMGB1),a cytokine associated with biomineralizing process under physiological and pathological conditi...AIM: Early calcification of atherosclerotic plaques are colocalized with macrophage and high mobility group box 1( HMGB1),a cytokine associated with biomineralizing process under physiological and pathological conditions. Our study aims to evaluate whether HMGB1 induces ectopic mineralization via promoting the secretion of matrix vesicles( MVs) from macrophages. METHODS: HMGB1 was added to the medium of macrophages,the secretion of MVs in the supernatant was tested by flow cytometry analysis. The mineral deposition in calcifying medium was detected by Alizarin Red staining and von Kossa staining. Transmission electron microscopy showed the formation of hydroxyapatite crystals in MVs. Then we subcutaneous injection into mice with MVs to induce regional mineralization. RESULTS: HMGB1 significantly promoted secretion of MVs from macrophages as raveled by flow cytometry analysis. TNAP activity,considered as a marker of MVs maturation,was higher in HMGB1-induced MVs compared to the control-MVs. HMGB1-MVs also led to mineral deposition in an in vitro MVs-collagen mineralization model. Subcutaneous injection into mice with MVs derived from HMGB1-treated cells showed a greater potential to initiate regional mineralization. Mechanistic experiments revealed that HMGB1 activated neutral sphingomyelinase 2( n SMase2) that involved the receptor for advanced glycation end products( RAGE) and p38MAPK( upstream of n SMase2). Inhibition of n SMase2 with GW4869 or p38 MAPK with SB-239063 prevented MVs secretion and mineral deposition. CONCLUSIONS: HMGB1 induces MVs secretion from macrophages at least in part,via the RAGE / p38 MAPK /n SMase2 signaling pathway. Our findings thus reveal a novel mechanism by which HMGB1 may participated in the early calcification of atherosclerotic plaques.展开更多
基金Supported by the grants from the National Natural Science Foundation of China(No.81270362No.81470561)State Project For Essential Drug Research and Development(No.2013ZX09103003-001)
文摘AIM: Early calcification of atherosclerotic plaques are colocalized with macrophage and high mobility group box 1( HMGB1),a cytokine associated with biomineralizing process under physiological and pathological conditions. Our study aims to evaluate whether HMGB1 induces ectopic mineralization via promoting the secretion of matrix vesicles( MVs) from macrophages. METHODS: HMGB1 was added to the medium of macrophages,the secretion of MVs in the supernatant was tested by flow cytometry analysis. The mineral deposition in calcifying medium was detected by Alizarin Red staining and von Kossa staining. Transmission electron microscopy showed the formation of hydroxyapatite crystals in MVs. Then we subcutaneous injection into mice with MVs to induce regional mineralization. RESULTS: HMGB1 significantly promoted secretion of MVs from macrophages as raveled by flow cytometry analysis. TNAP activity,considered as a marker of MVs maturation,was higher in HMGB1-induced MVs compared to the control-MVs. HMGB1-MVs also led to mineral deposition in an in vitro MVs-collagen mineralization model. Subcutaneous injection into mice with MVs derived from HMGB1-treated cells showed a greater potential to initiate regional mineralization. Mechanistic experiments revealed that HMGB1 activated neutral sphingomyelinase 2( n SMase2) that involved the receptor for advanced glycation end products( RAGE) and p38MAPK( upstream of n SMase2). Inhibition of n SMase2 with GW4869 or p38 MAPK with SB-239063 prevented MVs secretion and mineral deposition. CONCLUSIONS: HMGB1 induces MVs secretion from macrophages at least in part,via the RAGE / p38 MAPK /n SMase2 signaling pathway. Our findings thus reveal a novel mechanism by which HMGB1 may participated in the early calcification of atherosclerotic plaques.
