Effects of advanced glycated end–products on secretion of extracellular matrix metalloproteinase inducer and activity of matrix metalloproteinase–2 in mouse osteoblasts

Objective To study the effect of advanced glycated end products (AGEs) on the secretion of extracellular matrix metalloproteinase inducer (EMMPRIN) and the activity of matrix metalloproteinase–2 (MMP–2) in cultured mouse embryo/fetus calvaria osteoldasts (MC3T3–E1). Methods The AGEs–BSA was prepared by incubating bovine serum albumin (BSA) with glucose. The cultured MC3T3–E1 was added with AGEs–BSA (50, 100, 200, and 400 mg/L) for 24 h or 200 mg/L AGEs–BSA for 12, 24, and 48 h, respectively, taking DMEM and BSA as negative control. The concentration of EMMPRIN in the supernatant was quantified by ELISA. The activity of MMP–2 in MC3T3–E1 was determined by gelatin enzymogram method. MC3T3–E1 was cultured in the presence of DMEM and AGEs–BSA (50 mg/L) with or without Anti–EMMPRIN antibody. The activity of MMP–2 in MC3T3–E1 was determined by gelatin enzymogram method. All statistical analyses were carried out with the SPSS 13.0. Statistical analysis was done by one–way analysis of variance (ANOVA). Results In different dose of AGEs group, the concentration of EMMPRIN in the supernatant ((7.34±0.11), (10.86±0.07), (14.48±0.14), and (15.43±0.23) μg/L) was higher than BSA group (q was 3.111, 3.090, 2.921, and 4.387; P<0.05) and the activity of MMP–2 ((225.12±5.01), (305.83±5.21), (363.04±8.04), and (410.63±16.84)INT·mm2)was significantly increased than BSA group( q was 3.109, 3.545, 5.912, and 5.895; P<0.05). In different time of AGEs group, the concentration of EMMPRIN in the supernatant ((12.41±0.02), (17.88±0.35), and (18.88±0.36) μg/L) was higher than BSA group (q was 5.522, and 7.462, 7.323, P<0.05) and the activity of MMP–2 ((222.18±14.53), and (246.53±5.96) INT·mm2) was increased compared to BSA group (q was 4.159, and 4.321; P<0.05). The activity of MMP–2 was significantly decreased in anti–EMMPRIN antibody–blocking group compared to the control groups ((543.21±67.90) and (867.95±113.46) INT·mm2, q=6.354, P<0.05) and 50 mg/L AGEs group ((127.63±11.36) and (160.76±17.45) INT·mm2, q=7.742, P<0.05). Conclusions The AGEs could stimulate the expression of EMMPRIN and the activity of MMP–2 in cultured MC3T3–E1, which may be partially inhibited by anti–EMMPRIN antibody. These findings suggest that EMMPRIN might mediate the role of AGEs in the development of osteoporosis by MMP–2. Key words: Glycosylation end products, advanced; Osteoblast; Osteoporosis; Matrix metalloproteinases