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作 者:金晖[1] 蔡若男[2] 朱紫薇[1] 孙子林[1]
机构地区:[1]东南大学附属中大医院内分泌科,南京210009 [2]徐州市第一人民医院内分泌科,徐州221002
出 处:《中华骨质疏松和骨矿盐疾病杂志》2012年第4期271-275,共5页Chinese Journal Of Osteoporosis And Bone Mineral Research
摘 要:目的探讨糖基化终产物是否通过EMMPRIN/MMPs途径影响Ⅰ型胶原的代谢。方法利用小鼠成骨样细胞(MC3T3E1)和小鼠巨噬细胞(RAW264.7)构建共培养体系。应用AGE-BSA(50mg/L)、EMMPRIN抗体(5mg/L)、AGE-BSA+EMMPRIN抗体分别干预共培养体系24h,用ELISA法检测上清液中Ⅰ型胶原含量。不同浓度AGE-BSA(0、50、100、200、400mg/L)干预共培养细胞24h,收集细胞及上清液,用半定量RT-PCR法检测成骨细胞Ⅰ型胶原mRNA表达,用明胶酶谱法测定上清液中MMP-2和MMP-9的分泌量。结果加入AGE-BSA(50mg/L)干预后上清液中Ⅰ型胶原含量显著降低(P<0.05),EMMPRIN抗体使Ⅰ型胶原含量增加(P<0.05),EMMPRIN抗体+AGE-BSA组使Ⅰ型胶原水平显著增加(P<0.05)。不同浓度AGE-BSA干预共培养体系后,Ⅰ型胶原mRNA表达与对照组相比显著增加(P<0.05),且随AGE-BSA干预浓度增加而增加(P<0.05)。MMP-2、MMP-9分泌水平均较对照组显著增加(P<0.05),并随AGE-BSA干预浓度增加而增加(P<0.05)。结论在体外AGEs可增加成骨细胞Ⅰ型胶原的合成,同时促进Ⅰ型胶原的降解,提示AGEs可能通过打破Ⅰ型胶原合成与降解之间的平衡使降解多于合成,进而降低骨强度。Objective To observe wether the effects of advanced glycation end products (AGEs) on the metabolism of type Ⅰcollagen is through the EMMPRIN/MMPs pathway. Methods To establish the co-cultured system of MC3T3E1 and RAW264. 7. The co-cultured system of MC3T3E1 and RAW264. 7 was intervented with AGE-BSA (50 mg/L) or the antibody of EMMPRIN (5 mg/L) or AGE-BSA + the antibody of EMMPRIN for 24 hours, the level of collagen Ⅰin the supernatant was analyzed by ELISA. Different concentrations (0, 50, 100, 200, 400 mg/L) of AGEBSA was added into the co-cultured system for 24 hours. The mRNA expression of type Ⅰ collagen of osteoblastic cells were detected by semi-quantitative RT-PCR and the excretion of supernatant MMP-2 and MMP-9 was measured by gelatin enzymogram method. Results Compared with α-MEM group, the level of type I collagen from the supernatant significantly decreased in AGE-BSA group ( P 〈 0.05 ), increased in both of the antibody of EMMPRIN group (P 〈 0. 05 ) and AGE-BSA + the antibody of EMMPRIN group. With the 24-hour intervention of different concentrations of AGE-BSA compared with α-MEM group and BSA group, the mRNA level of type I collagen was increased (P 〈 0. 05) in a dose-dependent manner (P 〈 0. 05), the secretion of MMP-2 and MMP-9 were both increased (P 〈 0. 05) in a dose-dependent manner (P 〈 0. 05) in co-cultured system. Conclusion In vitro, AGEs increase the synthesis of type Ⅰ collagen in osteoblastic cells, while promoting the degradation of type Ⅰ collagen. It is indicated that AGEs could break the balance between synthesis and degradation of type Ⅰcollagen, leading to degradation than synthesis, and then to more decrease bone strength.
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