降钙素基因相关肽对MC3T3-E1成骨细胞氧化损伤的保护作用研究

目的 探讨降钙素基因相关肽（CGRP）对MC3T3-E1成骨细胞氧化损伤的保护作用及其潜在机制。方法1）构建细胞氧化损伤模型，将实验分为4组，H₂O₂组使用含不同浓度（10^-1、10^-2、10^-3、10^-4、10^-5 mol·L^-1）H₂O₂的培养液，CGRP+H₂O₂组使用含不同浓度（10^-6、10^-7、10^-8、10^-9、10^-10 mol·L^-1）CGRP的培养液预处理后再加入含10^-4 mol·L^-1 H₂O₂的培养液，对照组为常规培养液，空白组不接种细胞。培养12、24、36、48 h后，CCK-8法检测细胞增殖活性，筛选最佳建模浓度和CGRP对成骨细胞氧化损伤的最佳保护浓度。2）采用含10^-4 mol·L^-1 H₂O₂（H₂O₂组）、10^-8 mol·L^-1 CGRP（CGRP组）的培养液和10^-8 mol·L^-1 CGRP+10^-4 mol·L^-1 H₂O₂培养液（CGRP+H₂O₂组）、常规培养液（对照组）培养成骨细胞，测定超氧化物歧化酶（SOD）的活性、活性氧（ROS）的含量以及炎症因子肿瘤坏死因子（TNF）-α、白细胞介素（IL）-1β、IL-6的水平。结果 1）在10^-4 mol·L^-1 H²O²时细胞增殖开始出现抑制（P<0.01），并呈浓度和时间依赖性；10^-8 mol·L^-1 CGRP预处理后细胞增殖活性最高，与只加入10^-4 mol·L^-1 H₂O₂有统计学差异（P<0.01）。2）与H²O²组相比，CGRP+H₂O₂组SOD活性升高（P<0.01），TNF-α、IL-1β、IL-6水平降低（P<0.05），ROS含量降低（P<0.01）。结论 H₂O₂会对MC3T3-E1成骨细胞造成氧化损伤，CGRP对MC3T3-E1成骨细胞氧化损伤具有保护作用。

Protective effect of calcitonin gene-related peptide against oxidative damage in MC3T3-E1 osteoblasts

Objective This study aimed to observe the protective effect of calcitonin gene-related peptide (CGRP), as well as its potential mechanism, against oxidative damage in MC3T3-E1 osteoblasts.Methods 1) MC3T3-E1 osteoblasts were treated with different hydrogen peroxide (H2O2) concentrations (10-1, 10-2, 10-3, 10-4, and 10-5mol·L-1) for 12, 24, 36, and 48 h to build an oxidative damage model, to determine cell proliferation activity in each group by using CCK-8 assay, and to deter-mine the optimal modeling concentration. MC3T3-E1 osteoblasts were pretreated for 1 h with different CGRP concentrations (10-6, 10-7, 10-8, 10-9, and 10-10mol·L-1) followed by treatment with H2O2 (10-4 mol·L-1). After 12, 24, 36, and 48 h, the CGRP expression and activity of osteoblasts were detected using the CCK-8 method to determine the optimal CGRP concentration that provides the best protective effect against oxidative damage. 2) Superoxide dismutase (SOD) activity, reactive oxygen species (ROS) content, and the levels of the inflammatory cytokines tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6 of the groups treated with CGRP, H2O2, CGRP+H2O2 were determined.Results 1) Compared with the control group, treatment with 10-4 mol·L-1 H2O2 significantly started to inhibite the proliferation of osteoblasts (P<0.01) in a dose- and time-dependent manner. Compared with 10-4 mol·L-1 H2O2 group, pretreatment with 10-8 mol·L-1 CGRP significantly increased the proliferation of osteoblasts (P<0.01). 2) Compared with H2O2 group, CGRP+H2O2 group significantly increased the SOD activity (P<0.01), ROS content significantly decreased (P<0.01), TNF-α, IL-1β, and IL-6 secretion significantly decreased (P<0.05).Conclusion H2O2 can cause oxidative damage to MC3T3-E1 osteoblasts, whereas CGRP exerts protective effect against oxidative damage in MC3T3-E1 osteoblasts.