新型含Mn生物陶瓷粉体减轻软骨细胞的氧化应激损伤  

Alleviation of oxidative stress damage in chondrocytes by a new Mn-containing bioceramic powder

作　　者：张梓宁 邓荣辉 余家阔[1,2] Zhang Zining;Deng Ronghui;Yu Jiakuo(Department of Sports Medicine,Peking University Third Hospital,Beijing 100191,China;Department of Orthopedic and Sports Medicine Center,Beijing Tsinghua Changgung Hospital Medical Center,Medical Center of Tsinghua University,Beijing 102218,China)

机构地区：[1]北京大学第三医院运动医学科,北京市100191 [2]北京清华长庚医院骨科与运动医学中心,清华大学医学中心,北京市102218

出　　处：《中国组织工程研究》2025年第16期3335-3342,共8页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金-北京市自然科学基金区域新联合发展基金项目(U22A20283),项目负责人:余家阔。

摘　　要：背景:Mn可参与到多种生物体内氧化还原反应中,比如作为超氧化物歧化酶2中的金属辅助基团发挥辅助清除活性氧的作用,因此近些年开发含Mn的新型抗氧化应激材料成为研究的热点。目的:探讨含Mn生物陶瓷粉体通过降低活性氧途径对软骨细胞氧化应激损伤的保护作用。方法:采用熔盐法制备含Mn生物陶瓷粉体。分离培养小鼠原代软骨细胞。在H_(2)O_(2)溶液中分别加入含0,0.15,0.30 mg/mL Mn生物陶瓷粉体,避光孵育后检测H_(2)O_(2)清除率。将第2-4代软骨细胞分别与含不同质量浓度(0,0.15,0.30 mg/mL)含Mn生物陶瓷粉体的完全培养基共培养,采用细胞活死染色检测细胞活性。将第2-4代软骨细胞(或软骨组织)分4组干预:空白对照组加入完全培养基培养,H_(2)O_(2)组加入用含H_(2)O_(2)的完全培养基培养,H_(2)O_(2)+低质量浓度Mn粉体组加入含H_(2)O_(2)+0.15 mg/mL含Mn生物陶瓷粉体的完全培养基培养,H_(2)O_(2)+高质量浓度Mn粉体组加入含H_(2)O_(2)+0.30 mg/mL含Mn生物陶瓷粉体的完全培养基,采用CCK-8法检测软骨细胞活力,2,7-二氯荧光素二乙酸酯探针检测软骨细胞活性氧生成,qRT-PCR检测软骨细胞相关因子表达,甲苯胺蓝染色检测软骨组织结构与功能。结果与结论:①两种剂量的含Mn生物陶瓷粉体均可以于体外显著清除H_(2)O_(2),并且具有质量浓度依赖性;细胞活死染色结果显示,0.15 mg/mL含Mn生物陶瓷粉体具有软骨细胞安全性,0.30 mg/mL含Mn生物陶瓷粉体存在软骨细胞毒性;②CCK-8检测结果显示,两种质量浓度的含Mn生物陶瓷粉体均可以显著减轻H_(2)O_(2)对软骨细胞活力的抑制作用,并可抑制H_(2)O_(2)诱导的软骨细胞活性氧生成,并且具有质量浓度依赖性;两种质量浓度的含Mn生物陶瓷粉体均可逆转H_(2)O_(2)诱导的软骨细胞血小板反应蛋白解整合素金属肽酶5 mRNA表达升高与蛋白聚糖mRNA表达降低;③甲苯胺蓝染色结果显�BACKGROUND:Mn can participate in oxidation-reduction reactions in various organisms.For example,as a metal-assisted group in superoxide dismutase 2,Mn plays a role in helping to remove reactive oxygen species.Therefore,the development of novel anti-oxidative stress materials containing Mn has become a research focus in recent years.OBJECTIVE:To investigate the protective effect of Mn bioceramic powder material on oxidative stress damage to chondrocytes by reducing the reactive oxygen species pathway.METHODS:Bioceramic powders containing Mn were prepared by molten salt method.Primary mouse chondrocytes were isolated and cultured.Bioceramic powder containing 0,0.15,and 0.30 mg/mL of Mn was added into H_(2)O_(2)solution.The H_(2)O_(2)clearance rate was detected after incubation without light.The passage 2 to passage 4 chondrocytes were co-cultured with complete media containing Mn-containing bioceramic powder with different mass concentrations(0,0.15,and 0.30 mg/mL).Cell viability was detected by cell live/dead staining.The passage 2-4 chondrocytes(or cartilage tissue)were divided into four groups for intervention:Complete culture medium was added to the blank control group.The H_(2)O_(2)group was added and cultured with complete medium containing H_(2)O_(2).H_(2)O_(2)+low mass concentration Mn powder group was cultured by adding H_(2)O_(2)+0.15 mg/mL Mn-containing bioceramic powder.The complete medium containing H_(2)O_(2)+0.30 mg/mL Mn-containing bioceramic powder was added to the H_(2)O_(2)+high mass concentration Mn powder group.Viability of chondrocytes was detected by CCK-8 assay.Generation of reactive oxygen species of chondrocytes was detected by 2,7-dichlorofluorescein diacetate probe.Expression of chondrocyte-related factors was detected by qRT-PCR.The tissue structure and function of cartilage were detected by toluidine blue staining.RESULTS AND CONCLUSION:(1)Both doses of Mn-containing bioceramic powders could significantly remove H_(2)O_(2)in vitro,and they were concentration dependent.The results of ce

关 键 词：生物陶瓷 Mn粉体材料 软骨损伤 氧化应激 活性氧 骨关节炎 细胞外基质 生物安全性 

分 类 号：R459.9[医药卫生—治疗学] R318.08[医药卫生—临床医学] R684.3