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铁蓄积通过还原型烟酰胺嘌呤二核苷酸磷酸氧化酶4激活活性氧自由基抑制间充质干细胞与体内成骨功能

Iron accumulation induces reactive oxide species to inhibits mesenchymal stem cells and osteogenesis via nicotinamide adenine dinucleotide phosphate reduced oxidase 4 activation

摘要:

目的 研究铁蓄积对间充质干细胞(MSCs)影响,探索铁蓄积在MSCs层面对成骨功能抑制的可能机制.方法 体内实验,将8周龄C57雄性小鼠,分为对照组(Ctrl)和实验组(FAC),实验组腹腔注射枸橼酸铁铵(FAC)每周0.1 g/kg,8周后两组均检测:血清铁蛋白(FER)、成骨标志物(P1NP)、股骨远端骨小梁三维形态重建和空间结构参数、MSCs占骨髓细胞比例.体外实验,取C57雄性小鼠骨髓分离培养MSCs细胞,分为细胞对照组和细胞实验组,细胞实验组加FAC干预24h,两组均行活性氧自由基(ROS)水平、还原型烟酰胺嘌呤二核苷酸磷酸氧化酶4(NOX4)表达水平检测.结果 FAC组血清铁蛋白[(14.72±0.97)μg/L]高于对照组[(4.92±0.82) μg/L],P=0.002;FAC组骨髓MSCs占骨髓比例[(2.08±0.98)%]低于对照组[(3.98±0.82)%],P=0.004;FAC组细胞ROS的Mean值(56.26 ±8.36)高于对照组(257.65±12.69),P=0.000;FAC组细胞NOX4蛋白表达上升;FAC组P1NP[(4.81±0.51)ng/ml]低于对照组[(10.06±0.96) ng/ml],P=0.001.micro-CT检测的FAC组骨密度[(53.12±9.86) mg/mm3]低于对照组[(103.76±7.92) mg/mm3],P =0.001;FAC组骨小梁空间结构参数显示:FAC组骨体积分数(BV/TV):[(11.23±1.86)%]低于对照组[(18.63±2.02)%],P=0.002;FAC组骨小梁厚度(Tb.Th):[(0.057±0.018) mm]低于对照组[(0.083±0.006) mm],P=0.003;FAC组骨小梁数目(Tb.N):[(0.92±0.22) N/mm]低于对照组[(1.25 ±0.18) N/mm],P=0.004.结论 铁蓄积后,成骨指标降低可能与MSCs受抑制有关,MSCs受抑制可能与铁蓄积诱导NOX4激活ROS相关.

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abstracts:

Objective To study iron accumulation affect on mesenchymal stem cells (MSCs),and to explore iron accumulation in MSCs level pathogenesis.Methods In vivo,6-8 weeks C57 male mice were devived into control group and experimental group,the experimental group was intraperitoneally injected with ferric ammonium citrate (FAC) of 0.1 g/kg/week intervention in eight weeks,the two groups were tested:serum ferritin (FER),osteogenesis markers procollagen type 1 amino-terminal propeptide (P1NP),distal femur trabecular bone reconstruction of three dimensional form and spatial structure parameter,MSCs accounted for the proportion of bone marrow cells.In vitro,6-8 weeks male C57 mice bone marrow tissue were separation,MSCs were cultivated and devided after reaching a certain number in the control group and experimental group.After 24 h FAC intervention in experimental group,two groups were tested:reactive oxygen species (ROS),nicotinamide adenine dinucleotide phosphate reduced oxidase 4 (NOX4) expression level.Statistical analyses were performed.Results FAC group serum FER [(14.72 ±0.97) μg/L] was elevated compared to control group [(4.92 ± 0.82) μg/L],P =0.002.FAC group bone marrow MSCs [(2.08 ± 0.98) %] was reduced compared to control group [(3.98 ± 0.82%)%],P =0.004.FAC group MSCs ROS mean value (56.26 ±8.36) was exceed to control group (257.65 ± 12.69),P =0.000;MSCs NOX4 cell protein was increased compared with the control group;Osteogenesis index P1NP in FAC group [(4.81 ±0.51) ng/ml] was decreased compared to control group [(10.06 ±0.96) ng/ml],P=0.001.The micro-CT detection of bone mineral density in FAC group [(53.12 ± 9.86) mg/mm3] was decreased compared to control group [(103.76 ± 7.92) mg/mm3].FAC group spatial structure parameter showed bone volume/ tissue volume (BV/TV):[(11.23 ± 1.86)%] was decreased compared to control group [(18.63 ±2.02)%],P =0.002;FAC group trabecular thickness (Tb.Th):[(0.057± 0.018)mm] was decreased to control group [(0.083± 0.006) mm],P=0.003;FAC group trabecular number (Tb.N):[(0.92±0.22) N/mm] was decreased compared to control group [(1.25 ± 0.18) N/mm],P =0.004.Conclusion The osteogenetic activity is restrained after iron accumulation may be associated with increased MSCs ROS,MSCs ROS increased may be related to iron accumulation induced NOX4 activation.

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作者: 袁晔 [1] 费蓓蓓 [2] 俞晨 [1] 徐非 [3] 王亮 [1] 杨帆 [4] 张辉 [1] 李光飞 [1] 张志芳 [5] 高焱 [1] 刘功稳 [1] 徐又佳 [1]
期刊: 《中华实验外科杂志》2018年35卷11期 1991-1994页 ISTICPKU
栏目名称: 骨外科
DOI: 10.3760/cma.j.issn.1001-9030.2018.11.002
发布时间: 2018-11-30
基金项目:
国家自然科学基金 江苏省科技厅临床医学技术专项 苏州大学附属第二医院优势学科群 苏州市临床医学中心 苏州市民生科技项目 苏州大学附属第二医院预研基金 苏州大学青年教师科学基金项目 江南大学2018青年基金 江苏省研究生科研与实践创新计划项目(KYCX18_2531)National Natural Science Foundation of China The Clinical Medicine Technology Project of Jiangsu Province Advantage Discipline Groups of the Second Affiliated Hospital of Soochow University The Clinical Medical Center of Suzhou The Minsheng Science and Technology Project of Suzhou City The Second Affiliate Hospital of Soochow University Pre-research Foundation The Soochow University Youth Teacher Science Fund Project The Jiangnan University Youth Teacher Science Fund Project Postgraduate Research & Practice Innovation Program of Jiangsu Province
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