超顺磁氧化铁标记对干细胞转铁蛋白受体基因和蛋白表达的影响  

Effect of SPIO labeling on gene and protein expresion of transferrin receptor in stem cells

作　　者：颜丽芬[1,2] 刘再毅[1] 王广谊[1] 贾乾君[1] 梁长虹[1] 

机构地区：[1]广东省人民医院放射科,广东省医学科学院,广东广州510080 [2]南方医科大学,广东广州510515

出　　处：《暨南大学学报（自然科学与医学版）》2012年第4期395-401,共7页Journal of Jinan University(Natural Science & Medicine Edition)

基　　金：国家自然科学基金项目(30970798);广东省医学科学研究基金(A2011036)

摘　　要：目的:探讨超顺磁氧化铁(SPIO)标记对大鼠脂肪干细胞(ADSCs)内转铁蛋白受体(TfR)基因和蛋白表达的影响,及SPIO标记与TfR表达的剂量-效应关系。方法:从3周龄雄性SD大鼠脂肪组织中提取干细胞,用多聚赖胺酸(PLL)介导SPIO(质量浓度分别为12.5～、25～、50～、75～、100～μg/mL,PLL/SPIO为1∶0.03)标记大鼠ADSCs 12 h后,用普鲁士蓝染色、台盼蓝及噻唑蓝法(MTT)试验检测SPIO标记率、细胞活力及增殖力,并用逆转录聚合酶链反应(RT-PCR)、Western blot技术检测各质量浓度SPIO标记细胞内TfR表达情况。结果:PLL介导SPIO可以简单、高效地标记大鼠ADSCs。当SPIO终质量浓度为25～100μg/mL时,SPIO标记率达到近100%;各质量浓度SPIO标记可抑制细胞增殖,但对细胞活力没有明显影响。SPIO标记大鼠ADSCs内TfR表达水平下调,其中TfR mRNA表达量以标记后24 h时最低,TfR蛋白表达量以标记后7 d时最低。随着SPIO标记质量浓度的增加,TfR表达水平越低,持续时间越长,但当SPIO标记质量浓度达25μg/mL及以上时,TfR mRNA表达维持在一定的水平。结论:PLL介导SPIO可以简单、高效的标记大鼠ADSCs。PLL介导SPIO标记可引起大鼠ADSCs内TfR表达水平暂时性降低以减少对细胞外铁的摄取,且随着SPIO标记质量浓度的增加,TfR表达水平越低,低水平表达持续时间越长。Aim：To investigate the effects of SPIO labeling on TfR gene and protein expression in rat ADSCs, and the dose-response relationship of SPIO labeling and TfR expression. Methods： ADSCs were isolated from adipose tissue of 3 weeks old SD rats. Second-passaged rat ADSCs were labeled with 0 - , 12.5 - , 25 ~, 35 ~, 50 - ,75 ~ ,and 100 -Ixg/mL concentrations of SPIO（ the ratio of SPIO-to-PLL was 1：0. 03） for 12 hours and then were continued to incubate to certain time points. PLL-SPIO labeling efficiency was evaluated using Perl＇ s Prussian blue staining. Cell viability and proliferation were evalua- ted by Typan blue exclusion testing and MTr testing. Real-time quantitative RT-PCR and western blot were performed to evaluate TfR expression of SPIO labeled cells. Results： It was easy and efficient to la- bel ceils with SPIO by using PLL. The labeling efficiency was approximately 100% in the groups labeled with 25 ~ , 50 - , 75 - , and 100 ~ p^g/mL SPIO. SPIO labeling did not affect ceil viability, but could inhibit cell proliferation capability. SPIO-labeled ADSCs transiently down-regulated gene and protein ex- pression of TfR, and the lowest expression level of TfR mRNA was presented at 24th hour after SPIO labe- ling, but the lowest expression level of TfR protein expression was presented at 7th day after SPIO labe- ling. The gene and protein expression were decreased within the increase of SPIO labeling concentration, but when the SPIO labeling concentration reached 25 p^g/mL and above ,TfR mRNA expression retained at a steady level. Conclusion：It was easy and efficient to label ceils with SPIO by using PLL. Rat ADSCs can transiently down-regulate TfR expression in response to iron-overload after SPIO labeling, and within the increase of SPIO labeling concentration, the TfR expression level becomes lower.

关 键 词：超顺磁氧化铁 脂肪干细胞 标记 转铁蛋白受体 

分 类 号：R445.2[医药卫生—影像医学与核医学]