首页> 中文期刊>暨南大学学报(自然科学与医学版) >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

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

     

摘要

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 ~ jjug/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 evaluated by Typan blue exclusion testing and MTT 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 label cells with SPIO by using PLL. The labeling efficiency was approximately 100% in the groups labeled with 25 ~ , 50 ~ , 75 ~ , and 100 ~ jxg/mL SPIO. SPIO labeling did not affect cell viability, but could inhibit cell proliferation capability. SPIO-labeled ADSCs transiently down-regulated gene and protein expression of TfR, and the lowest expression level of TfR mRNA was presented at 24th hour after SPIO labeling, but the lowest expression level of TfR protein expression was presented at 7th day after SPIO labeling. The gene and protein expression were decreased within the increase of SPIO labeling concentration, but when the SPIO labeling concentration reached 25μg/mL and above,TfR mRNA expression retained at a steady level. Conclusion: It was easy and efficient to label cells 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.%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(MTr)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25~100 μg/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rfR mRNA表达量以标记后24h时最低,TfR蛋白表达量以标记后7d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.

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