摘要
目的大鼠脂肪来源的成体干细胞(ADSC)转染人超极化激活环核苷酸门控离子通道-2(hHCN2)基因,观察其作为起搏细胞的可行性。方法取生长良好的ADSC转染hHCN2基因,通过RT-PCR、Western blot分析、免疫荧光技术检测hHCN2基因及蛋白的表达,用膜片钳技术检测细胞的电生理特征并记录其内向电流。将转染hHCN2基因的ADSC与心室肌细胞共培养,观察其对心室肌细胞自发性搏动的影响。结果 hHCN2基因修饰的大鼠ADSC能表达较强的hHCN2基因及蛋白,还能够产生超级化激活内向离子流。将其与心室肌细胞共培养,能使心室肌细胞的自发性搏动明显增快增强。结论将hHCN2基因转染ADSC能够使其分化为具有起搏功能的起搏样细胞,为进一步优化生物起搏器的研究提供了材料。
Objective To transfect rat adipose tissue-derived adult stem cells （ADSCs） with the pacemaker gene of human hyperpolarization-activated cyclic nucleotide-gated ion channel 2 （ hHCN2）, and observe its feasibility of functioning as pacemaker cells. Methods First of all, healthy ADSCs were harvested and transfected with hHCN2 gene. Thereafter, we detected the expression of hHCN2 gene and protein by quantitative real-time PCR （qRT-PCR）, Western blot analysis and immunofluorescence cytochemistry and measured the electrophysiological properties and recorded the inward currents using patch clamp techniques. In addition, we mixed and co-cultured the transfected ADSCs and rat cadiomyocytes to observe the influence on the spontaneous beating rate of cadiomyocytes. Results Compared with the control ADSCs, the transfected ADSCs expressed the higher level of hHCN2 gene and protein （ P 〈 0.05）, and they could produce pacemaker current. When co-cultured with rat ventricular myocytes, they could accelerate the spontaneous beating rate of the ventricular myocytes （P〈0.05）. Conclusion After transfected with hHCN2 gene, ADSCs can work as pacemaker-like cells, which provides materials for the research of biological pacemakers.
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2013年第9期901-904,909,共5页
Chinese Journal of Cellular and Molecular Immunology
基金
江西省自然科学基金(2008GZY0045)
国家自然科学基金(81041097)
关键词
脂肪来源的干细胞
起搏样细胞
生物起搏
超极化激活环核苷酸门控离子通道-2
adipose tissue-derived adult stem cells~ pacemaker cells~ biological pacemaker~ hyperpolarization-activatedcyclic nucleotide-gated ion channel 2
