Open AccessTracing type 1 diabetic T ibet miniature pig's bone marrow mesenchymal stem cells in vitro by magnetic resonance imaging (1型糖尿病西藏小型猪骨髓间充质干细胞体外磁共振成像示踪)
Author(s) -
Tang KuanXiao,
Yan JinHua,
Shen YunFeng,
Li BaoYing,
Chen YingMing,
Liu DaYue,
Ma DeDong,
Li Jie,
Liang Hua,
Weng JianPing
Publication year - 2014
Publication title -
journal of diabetes
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.949
H-Index - 43
eISSN - 1753-0407
pISSN - 1753-0393
DOI - 10.1111/1753-0407.12084
Subject(s) - mesenchymal stem cell , trypan blue , green fluorescent protein , in vitro , bone marrow , transfection , pathology , flow cytometry , magnetic resonance imaging , microbiology and biotechnology , staining , medicine , chemistry , cell culture , biology , biochemistry , genetics , radiology , gene
Background Traditional cell‐tracking methods fail to meet the needs of preclinical or clinical research. Thus, the aim of the present study was to establish a new method of double labeling bone marrow mesenchymal stem cells ( BMSCs ) from type 1 diabetic ( T1D ) minipigs with super‐paramagnetic iron oxide ( SPIO ) and enhanced green fluorescent protein ( eGFP ) and tracing them using MRI in vitro. Methods Isolated BMSCs from T1D minipigs were labeled with eGFP and different concentrations of SPIO . The effects of lentivirus ( LV )‐ eGFP transfection and SPIO on the viability and growth curves of BMSCs were determined by T rypan blue exclusion, the 3‐(4,5‐dimethyl‐2 thiazoyl)‐2,5‐diphenyl‐ 2H ‐tetrazolium bromide assay and flow cytometry. Cellular ultrastructure was evaluated by transmission electron microscopy. Magnetic resonance imaging was used to evaluate BMSCs labeled with SPIO ‐ eGFP complexes 6 weeks after labeling. Results Expression of eGFP in BMSCs peaked 96 h after transfection with LV ‐ eGFP . Prussian blue staining revealed scattered blue granules in the cytoplasm of SPIO ‐labeled cells. Transmission electron microscopy revealed that the dense granules aggregated mainly in secondary lysosomes. On MRI , T 2* ‐weighted imaging was far more sensitive for SPIO ‐labeled BMSCs than other image sequences 3 and 6 weeks after the cells had been labeled with SPIO ‐ eGFP . Conclusions We have developed a relatively simple and safe method for double labeling of BMSCs from T1D minipigs using SPIO and LV ‐ eGFP and tracing them in vitro by MRI for 6 weeks.