Triple-modal imaging of stem-cells labeled with multimodal nanoparticles, applied in a stroke model

查看全文 作　　者：Helio Rodrigues da [1]Silva;Javier Bustamante [1]Mamani;Mariana Penteado [2]Nucci;Leopoldo Penteado [3]Nucci;Andrea Tiemi [1]Kondo;Daianne Maciely Carvalho [4]Fantacini;Lucas Eduardo Botelho de [4]Souza;Virginia Picanço-[4]Castro;Dimas Tadeu [4]Covas;José Mauro [1]Kutner;Fernando Anselmo de [1]Oliveira;Nelson [1]Hamerschlak;Lionel Fernel [1]Gamarra 机构地区：[1]Hospital Israelita Albert Einstein;[2]LIM44, Hospital das Clinicas HCFMUSP, Faculdade de Medicina,Universidade de São Paulo;[3]Centro Universitário do Planalto Central;[4]Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo 出　　处：《World Journal of Stem Cells》索引2019年第11卷第2期,共24页 基　　金：Supported by Conselho Nacional de Desenvolvimento Científico e Tecnológico,No.CNPq-465259/2014-6,and No.CNPq-400856/2016-6;São Paulo State Research Support Foundation,No.2014/50983-3,and No.2016/21470-3 摘　　要：BACKGROUND Mesenchymal stem cells(MSCs) have been widely tested for their therapeutic efficacy in the ischemic brain and have been shown to provide several benefits. A major obstacle to the clinical translation of these therapies has been the inability to noninvasively monitor the best route, cell doses, and collateral effects while ensuring the survival and effective biological functioning of the transplanted stem cells. Technological advances in multimodal imaging have allowed in vivo monitoring of the biodistribution and viability of transplanted stem cells due to a combination of imaging technologies associated with multimodal nanoparticles(MNPs) using new labels and covers to achieve low toxicity and longtime residence in cells.AIM To evaluate the sensitivity of triple-modal imaging of stem cells labeled with MNPs and applied in a stroke model.METHODS After the isolation and immunophenotypic characterization of human bonemarrow MSCs(hBM-MSCs), our team carried out lentiviral transduction of these cells for the evaluation of bioluminescent images(BLIs) in vitro and in vivo. In addition, MNPs that were previously characterized(regarding hydrodynamic size, zeta potential, and optical properties), and were used to label these cells,analyze cell viability via the 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide assay and BLI analysis, and quantify the internalization process and iron load in different concentrations of MNPs via magnetic resonance imaging(MRI),near-infrared fluorescence(NIRF), and inductively coupled plasma-mass spectrometry(ICP-MS). In in vivo analyses, the same labeled cells were implanted in a sham group and a stroke group at different times and under different MNP concentrations(after 4 h or 6 d of cell implantation) to evaluate the sensitivity of triple-modal images.RESULTS hBM-MSC collection and isolation after immunophenotypic characterization were demonstrated to be adequate in hBM samples. After transduction of these cells with luciferase(hBM-MSCLuc), we detected a maximum BLI intensity of 2.0 x10^8 photons/s in samples of 10~6 hBM-MSCs. Analysis of the physicochemical characteristics of the MNPs showed an average hydrodynamic diameter of 38.2 ±0.5 nm, zeta potential of 29.2 ± 1.9 mV and adequate colloidal stability without agglomeration over 18 h. The signal of iron load internalization in hBM-MSCLuc showed a close relationship with the corresponding MNP-labeling concentrations based on MRI, ICP-MS and NIRF. Under the highest MNP concentration, cellular viability showed a reduction of less than 10% compared to the control.Correlation analysis of the MNP load internalized into hBM-MSCLuc determined via the MRI, ICP-MS and NIRF techniques showed the same correlation coefficient of 0.99. Evaluation of the BLI, NIRF, and MRI signals in vivo and ex vivo after labeled hBM-MSCLuc were implanted into animals showed differences between different MNP concentrations and signals associated with different techniques(MRI and NIRF; 5 and 20 μg Fe/mL; P < 0.05) in the sham groups at 4 h as well as a time effect(4 h and 6 d; P < 0.001) and differences between the sham and stroke groups in all images signals(P < 0.001).CONCLUSION This study highlighted the importance of quantifying MNPs internalized into cells and the efficacy of signal detection under the triple-image modality in a stroke model. 关 键 词：Multimodal nanoparticles Human bone marrow mesenchymal stem cells Near-infrared fluorescence image Magnetic resonance image BIOLUMINESCENCE STROKE