Dodd C H, Hsu H C, Chu W J, Yang P, Zhang H G, Mountz J D, Zinn K, Forder J, Josephson L, Weissleder R, Mountz J M, Mountz J D
Division of Clinical Immunology and Rheumatology, Department of Medicine, The University of Alabama at Birmingham, 35294, USA.
J Immunol Methods. 2001 Oct 1;256(1-2):89-105. doi: 10.1016/s0022-1759(01)00433-1.
The present study analyzed the feasibility of using magnetic resonance imaging (MRI) to monitor T-cell homing in vivo after loading T cells with superparamagnetic iron oxide (CLIO) nanoparticles derivatized with a peptide sequence from the transactivator protein (Tat) of HIV-1. T cells were isolated from C57BL/6 (B6) mice and loaded with 0, 400, 800, 1600, or 8000 ng/ml of FITC conjugated CLIO-Tat (FITC-CLIO-Tat). There was a dose-dependent uptake of FITC-CLIO-Tat by T cells. Stimulation of FITC-CLIO-Tat loaded T cells with anti-CD3 (0.1 microg/ml) plus IL-2 (5 ng/ml) elicited normal activation and activation-induced cell death (AICD) responses, and normal upregulation of CD69, ICAM-1 (CD54), L-selectin (CD62L), and Fas. The FITC-CLIO-Tat loaded T cells (3 x 10(7)) were transferred intravenously (i.v.) into B6 mice and the in vivo MRI of mice was acquired using a spin-echo pulse sequence at 4.7 T with a Bruker Biospec system. Homing of T cells into the spleen was observed by a decrease in MRI signal intensity within 1 h after the transfer, which remained decreased for 2-24 h after transfer. These homing data were confirmed by FACS analysis and biodistribution analysis using 125I-CLIO-Tat. Thus, T cells can be efficiently loaded with FITC-CLIO-Tat without interfering with their normal activation and AICD, or homing to the spleen, and the biodistribution of FITC-CLIO-Tat loaded T cells can be monitored in vivo over time by MRI.
本研究分析了使用磁共振成像（MRI）监测经用来自HIV-1反式激活蛋白（Tat）的肽序列衍生化的超顺磁性氧化铁（CLIO）纳米颗粒加载T细胞后体内T细胞归巢的可行性。从C57BL/6（B6）小鼠中分离T细胞，并用0、400、800、1600或8000 ng/ml的异硫氰酸荧光素（FITC）偶联的CLIO-Tat（FITC-CLIO-Tat）加载。T细胞对FITC-CLIO-Tat的摄取呈剂量依赖性。用抗CD3（0.1μg/ml）加白细胞介素-2（5 ng/ml）刺激加载了FITC-CLIO-Tat的T细胞，引发正常的激活和激活诱导的细胞死亡（AICD）反应，以及CD69、细胞间黏附分子-1（ICAM-1，CD54）、L-选择素（CD62L）和Fas的正常上调。将加载了FITC-CLIO-Tat的T细胞（3×10⁷）静脉内（i.v.）转移到B6小鼠中，并使用Bruker Biospec系统在4.7 T下通过自旋回波脉冲序列获取小鼠的体内MRI。转移后1小时内通过MRI信号强度降低观察到T细胞归巢到脾脏，转移后2 - 24小时内信号强度仍保持降低。这些归巢数据通过流式细胞术分析和使用¹²⁵I-CLIO-Tat的生物分布分析得到证实。因此，T细胞可以有效地加载FITC-CLIO-Tat，而不会干扰其正常激活和AICD，或归巢到脾脏，并且加载了FITC-CLIO-Tat的T细胞的生物分布可以通过MRI在体内随时间进行监测。
