Ex Vivo Magnetic Resonance Imaging of Transplanted Hepatocytes in a Rat Model of Acute Liver Failure
Ex Vivo Magnetic Resonance Imaging of Transplanted Hepatocytes in a Rat Model of Acute Liver Failure
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DOI：
10.3727/096368913x663596
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发表时间：
2014-01-01
影响因子：
3.3
通讯作者：
Hughes, Robin D.
中科院分区：
文献类型：
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作者：
Puppi, Juliana;Modo, Michel;Hughes, Robin D.
肝细胞移植正在被评估为肝移植的替代方案。然而，肝细胞移植后的命运还没有很好的定义。本研究的目的是提高肝细胞标记在体外使用超顺磁性氧化铁纳米粒子（SPIO）和磁共振成像（MRI）跟踪标记的细胞进行体内实验。用临床批准的SPIO（12.5 μ g Fe/ml）和作为转染剂的硫酸鱼精蛋白（3 μ g/ml）体外标记人和大鼠肝细胞16小时。增加细胞铁的摄取，获得和细胞活力和功能显示不受影响的标记。标记细胞（2，000/μ l）可在体外使用7 T MR扫描仪在T-2加权图像上检测到。在急性肝衰竭（ALE）大鼠模型中，雌性受体在腹腔注射D-半乳糖胺（1.2 g/kg）后28-30 h接受2 x 107雄性大鼠肝细胞脾内移植。有四个组（每组n=4）：溶剂注射、用CM-DiI标记的新鲜分离的细胞注射、用CM-DiI标记的培养细胞注射以及用SPTO和CM-DiI两者标记的培养细胞注射。在ALF诱导后第7天获得7 T MRI的离体T*（2）加权梯度回波图像。移植后6天，在大鼠肝脏中检测到SPIO，表现为存活动物的MRI信号强度降低。在组织学上，大多数SPIO位于枯否细胞中，表明标记的肝细胞被清除。此外，通过荧光染料或Y染色体（Sry-2基因）的PCR，在肝脏中无法检测到标记细胞。总之，建立了用SPIO标记人肝细胞的最佳条件，并且不影响细胞活力或代谢功能，并且足以用于体外MRI检测。然而，移植后肝细胞的清除限制了MRI评估长期肝细胞植入的价值。
Hepatocyte transplantation is being evaluated as an alternative to liver transplantation. However, the fate of hepatocytes after transplantation is not well defined. The aims of the study were to improve hepatocyte labeling in vitro using superparamagnetic iron oxide nanoparticles (SPIOs) and to perform in vivo experiments on tracking labeled cells by magnetic resonance imaging (MRI). Human and rat hepatocytes were labeled in vitro for 16 h with clinically approved SPIOs (12.5 mu g Fe/ml) and protamine sulfate (3 mu g/ml) as a transfection agent. Increased cellular iron uptake was obtained, and cell viability and function were shown not to be affected by labeling. Labeled cells (2,000/mu l) could be detected on T-2-weighted images in vitro using a 7T MR scanner. In a rat model of acute liver failure (ALE), female recipients received intrasplenic transplantation of 2 x 10(7) male rat hepatocytes 28-30 h after intraperitoneal injection of D-galactosamine (1.2 g/kg). There were four groups (n=4 each): vehicle injection, injection of freshly isolated cells labeled with CM-DiI, injection of cultured cells labeled with CM-DiI, and injection of cultured cells labeled with both SPTOs and CM-DiI. Ex vivo T*(2)-weighted gradientecho images at 7T MRI were acquired at day 7 post-ALF induction. Six days after transplantation, SPIOs were detected in the rat liver as a decrease in the MRI signal intensity in the surviving animals. Histologically, most of the SPIOs were located in Kupffer cells, indicating clearance of labeled hepatocytes. Furthermore, labeled cells could not be detected in the liver by the fluorescent dye or by PCR for the Y-chromosome (Sry-2 gene). In conclusion, optimum conditions to label human hepatocytes with SPIOs were established and did not affect cell viability or metabolic function and were sufficient for in vitro MRI detection. However, the clearance of hepatocytes after transplantation limits the value of MRI for assessing long-term hepatocyte engraftment.