Cell Tracking and imaging gene expression in the brain

描述（由申请人提供）：脑的代谢紊乱和遗传性疾病可以通过神经干细胞移植潜在地治愈。这些细胞可以迁移并分化成神经元、星形胶质细胞或少突胶质细胞，从而可以传递治疗基因，甚至取代大脑中的缺陷细胞。发展干细胞治疗的主要挑战是非侵入性地监测这些供体细胞的迁移和表达。绿色荧光蛋白和荧光素酶仅适用于浅表器官，而单纯疱疹病毒-胸苷激酶gone需要与不穿过血脑屏障的阿昔洛韦配体结合。这些技术的灵敏度很高，但空间分辨率非常有限。用氧化铁颗粒标记的干细胞可以使用磁共振成像在体内检测。这些细胞也可以用慢病毒多巴胺2型受体标记，并可以通过PET成像以监测其体内基因表达。该提案的总体目标是评估磁共振成像（MRI）和正电子发射断层扫描（PET）在脑细胞跟踪和成像基因表达中的效用。该假设将通过以下具体目的进行测试：目的1：使用高分辨率体内MR成像在小鼠脑中连续监测用临床批准的超顺磁性氧化铁（SPIO）颗粒标记的C17.2神经祖细胞的迁移和植入。目标二：使用突变型多巴胺2型受体（D2 R-80 A）和3-（2 '-[18 F]-氟乙基）-螺哌隆（FESP）配体，通过微型PET成像对小鼠脑中供体C17.2神经祖细胞的基因表达进行成像。目的3：使用体内MR和PET成像，共同记录C17.2细胞的迁移与D2 R受体的基因表达。 神经C17.2祖细胞将用慢病毒D2 R-80 A受体和临床批准的氧化铁颗粒转导。C17.2细胞从克隆整合位点表达β-半乳糖苷酶基因，其将用作独立的供体细胞标志物。将细胞颅内移植到成年小鼠的海马体和新生小鼠的脑室中。将使用自旋回波和梯度回波MR成像连续监测这些细胞的迁移。D2 R受体的Gone表达将随后与FESP配体结合，并将使用微型PET成像。将使用解剖标记物和β-半乳糖苷酶活性的免疫组织化学染色在体内和体外进行MRI和PET图像的相关性。

DESCRIPTION (provided by applicant): Metabolic disorders and genetic diseases of the brain can potentially be cured by transplantation of neural stem cells. These cells can migrate and differentiate into neurons, astroglia or oligodendrocytes and thereby can deliver therapeutic genes or even replace defective cells in the brain. A major challenge towards development of stem cell therapy has been to non-invasively monitor the migration and gone expression of these donor cells. Green fluorescence protein and luciferase are suitable only for superficial organs while herpes-simplex virus-thymidine kinase gone requires binding to acyclovir ligands that do not cross the blood brain barrier. The sensitivity of these techniques is high but spatial resolution is very limited. Stem cells labeled with iron oxide particles can be detected in vivo using magnetic resonance imaging. These cells can also be labeled with a lentiviral dopamine type 2 receptor and can be imaged by PET to monitor their gene expression in vivo. The overall goal for this proposal is to evaluate the utility of magnetic resonance imaging (MRI) and positron emission tomography (PET) for cell tracking and imaging gene expression in the brain. This hypothesis will be tested with the following specific aims: Aim 1: To serially monitor migration and engraftment of C17.2 neural progenitor cells tagged with clinically approved super paramagnetic iron oxide (SPIO) particles using high resolution in vivo MR imaging in a mouse brain. Aim 2: To image gene expression in donor C17.2 neural progenitor cells in the mouse brain using the mutant dopamine type 2 receptor (D2R-80A) and 3-(2'-[18F]-fluoroethyl)-spiperone (FESP) ligand with micro PET imaging. Aim 3: To co-register migration of C17.2 cells with gene expression of the D2R receptor using in vivo MR and PET imaging. Neural C17.2 progenitor cells will be transduced with the lentiviral D2R-80A receptor and with clinically approved iron oxide particles. C17.2 cells express the beta-galactosidase gene from a clonally integrated site which will be used as an independent donor cell marker. The cells will be transplanted intracranially in the hippocampus of adult mice and in the ventricles of neonatal mice. Migration of these cells will be monitored serially using spin echo and gradient echo MR imaging. Gone expression of the D2R receptor will be followed by its binding with FESP ligand and will be imaged using a micro PET. Correlation of MRI and PET images will be performed in vivo and in vitro using anatomical markers and with immunohistochemical staining for the beta-galactosidase activity.