Advanced Magnetic Resonance Center, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104, USA.
Free Radic Biol Med. 2010 Mar 1;48(5):691-703. doi: 10.1016/j.freeradbiomed.2009.12.012. Epub 2009 Dec 23.
Increased iNOS expression is often found in brain tumors, such as gliomas. The goal of this study was to develop and assess a novel molecular MRI (mMRI) probe for in vivo detection of iNOS in rodent models for gliomas (intracerebral implantation of rat C6 or RG2 cells or ethyl nitrosourea-induced glioma). The probe we used incorporated a Gd-DTPA (gadolinium(III) complex of diethylenetriamine-N,N,N',N'',N''-pentaacetate) backbone with albumin and biotin moieties and covalent binding of an anti-iNOS antibody (Ab) to albumin (anti-iNOS probe). We used mMRI with the anti-iNOS probe to detect in vivo iNOS levels in gliomas. Nonimmune normal rat IgG coupled to albumin-Gd-DTPA-biotin was used as a control nonspecific contrast agent. By targeting the biotin component of the anti-iNOS probe with streptavidin Cy3, fluorescence imaging confirmed the specificity of the probe for iNOS in glioma tissue. iNOS levels in glioma tumors were also confirmed via Western blots and immunohistochemistry. The presence of plasma membrane-associated iNOS in glioma cells was established by transmission electron microscopy and gold-labeled anti-iNOS Ab. The more aggressive RG2 glioma was not found to have higher levels of iNOS compared to C6. Differences in glioma vascularization and blood-brain barrier permeability between the C6 and the RG2 gliomas are discussed. In vivo assessment of iNOS levels associated with tumor development is quite feasible in heterogeneous tissues with mMRI.
在脑肿瘤（如神经胶质瘤）中经常发现 iNOS 表达增加。本研究的目的是开发和评估一种新型的分子 MRI（mMRI）探针，用于在神经胶质瘤的啮齿动物模型中（脑内植入大鼠 C6 或 RG2 细胞或乙基亚硝脲诱导的神经胶质瘤）体内检测 iNOS。我们使用的探针结合了 Gd-DTPA（二乙三胺五乙酸的钆(III)络合物）骨架、白蛋白和生物素部分，并将抗 iNOS 抗体（Ab）共价结合到白蛋白上（抗 iNOS 探针）。我们使用 mMRI 结合抗 iNOS 探针来检测神经胶质瘤中的体内 iNOS 水平。将与白蛋白-Gd-DTPA-生物素偶联的非免疫正常大鼠 IgG 用作对照非特异性对比剂。通过用链霉亲和素 Cy3 靶向抗 iNOS 探针的生物素部分，荧光成像证实了探针在神经胶质瘤组织中对 iNOS 的特异性。还通过 Western blot 和免疫组织化学证实了神经胶质瘤肿瘤中的 iNOS 水平。通过透射电子显微镜和金标记的抗 iNOS Ab 证实了神经胶质瘤细胞中存在与质膜相关的 iNOS。与 C6 相比，侵袭性更强的 RG2 神经胶质瘤并未发现 iNOS 水平更高。讨论了 C6 和 RG2 神经胶质瘤之间的肿瘤血管生成和血脑屏障通透性的差异。使用 mMRI 在异质组织中对与肿瘤发展相关的 iNOS 水平进行体内评估是相当可行的。
