Semi-automated rapid quantification of brain vessel density utilizing fluorescent microscopy.
Semi-automated rapid quantification of brain vessel density utilizing fluorescent microscopy.
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DOI：
10.1016/j.jneumeth.2016.06.012
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发表时间：
2016-09-01
影响因子：
3
通讯作者：
Lockman PR
中科院分区：
文献类型：
--
作者：
Bohn KA;Adkins CE;Mittapalli RK;Terrell-Hall TB;Mohammad AS;Shah N;Dolan EL;Nounou MI;Lockman PR
血管密度的测量在表征健康和病变组织方面具有重要价值，特别是在血管密度在不同区域之间变化的大脑中。此外，对脑血管大小的了解有助于区分毛细血管和更大的血管，如小动脉和小静脉。不幸的是，很少有尖端的方法可用于实验室快速量化血管密度。我们开发了一种快速显微方法，可以量化大脑血管的数量和直径。利用这种方法，我们用三种示踪剂对小鼠和大鼠两种肿瘤模型的五个脑区血管密度进行了表征。我们观察了不同脑区的切片数/mm2：胼胝体（161±7），海马（266±18），上丘（300±24），额皮质（391±55），下丘（692±18）（n=5只动物）。不同物种间、不同示踪剂（70kDa和2000kDa Texas Red， p>0.05）脑区域数据无显著差异（p>0.05）。临床前脑转移瘤的血管密度降低（62% - 79%），但在大鼠胶质瘤模型中血管密度增加（62%）。我们的值与已发表的文献相似（p < 0.05）。我们将这种方法应用于脑肿瘤，观察到与正常皮质区相比，乳腺癌脑转移灶的血管/mm2减少了2.5倍（p>0.05）。相比之下，胶质瘤模型的血管密度明显更高（切片/mm2 736±84；p<0.05）。总之，我们提出了一种血管密度计数方法，它是快速，敏感的，并使用荧光显微镜无抗体。血管密度在脑区域之间以及健康和病变组织之间存在显著差异。然而，目前的方法既费力又耗时。本文提出了一种快速简便的定量脑内血管密度的方法。正常的脑血管系统可见吲哚菁绿色荧光。
Measurement of vascular density has significant value in characterizing healthy and diseased tissue, particularly in brain where vascular density varies among regions. Further, an understanding of brain vessel size helps distinguish between capillaries and larger vessels like arterioles and venules. Unfortunately, few cutting edge methodologies are available to laboratories to rapidly quantify vessel density. We developed a rapid microscopic method, which quantifies the numbers and diameters of blood vessels in brain. Utilizing this method we characterized vascular density of five brain regions in both mice and rats, in two tumor models, using three tracers. We observed the number of sections/mm2 in various brain regions: genu of corpus callosum 161 ± 7, hippocampus 266 ± 18, superior colliculus 300 ± 24, frontal cortex 391 ± 55, and inferior colliculus 692 ± 18 (n=5 animals). Regional brain data were not significantly different between species (p>0.05) or when using different tracers (70kDa and 2000kDa Texas Red; p>0.05). Vascular density decreased (62–79%) in preclinical brain metastases but increased (62%) a rat glioma model. Our values were similar (p>0.05) to published literature. We applied this method to brain-tumors and observed brain metastases of breast cancer to have a ~2.5-fold reduction (p>0.05) in vessels/mm2 compared to normal cortical regions. In contrast, vascular density in a glioma model was significantly higher (sections/mm2 736 ± 84; p<0.05). In summary, we present a vascular density counting method that is rapid, sensitive, and uses fluorescence microscopy without antibodies. Vascular density varies significantly between brain regions and between healthy and diseased tissue. However, current methods are arduous and time consuming. Herein we present a rapid simple method to quantify vascular density in brain. Normal brain vasculature is seen with indocyanine green fluorescence within the image.