Comparative transcriptome of brain blood vessels: age, hypertension and diabetes

描述(申请人提供)：脑血管在中风的病理生理学中起着关键作用。我们都“知道”，脑血管很可能不同于其他器官的血管。例如，脑血管有独特的紧密连接，有助于形成血脑屏障。但除了这一障碍表型之外，人们对其他方面的了解也不多。令人惊讶的是，从未对脑血管进行系统分析，将它们与其他器官的脑血管进行比较。在这项RC2提案中，我们寻求填补这一重要的知识空白。我们的总体目标是组装小鼠脑、心脏和肾脏内皮细胞和平滑肌细胞的比较转录组(信使核糖核酸表达谱)。我们将提出以下问题：1.小鼠脑灰质和白质的血管基因表达是否有差异？2.脑血管的基因表达谱与心脏和肾脏的基因表达谱有何不同？3.年龄、高血压和糖尿病对脑血管的影响是否与心脏和肾脏的不同？将通过两种互补的方式收集内皮细胞和平滑肌细胞：激光捕获新鲜冰冻组织的显微解剖，以及从新鲜制备的整个组织匀浆中分离出原代细胞。基因表达将用Affymetrix小鼠基因组430 2.0芯片进行分析。对于年龄，我们将比较“年轻”(5个月)和“老年”(15个月)C57BL6小鼠。对于高血压，我们将比较正常血压的BPN/3J小鼠和自发性高血压的BPH/2J小鼠。对于糖尿病，我们将比较匹配的野生型对照和糖尿病db/db小鼠。这个项目应该产生一个新的数据库，可以被所有中风研究人员用来研究新的假设、机制和靶点。就血管功能障碍可能在神经退行性变中起作用而言，这些结果也可以影响其他广泛的中枢神经系统疾病。最后，尽管我们在这里主要关注的是中风，但了解两种主要疾病(高血压和糖尿病)如何系统性地影响血管基因表达，应该对许多其他生物医学领域也有广泛的帮助。 公共卫生相关性：目前还没有治疗中风的临床有效的神经保护疗法。最近的数据表明，仅关注神经元是不够的，大脑内的血管非常重要。然而，没有人知道这些脑血管与我们身体其他器官的血管有什么不同。在这里，我们试图找出脑血管的完整基因表达谱，并将其与心脏和肾脏进行比较。我们还将发现衰老、高血压和糖尿病是如何改变这些基因模式的。这个数据库对于中风的研究，在未来寻找新的治疗靶点方面，应该是无价的。

DESCRIPTION (Provided by Applicant): Cerebral blood vessels play a key role in the pathophysiology of stroke. We all "know" that brain blood vessels are likely to be different from vessels in other organs. For example, brain blood vessels have unique tight junctions that contribute to the blood-brain barrier. But beyond this barrier phenotype, not much else is known. Surprisingly, a systematic analysis of brain blood vessels comparing them with those from other organs has never been performed. In this RC2 proposal, we seek to fill this important gap in knowledge. Our overall goal is to assemble a comparative transcriptome (mRNA expression profiles) of endothelial and smooth muscle cells in mouse brain, heart and kidney. We will ask the following questions: 1. Are there differences in vascular gene expression in gray matter vs. white matter of mouse brain? 2. Are gene expression profiles of brain blood vessels different from those in heart and kidney? 3. Will age, hypertension and diabetes affect brain blood vessels differently compared to those in heart and kidney? Endothelial and smooth muscle cells will be collected in two complementary ways: laser capture microdissection of fresh-frozen tissue, and primary cell isolation from freshly prepared whole tissue homogenates. Gene expression will be analyzed with the Affymetrix Mouse Genome 430 2.0 chip. For age, we will compare "young" (5 month) vs. "old" (15 month) C57Bl6 mice. For hypertension, we will compare normotensive BPN/3J mice vs. spontaneously hypertensive BPH/2J mice. For diabetes, we will compare matching wildtype controls vs. diabetic db/db mice. This project should yield a novel database that can be used by all stroke researchers to investigate new hypotheses, mechanisms and targets. Insofar as vascular dysfunction may play a role in neurodegeneration, these results can also impact a wide spectrum of other CNS disorders. And finally, although our primary focus here is on stroke, gaining an understanding of how two major diseases (hypertension and diabetes) systematically affect vascular gene expression should be broadly useful for many other biomedical fields as well. PUBLIC HEALTH RELEVANCE: There is no clinically effective neuroprotective therapy for stroke. Recent data suggest that focusing on neurons alone is not enough, and that blood vessels within the brain are very important. Yet, no one knows how these brain blood vessels are different from blood vessels in other organs in our body. Here, we seek to find out the full gene expression profile of brain blood vessels, and compare it with heart and kidney. And we will also find out how aging, hypertension and diabetes alter these gene patterns. This database should be invaluable for stroke research, in terms of finding new therapeutic targets in the future.