靶向整合素αvβ3分子影像探针定量评估糖尿病肾病早期微循环灌注改变的基础研究

To make sensitive and accurate evaluation of early perfusion changes in renal cortex of diabetic nephropathy (DN) is of vital importance in clinical practice. Integrin αvβ3 was proved to be important in the progression of diabetic nephropathy, which could be used to make quantitative analysis of abnormal microvascular hyperplasia in DN patients. It was considered as potential target for molecular imaging of angiogenesis in early stage of DN. In our previous researches, we had applied real time contrast-enhanced ultrasound and quantitative analysis to evaluate perfusion changes in renal cortex of early stage DN patients. We had built the hollow periodic mesoporous organosilicas (HPMOs) as targeted microbubbles. In our current research, we conceive to use HPMOs microbubbles targeted to integrin αvβ3 as molecular imaging probe. We want to apply targeted contrast-enhanced ultrasound method to evaluate early microvascular changes of renal cortex in DN rat model. We aimed to analyze in details the targeting imaging mechanism and various influence factors of quantitative process, to optimize quantitative fitted equation and quantitative methods. Our purpose is to screening of a reliable quantitative method and sensitive quantitative indexes. Various methods such as immunohistochemical staining, confocal laser scanning microscopy and Motic Advanced quantitative method would be used to verify the practicability and accuracy of our quantitative results. Our aim is to provide theory evidence and technical support for this method to be used clinically in early and specific evaluation of DN patients.

如何敏感、准确地评估糖尿病肾病（DN）早期肾皮质微循环灌注改变是提高糖尿病患者生存期的关键。研究证实单核/巨噬细胞系统参与的炎症反应和微血管病变是导致DN早期微循环灌注异常的重要因素，且两者均高表达整合素αvβ3，这使其成为潜在的分子成像的靶点。我们在前期利用超声造影定量分析DN患者肾皮质微循环灌注改变的基础上，研发了有序介孔有机硅（HPMOs）纳米粒子。本课题拟构建以HPMOs为载体的携抗整合素αvβ3的靶向特异性分子探针，应用靶向超声造影定量分析技术观察大鼠DN模型早期肾皮质微循环灌注的变化，深入研究靶向超声造影机制，优化定量拟合方程、筛选可靠的定量方法和敏感定量参数，并利用免疫组化、激光共聚焦显微镜、Motic Advanced等技术验证定量结果的可行性与准确性，为这一新技术用于临床早期、特异性评估糖尿病肾病肾皮质微循环灌注及其功能状况提供理论依据和技术支撑。

本项目以纳米超声造影剂微泡为载体，构建了针对肾小球血管内皮细胞整合素αvβ3 靶点的特异性分子探针，制备对靶向整合素αvβ3 的分子探针的物理性质、稳定性、体外寻靶能力等作出了初步评价。在大鼠糖尿病肾病（DN）动物模型的基础上，对早期DN大鼠肾皮质靶向超声造影的感兴趣区（region of interest, ROI）进行时间-强度曲线分析，生成ROI 内微泡充盈-消退时间强度曲线（wash-in and wash-out TIC），建立可行的肾皮质血流动力学数学模型，分析并阐明肾皮质微循环灌注超声定量的各类影响因素。探究定量肾皮质血流灌注的最佳方法和最佳参数、优化定量拟合方程。并将靶向整合素αvβ3 分子探针超声造影显示的肾皮质微循环灌注情况、敏感的靶向定量参数与HE染色、PAS染色、免疫组化等结果进行相关分析。验证肾皮质整合素αvβ3 的表达和靶向超声造影定量参数的相关性，为靶向整合素αvβ3 的分子探针及超声造影定量分析DN 早期肾皮质微循环灌注的改变提供理论依据。