Second Harmonic Generation analysis of the ECM in idiopathic pulmonary fibrosis

 描述（由申请人提供）：特发性肺纤维化中ECM的二次谐波生成分析特发性肺纤维化（IPF）是一种以肺的无情瘢痕形成和硬化为特征的疾病，其导致美国每年估计有34，000人死亡。不幸的是，IPF患者的治疗选择非常有限，因为尚未发现有效的药物来阻止纤维化的进展。尽管胶原蛋白对正常和重塑ECM的结构组织都很重要，但对IPF中胶原蛋白结构与正常组织结构的差异知之甚少。显然需要开发高度特异性/灵敏度的技术来探测IPF组织中的胶原蛋白结构和组织。在这个项目中，我们将使用二次谐波产生（SHG）的高分辨率显微镜技术实现新的胶原蛋白特异性分析。这种方法是敏感的原纤组织，也次分辨率方面的宏观和超分子组装。在这里，我们将利用SHG显微镜来：1）确定病理性胶原组织如何（见于IPF）与正常组织不同; 2）识别和量化活动性纤维化区域（富含胶原蛋白III）从“老”或成熟的纤维化3）评估疾病进展期间弹性蛋白和胶原蛋白分布的变化;和4）将IPF组织学样品中高胶原III/I信号的区域与疾病活动的临床标志物相关联。作为项目的一部分， 我们将开发定制的自动化机器视觉程序，以根据严重程度自动对组织进行分类。我们将特别集中我们所有的努力，研究周围的成纤维细胞病灶，这将是确定由其他显微镜模式的结构。这些病灶被认为是ECM重塑的前沿，但其形成的动力学与整体纤维化过程的关系仍不清楚。我们假设这些结构变化将作为IPF的无标记生物标志物，并进一步假设胶原蛋白以与疾病进展相关的方式在病灶周围特异性改变。所获得的信息可能构成未来预后/诊断方案的基础。我们提出了2个目的：目的1极化分辨SHG以确定IPF的不同阶段中Col I/III和其他ECM变化的分布。 目标2.开发通过SHG可视化的IPF形态学变化的分类系统。

 DESCRIPTION (provided by applicant): Second Harmonic Generation analysis of the ECM in idiopathic pulmonary fibrosis Idiopathic pulmonary fibrosis (IPF) is a disorder characterized by unrelenting scarring and stiffening of the lungs that leads to the death of an estimated 34,000 individuals in the U.S. each year. Unfortunately, individuals with IPF have extremely limited treatment options, as no effective drugs have been identified to halt the progression of fibrosis. Despite the importance of collagens to the structural organization both normal and remodeled ECM, little is known about how collagen structure in IPF differs from that of normal tissue architecture. There is a clear need to develop highly specific/sensitive techniques to probe collagen structure and organization in IPF tissues. In this project we will implement new collagen specific analyses using the high resolution microscopy technique of Second Harmonic Generation (SHG). This method is sensitive to both the fibrillar organization and also sub-resolution aspects of macro and supramolecular assembly. Here we will utilize SHG microscopy to: 1) determine the how pathologic collagen organization (seen in IPF) differs from normal tissue; 2) identify and quantify areas of active fibrosis (enriched in collagen III) from "old" or mature fibrosis (high in collagen I) in IPF lung specimens; 3) assess changes in elastin and collagen distribution during disease progression; and 4) correlate areas of high collagen III/I signal in IPF histologic samples with clinical markers of disease activity. As part of the project, we will develop customized automated machine vision routines to automatically classify tissues in terms of severity. We will specifically focus all of our efforts on studying structure around fibroblastic foci, which will be identified by other microscope modalities. These foci are thought to be at the leading edge of ECM remodeling but the dynamics of their formation in relationship to the overall fibrotic process remain unclear. We hypothesize that these structural changes will serve as label- free biomarkers of IPF and further hypothesize that the collagen is altered specifically around foci in a manner which is associated with disease progression. The information gained may form the basis of future prognostic/diagnostic schemes. We propose 2 Aims: Aim 1 Polarization resolved SHG to determine distribution of Col I/III and other ECM changes in different stages of IPF. Aim 2. Develop classification system of morphological changes in IPF visualized by SHG.