High-Resolution Morphological Approach to Analyse Elastic Laminae Injuries of the Ascending Aorta in a Murine Model of Marfan Syndrome

Júlia López-Guimet,Gustavo Egea,Pablo Loza-Alvarez,Jordi Andilla

doi:10.1038/s41598-017-01620-8

Abstract

In Marfan syndrome, the tunica media is disrupted, which leads to the formation of ascending aortic aneurysms. Marfan aortic samples are histologically characterized by the fragmentation of elastic laminae. However, conventional histological techniques using transverse sections provide limited information about the precise location, progression and 3D extension of the microstructural changes that occur in each lamina. We implemented a method using multiphoton excitation fluorescence microscopy and computational image processing, which provides high-resolution en-face images of segmented individual laminae from unstained whole aortic samples. We showed that internal elastic laminae and successive 2nd laminae are injured to a different extent in murine Marfan aortae; in particular, the density and size of fenestrae changed. Moreover, microstructural injuries were concentrated in the aortic proximal and convex anatomical regions. Other parameters such as the waviness and thickness of each lamina remained unaltered. In conclusion, the method reported here is a useful, unique tool for en-face laminae microstructure assessment that can obtain quantitative three-dimensional information about vascular tissue. The application of this method to murine Marfan aortae clearly shows that the microstructural damage in elastic laminae is not equal throughout the thickness of the tunica media and in the different anatomical regions of the ascending aorta.

Highlights

Vessel, animal species and age[8,9,10]
Based on endogenous tissue sources of nonlinear signals, the two-photon excitation fluorescence (TPEF) signal arises from the elastin content in elastic lamellae and the second harmonic generation (SHG) signal originates from collagen fibres located at the adventitia and interlamellar spaces
The transverse image of the aortic tissue can be obtained by visualizing the 3D image volume from the XZ or YZ perspectives (Fig. 1C and D), which was comparable with conventional histological preparations visualized by bright field microscopy (Fig. 1E and F)

Summary

Introduction

Vessel, animal species and age[8,9,10]. Their role is not yet well-established, but it is thought that they facilitate the flow of nutrients and the connection between cells located in different interlamellar spaces, and contribute to the developmental modelling of the IEL11. No further data is available on lamellae 3D microstructure features in health or disease In this context, our aim was to provide further insights into the characterization of elastic lamellae microstructure, including fenestrae features, using the ascending aorta of MFS Fbn1C1039G/+ mice as a histopathological model of tunica media alterations that typically occur in aortic aneurysms. Our aim was to provide further insights into the characterization of elastic lamellae microstructure, including fenestrae features, using the ascending aorta of MFS Fbn1C1039G/+ mice as a histopathological model of tunica media alterations that typically occur in aortic aneurysms To achieve this aim, we applied a recently implemented microscopy and computational method that we developed, which provides the en-face TPEF image of segmented individual lamellae from unstained whole aortic samples. The application of our methodology to MFS murine aortae showed relevant lamellae fenestrae differences between IEL and 2nd laminae, and between the proximal-concavity and the rest of the anatomical regions in the ascending aorta

Objectives

Methods

Results

Conclusion