Collagen Organization, Polarization Sensitivity and Image Quality in Human Corneas using Second Harmonic Generation Microscopy

Abstract

In this paper, a Second-Harmonic-Generation (SHG) microscope was used to study the relationship between collagen structural arrangement, image quality and polarization sensitivity in human corneas with different organizations. The degree of order (or alternatively, the Structural Dispersion, SD) was quantified using the structure tensor method. SHG image quality was evaluated with different objective metrics. Dependence with polarization was quantified by means of a parameter defined as polarimetric modulation, which employs polarimetric SHG images acquired with four independent polarization states. There is a significant exponential relationship between the quality of the SHG images and the SD of the samples. Moreover, polarization sensitivity strongly depends on collagen arrangement. For quasi- or partially organized specimens, there is a polarization state that noticeably improves the image quality, providing additional information often not seen in other SHG images. This does not occur in non-organized samples. This fact is closely related to polarimetric modulation, which linearly decreases with the SD. Understanding in more detail the relationships that take place between collagen distribution, image quality and polarization sensitivity brings the potential to enable the development of optimized SHG image acquisition protocols and novel objective strategies for the analysis and detection of pathologies related to corneal collagen disorders, as well as surgery follow-ups.