Maturation of the Meniscal Collagen Structure Revealed by Polarization-Resolved and Directional Second Harmonic Generation Microscopy

Abstract

We report Polarization-resolved Second Harmonic Generation (P-SHG) and directional SHG (forward and backward, F/B) measurements of equine foetal and adult collagen in meniscus, over large field-of-views using sample-scanning. Large differences of collagen structure and fibril orientation with maturation are revealed, validating the potential for this novel methodology to track such changes in meniscal structure. The foetal menisci had a non-organized and more random collagen fibrillar structure when compared with adult using P-SHG. For the latter, clusters of homogeneous fibril orientation (inter-fibrillar areas) were revealed, separated by thick fibers. F/B SHG showed numerous different features in adults notably, in thick fibers compared to interfibrillar areas, unlike foetal menisci that showed similar patterns for both directions. This work confirms previous studies and improves the understanding of meniscal collagen structure and its maturation, and makes F/B and P-SHG good candidates for future studies aiming at revealing structural modifications to meniscus due to pathologies.

Circular dichroism second-harmonic generation microscopy probes the polarity distribution of collagen fibrils

“…SHG signals were detected in the forward direction (U-AAAC aplanetic condenser 1.4 NA, Olympus) using a photon-counting photomultiplier tube (P25PC, SensTech) and suitable spectral filters (FF01-720/SP, FF01-680/SP and FF01-427/10, Semrock). The second setup used for fascia imaging is very similar as described in [39].…”

Multiscale molecular profiling of pathological bone resolves sexually dimorphic control of extracellular matrix composition

“…Furthermore, p-SHG has been applied to investigate collagenous alterations that accompany ageing ( Van Gulick et al, 2019 ), wound repair and scar formation ( Mostaço-Guidolin et al, 2017 ). In a number of skeletal studies, second-harmonic generation (SHG) has also been used to (1) visualise woven bone formation during fracture repair ( Park et al, 2019 ), (2) elucidate the effects of mechanical loading on collagen formation ( Mansfield et al, 2019 ; Pinsard et al, 2019 ), and (3) differentiate between normal and OI bone tissue ( Nadiarnykh et al, 2007 ). To date, however, multiscale quantitative analyses of collagen organisation, fibre alignment or anisotropy combined with the assessment of bone mineral composition within pathological bone have not been studied.…”

Polarization‐dependent second‐harmonic generation for collagen‐based differentiation of breast cancer samples

“…21 The dependence of nonlinear signal from the incident polarization of the excitation radiation can provide additional quantitative information about the samples even at subcellular level. [22][23][24] This information is relevant with the orientation of sample's SHG emitters at molecular and supramolecular scale. At molecular level and for cylindrical symmetry molecules, polarization-dependent second-harmonic generation (PSHG) can provide the angle between SHG emitters and their molecular symmetry axis.…”

“…The dependence of nonlinear signal from the incident polarization of the excitation radiation can provide additional quantitative information about the samples even at subcellular level 22–24 . This information is relevant with the orientation of sample's SHG emitters at molecular and supramolecular scale.…”