gms | German Medical Science

Objective: Changes in tissue composition due to degenerative processes after spinal cord injury (SCI) involve a series of complex biochemical alterations. Vibrational optical spectroscopic mapping techniques may be used to probe molecular structure. They were investigated for their ability to characterize alterations in biochemical composition after SCI.

Methods: SCI was induced in adult wistar rats by T9 – T10 hemisection. Unstained cryosections of lesioned spinal cord were investigated with Fourier Transform-Infraredspectroscopy (FT-IR), Raman spectroscopy and Coherent Antistokes Raman Spectroscopy (CARS) in combination with microscopic mapping systems. Images of biochemical features were obtained by application k-means clustering and by plotting intensity variations of specific vibrational bands. Consecutive sections were submitted to histological analysis.

Results: We detected injury-induced modifications in the spectroscopic signature of the tissue with all three techniques tested. The most prominent change seen in lesioned tissue was the dramatic decrease in vibrational bands related to lipids (wavenumbers: FT-IR: 1465 cm^-1 and 1740 cm^-1; Raman: 1063 cm^-1, 1300 cm^-1 and 1440 cm^-1). Additionally, signals related to nuclear acids, polysaccharides and chondroitin sulphate proteoglycans (FT-IR: 993 cm^-1 and 1140 cm^-1 Raman: 840 cm^-1, 522 cm^-1 and 640 cm^-1) proved to be good markers for identification of the lesion and formation of the scar. Raman mapping proved to be most effective in detailed mapping of the lipid distribution. Several bands can be assigned to specific chemical bond vibrations of lipids, in particular of cholesterol (607 cm^-1 and 700 cm^-1), which allowed to generate distribution maps of these compounds. Clustering of FTIR-data reported different degrees of degeneration in the transition zone between lesion and nearly normal tissue that were lacking in Raman maps. CARS-imaging details the C-H stretching bonds of lipids and provides also morphological information in nearly real-time that cannot be retrieved from Raman or FT-IR maps.

Conclusions: We demonstrated that label-free purely optical spectroscopy techniques provide insight into the biochemical composition of nervous tissue, and enable to create visual maps of tissue degeneration after SCI. The application of these methods with cellular resolution to injured neuronal tissue will enable detailed monitoring of degeneration and objective cross-sample analysis.