Characterization of putative stem cell populations in the cornea using synchrotron infrared microspectroscopy

Abstract

Purpose: High-resolution synchrotron radiation-based Fourier transform infrared (SR-FTIR) microspectroscopy coupled with multivariate analysis was used to investigate the characteristics of putative adult stem cell (SC), transiently amplified (TA) cell, and terminally differentiated (TD) cell populations of the corneal epithelium.

Methods: Spectra of individual cells in situ in cryosections of bovine cornea were collected by using a synchrotron microspectroscopy facility at Daresbury Laboratory (United Kingdom). The resultant spectra were analyzed by multivariate analysis.

Results: The median spectra of the three different cell populations showed marked differences, which correlated with their degree of differentiation and proliferative capacity. Multivariate (principal component) analysis (PCA) showed that the three cell populations could be segregated into discrete clusters, with only a slight overlap between the SC and TA cell populations. Further analysis (Mann-Whitney test) indicated that the most significant (P<0.001) spectral differences between the SC and TA cell populations were chiefly associated with changes in nucleic acid conformation.

Conclusions: SR-FTIR microspectroscopy coupled with PCA appears to enable the identification of SC, TA cell, and TD cell populations. The results also suggest that a small subpopulation of cells in the corneal epithelial SC niche possess TA cell-like characteristics. The most significant spectral characteristics associated with the SCs appear to relate to differences in nucleic acid conformation. This finding is consistent with recent theories suggesting that the control of differentiation is related to major changes in chromatin structure.