Raman Spectroscopy Analysis of the Retina Reveals Changes in Glutamate, N-Acetyl-Aspartate, NADH and Phosphatidylcholine in Neuroinflammation (P01.166)

Objective: To assess the usefulness of Raman spectroscopy (RS) of the retina for monitoring molecular changes during neuroinflammation. Background Development of molecular imaging techniques will allow assessing molecular changes during disease course. RS offers this promise by providing such information from the retina. Design/Methods: RS was used to create a spectral library of eight pivotal biomolecules known to be involved in neuroinflammation; Nicotinamide Adenine Dinucliotide (NADH), Flavin Adenine Nucleotide (FAD), Lactate, Cytochrome C, Glutamate, N-Acetyl-Aspartate (NAA), Phosphotidylcholine, with Advanced Glycolization End Products (AGEs) analyzed as a reference. We obtained RS from mice retina organotypic cultures with and without stimulation with lipopolysaccharide (LPS) for inducing neuroinflammation. Results: Glutamate, Lactate, NAA, NADH, and Phosphotidylcholine yielded strong peaks in solution and were chosen for further analysis. These five molecular spectra were used to tentatively characterize the Raman peaks of spectra taken in the ganglion cell layer of murine retina organotypic culture. Neuroinflammation was induced by adding lypopolysacharide (LPS) to cultures. The loading plots of the Principal Component Analysis (PCA) of 50 spectra taken of murine retinal tissue culture undergoing neuroifnlamation and healthy controls were analyzed in order to characterize the molecular makeup of the inflammation. The loading plots revealed the influence of peaks related to Glutamate, NAA, NADH, and Phosphotidylcholine to neuroinflammation-related spectral changes, with each molecule yielding 2-3 highly varying peaks within the retinal spectra. Partial Least Squares- Discriminant analysis (PLS-DA) was performed to create a multivariate classifier for the spectral diagnosis of neuroinflammed tissue, which was validated using venetian blinds w/6 splits with high accuracy. Conclusions: we provide a proof of concept that Raman spectrosocpy is abl to identify changes in retina metabolites during inflammation, becoming a promissing technology for molecular imaging of the retina for the study of brain diseases. Supported by: Spanish Ministry of Science - Instituto de Salud Carlos III: Red Esapnola de Esclerosis Multiple. Disclosure: Dr. Villoslada has received personal compensation for activities with Roche Diagnostics Corporation, Novartis, MedImmune, Neurotek, Bionure, an Digna Biotech. Dr. Villoslada has received research support from Digna Biotech. Dr. Petrov has nothing to disclose. Dr. Taubes has nothing to disclose. Dr. Moreno has nothing to disclose. Dr. Petrov has nothing to disclose.