Room: Exhibit Hall | Forum 1
Purpose: Raman spectroscopy measures the change in the energy of scattered light compared to the excitation photons to quantify molecular changes. In this study, Raman spectroscopy was used to investigate molecular changes in mice following radiation-induced pulmonary fibrosis.
Methods: Fourteen C57BL/6J male mice were investigated. The whole lungs of seven mice were irradiated with 15 Gy in a single fraction using 6 MV photons. Seven additional mice were used as controls. At specified times after irradiation, 48 hour (3 irradiated and 3 control mice) and 6 months (4 irradiated and 4 control mice), lungs were excised, flash frozen, and sectioned. Alternating sections were either stained with hematoxylin and eosin and evaluated by a pathologist or unstained for Raman measurements. Raman data were acquired from areas of alveolar wall thickening or areas near hyaline membranes (48 hr) and severe distortion of structure (due, for example, to fibrosis at 6 months), and compared with that from normal lung parenchyma. A total of 244 Raman spectra were acquired and analyzed using principal component analysis (PCA), discriminant function analysis (DFA) and Raman intensity ratio metric method.
Results: PCA captured 95% of the variance in the Raman spectral features. PCA/DFA analysis showed severe distortion (fibrosis) was identifiable 92% of the time. Acute injury (48 hr) compared to normal lung parenchyma was identified with 100% sensitivity and specificity. A minor alveolar septal thickening in normal controls was noted between spectra collected at 48 hr and 6 month time points. Consistent with fibrosis, multiple Raman bands (850, 1002, 1309, 1448, 1656 cm-1) assignable to proteins and lipids increased whereas others (1223 and 1578 cm-1) assignable to nucleic acids decreased. Consistent with acute damage, increased Raman intensity ratio of 1448/1002 and 1658/1002 (protein) was observed.
Conclusion: Preliminary findings support the use of Raman spectroscopy to characterize radiation-induced fibrosis.
Funding Support, Disclosures, and Conflict of Interest: Work supported by Henry Ford Health System internal grant, HFCI/CRAG pilot funding