Two-photon imaging of endogenous biomolecules in freshly excised human cervical tissue biopsies captures pre-cancer induced changes in tissue metabolism and morphology

Christopher M. Polleys; Hong-Thao Thieu; Elizabeth M. Genega; Narges Jahanseir; Irene Georgakoudi

doi:10.1117/12.2650926

14 March 2023 Two-photon imaging of endogenous biomolecules in freshly excised human cervical tissue biopsies captures pre-cancer induced changes in tissue metabolism and morphology

Christopher M. Polleys, Hong-Thao Thieu, Elizabeth M. Genega, Narges Jahanseir, Irene Georgakoudi

Author Affiliations +

Proceedings Volume 12363, Multiscale Imaging and Spectroscopy IV; 1236302 (2023) https://doi.org/10.1117/12.2650926
Event: SPIE BiOS, 2023, San Francisco, California, United States

Abstract

Fiber endoscopes capable of making two-photon (2P) autofluorescence measurements, time-resolved fluorescence decay measurements, and collagen second harmonic generation (SHG) measurements have been applied to animal models. Clinical translation of such devices to internal human organs has the potential to overhaul conventional methods of disease diagnosis and monitoring. Previous work by our lab has established the potential to diagnose high-grade cervical precancers using 2P autofluorescence measurements. Other groups have demonstrated that 2P-based fluorescence lifetime imaging microscopy (FLIM) measurements of nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) and collagen SHG measurements have the potential to discriminate between cancerous and benign tissues. In this work, we demonstrate the potential to discern high-grade cervical precancerous lesions (HSILs) from benign tissues using fluorescence intensity measurements of NAD(P)H and oxidized flavoproteins, FLIM NAD(P)H measurements, and collagen SHG measurements. Consistent with previous results, benign tissues demonstrated increased depth-dependent heterogeneity in mitochondrial clustering, and increased overall and intrafield heterogeneity of oxido-reductive state relative to HSILs. FLIM phasor analysis demonstrated a relative decrease in NAD(P)H short and long lifetime, and a relative increase in NAD(P)H bound fraction for benign tissues compared to HSILs. Collagen SHG intensity in benign tissues was greater than that of HSIL tissues, along with overall intrafield variations in collagen fiber orientation. This work motivates the functionalization of a clinical 2P fiber endoscope capable of making SHG, autofluorescence intensity and lifetime measurements of metabolic coenzymes in the human cervix.

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Christopher M. Polleys, Hong-Thao Thieu, Elizabeth M. Genega, Narges Jahanseir, and Irene Georgakoudi "Two-photon imaging of endogenous biomolecules in freshly excised human cervical tissue biopsies captures pre-cancer induced changes in tissue metabolism and morphology", Proc. SPIE 12363, Multiscale Imaging and Spectroscopy IV, 1236302 (14 March 2023); https://doi.org/10.1117/12.2650926

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