Multiphoton microscopy to characterize macrophage function and collagen alignment

Linghao Hu; Dhavan Sharma; Te-An Chen; Feng Zhao; Alex Walsh

doi:10.1117/12.2619396

7 March 2022 Multiphoton microscopy to characterize macrophage function and collagen alignment

Linghao Hu, Dhavan Sharma, Te-An Chen, Feng Zhao, Alex Walsh

Proceedings Volume PC11965, Multiphoton Microscopy in the Biomedical Sciences XXII; PC119650K (2022) https://doi.org/10.1117/12.2619396
Event: SPIE BiOS, 2022, San Francisco, California, United States

Conference Poster

Abstract

The human skin dermis is vascularized soft tissue containing highly organized extracellular matrix (ECM) architecture. Macrophages mediate wound healing within the skin but technologies for detection of macrophage function simultaneously with ECM properties are limited. In this study, we use multiphoton microscopy to evaluate the metabolic function of macrophages and quantify collagen alignment. Multiphoton microscopy is label-free and non-damaging allowing time-course studies of turbid samples and in vivo measurements. The fluorescence lifetime imaging technique detects the time fluorophores remain in an excited state before relaxing to the ground state. Reduced nicotinamide dinucleotide (NADH) and oxidized flavin adenine dinucleotide (FAD) are auto-fluorescence coenzymes that participate in various metabolic pathways including glycolysis, oxidative phosphorylation, fatty acid synthesis, and fatty acid oxidation, and label-free fluorescence lifetime imaging is a functional method to track cellular metabolism. In this paper, macrophages were seeded on anisotropic and isotropic ECM. Fluorescence lifetime images of macrophages and second harmonic generation (SHG) images of collagen in the ECM were obtained with a two-photon fluorescence lifetime microscope every two days for one week. Significant changes in macrophage size and fluorescence lifetime features were observed with time. The free fraction of NAD(P)H and bound fraction of FAD are both higher for macrophages on random ECM compared with aligned collagen. In summary, multiphoton microscopy provides a label-free method to track metabolic variation of macrophages concurrently with imaging of the ECM.

Conference Presentation

Citation Download Citation

Linghao Hu, Dhavan Sharma, Te-An Chen, Feng Zhao, and Alex Walsh "Multiphoton microscopy to characterize macrophage function and collagen alignment", Proc. SPIE PC11965, Multiphoton Microscopy in the Biomedical Sciences XXII, PC119650K (7 March 2022); https://doi.org/10.1117/12.2619396

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