Immune cells, including macrophages and T cells, have a range of cytotoxic and immune-modulating functions depending on activation state and subtype. However, current methods to assess immune cell function use exogenous labels that are limiting for time-course studies of immune cell behavior in tumors. Label-free optical imaging is an attractive solution. Here, we use multiphoton imaging of NAD(P)H and FAD, co-enzymes of metabolism, to monitor the function of T cells and macrophages. T cells were isolated from the peripheral blood of human donors, polarized to functional subsets, and identity was confirmed with standard surface markers. Separately, human macrophages were co-cultured with patient-derived invasive breast cancer cells in a 3D microdevice to monitor metabolic changes with tumor-stimulated macrophage migration. NAD(P)H and FAD fluorescence intensity and lifetime were monitored on a single cell level in both systems using multiphoton microscopy. Significant differences in autofluorescence were observed between functional T cell subsets including naïve vs. activated, helper vs. cytotoxic, and naïve vs. memory. Macrophages that actively migrated to the tumor cells in the 3D microfluidic model showed increased NAD(P)H/FAD intensity (optical redox ratio) compared to passively-migrating macrophages. Studies of in vivo T cell and macrophage metabolism using mouse models of cancer are underway. Altogether, these results indicate that multiphoton imaging of NAD(P)H and FAD is a powerful method for label-free, non-destructive monitoring of immune cell metabolism within single cells in the tumor microenvironment. These methods could inform new immunotherapy approaches for cancer.
Melissa Skala received her BS in Physics at Washington State University in 2002, her M.S. in Biomedical Engineering at the University of Wisconsin, Madison in 2004, and her Ph.D. in Biomedical Engineering at Duke University in 2007. Her postdoctoral training was also in Biomedical Engineering at Duke University, from 2007-2010. From 2010-2016, she was an Assistant Professor of Biomedical Engineering at Vanderbilt University. Since 2016 she has been an Investigator at the Morgridge Institute for Research. She is also a Professor of Biomedical Engineering at the University of Wisconsin – Madison. Her lab focuses on developing biomedical optical imaging technologies, personalized cancer treatment technologies, and more effective cancer therapies.