{"database":"biostudies-literature","file_versions":[],"scores":null,"additional":{"submitter":["Bess SN"],"funding":["Arkansas Integrative Metabolic Research Center","NCI NIH HHS","National Institutes of Health","NIGMS NIH HHS","Arkansas Biosciences Institute","National Science Foundation"],"pagination":["210-223"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/S-EPMC10959688"],"repository":["biostudies-literature"],"omics_type":["Unknown"],"volume":["53(2)"],"pubmed_abstract":["<h4>Background</h4>The immune system has evolved to detect foreign antigens and deliver coordinated responses, while minimizing \"friendly fire.\" Until recently, studies investigating the behavior of immune cells were limited to static <i>in vitro</i> measurements. Although static measurements allow for real-time imaging, results are often difficult to translate to an <i>in vivo</i> setting. Multiphoton microscopy is an emerging method to capture spatial information on subcellular events and characterize the local microenvironment. Previous studies have shown that multiphoton microscopy can monitor changes in single-cell macrophage heterogeneity during differentiation. Therefore, there is a need to use multiphoton microscopy to monitor molecular interactions during immunological activities like phagocytosis. Here we investigate the correlation between phagocytic function and changes in endogenous optical reporters during phagocytosis.<h4>Methods</h4><i>In vitro</i> autofluorescence imaging of nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD) was used to detect metabolic changes in macrophages during phagocytosis. More specifically, optical redox ratio, mean NADH fluorescence lifetime and ratio of free to protein-bound NADH were used to quantify changes in metabolism.<h4>Results</h4>Results show that IFN-<i>γ</i> (M1) macrophages showed decreased optical redox ratios and mean NADH lifetime while phagocytosing immunogenic cancer cells compared to metastatic cells. To validate phagocytic function, a fluorescence microscopy-based protocol using a pH-sensitive fluorescent probe was used. Results indicate that M0 and M1 macrophages show similar trends in phagocytic potential.<h4>Conclusion</h4>Overall, this work demonstrates that <i>in vitro</i> multiphoton imaging can be used to longitudinally track changes in phagocytosis and endogenous metabolic cofactors."],"journal":["Immunological investigations"],"pubmed_title":["Live-Cell Imaging Quantifies Changes in Function and Metabolic NADH Autofluorescence During Macrophage-Mediated Phagocytosis of Tumor Cells."],"pmcid":["PMC10959688"],"funding_grant_id":["P20 GM139768","5P20GM139768-02","CBET 1751554","R15 CA238861"],"pubmed_authors":["Bess SN","Muldoon TJ","Igoe MJ"],"additional_accession":[]},"is_claimable":false,"name":"Live-Cell Imaging Quantifies Changes in Function and Metabolic NADH Autofluorescence During Macrophage-Mediated Phagocytosis of Tumor Cells.","description":"<h4>Background</h4>The immune system has evolved to detect foreign antigens and deliver coordinated responses, while minimizing \"friendly fire.\" Until recently, studies investigating the behavior of immune cells were limited to static <i>in vitro</i> measurements. Although static measurements allow for real-time imaging, results are often difficult to translate to an <i>in vivo</i> setting. Multiphoton microscopy is an emerging method to capture spatial information on subcellular events and characterize the local microenvironment. Previous studies have shown that multiphoton microscopy can monitor changes in single-cell macrophage heterogeneity during differentiation. Therefore, there is a need to use multiphoton microscopy to monitor molecular interactions during immunological activities like phagocytosis. Here we investigate the correlation between phagocytic function and changes in endogenous optical reporters during phagocytosis.<h4>Methods</h4><i>In vitro</i> autofluorescence imaging of nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD) was used to detect metabolic changes in macrophages during phagocytosis. More specifically, optical redox ratio, mean NADH fluorescence lifetime and ratio of free to protein-bound NADH were used to quantify changes in metabolism.<h4>Results</h4>Results show that IFN-<i>γ</i> (M1) macrophages showed decreased optical redox ratios and mean NADH lifetime while phagocytosing immunogenic cancer cells compared to metastatic cells. To validate phagocytic function, a fluorescence microscopy-based protocol using a pH-sensitive fluorescent probe was used. Results indicate that M0 and M1 macrophages show similar trends in phagocytic potential.<h4>Conclusion</h4>Overall, this work demonstrates that <i>in vitro</i> multiphoton imaging can be used to longitudinally track changes in phagocytosis and endogenous metabolic cofactors.","dates":{"release":"2024-01-01T00:00:00Z","publication":"2024 Feb","modification":"2025-04-05T10:19:13.034Z","creation":"2025-04-05T10:19:13.034Z"},"accession":"S-EPMC10959688","cross_references":{"pubmed":["37999933"],"doi":["10.1080/08820139.2023.2284369"]}}