title: In Vivo nonlinear optical imaging of immune responses: tissue injury and infection.
abstract: In this study, we demonstrate a noninvasive imaging approach based on multimodal nonlinear optical microscopy to in vivo image the responses of immune cells (neutrophils) to the tissue injury and bacterial infection in a zebrafish model. Specifically, the second harmonic generation from myosin thick filaments in sarcomere enabled a clear visualization of the muscle injury and infection. Two-photon excited fluorescence was used to track the behavior of the neutrophils that were transgenically labeled by red fluorescent protein. The corresponding reduced nicotinamide adenine dinucleotide (NADH) two-photon excited fluorescence images revealed a detailed morphological transformation process of individual neutrophils during muscle tissue injury and bacterial infection. The analysis of time-resolved NADH signals from the neutrophils provided important biological insights of the cellular energy metabolism during the immune responses. We found a significant increase of free/protein-bound NADH ratios in activated neutrophils in bacterial-infected tissue. In this study, we also discovered that, under 720 nm excitation, two wild-type strains (DH5? and BL21) of bacteria Escherichia coli emitted distinct endogenous fluorescence of double-peak at ?450 and ?520 nm, respectively. We demonstrated that the double-peak fluorescence signal could be used to differentiate the E. coli from surrounding tissues of dominant NADH signals, and to achieve label-free tracking of E. coli bacteria in vivo.
authors.affiliation: Biophotonics Research Laboratory, Department of Electronic and Computer Engineering, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong SAR, P.R. China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Zeng
authors.affiliation: State Key Laboratory of Molecular Neuroscience, Division of Life Science, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong SAR, P.R. China.
authors.forename: Bo
authors.initials: B
authors.lastname: Yan
authors.affiliation: State Key Laboratory of Molecular Neuroscience, Division of Life Science, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong SAR, P.R. China.
authors.forename: Jin
authors.initials: J
authors.lastname: Xu
authors.affiliation: Biophotonics Research Laboratory, Department of Electronic and Computer Engineering, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong SAR, P.R. China.
authors.forename: Qiqi
authors.initials: Q
authors.lastname: Sun
authors.affiliation: Biophotonics Research Laboratory, Department of Electronic and Computer Engineering, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong SAR, P.R. China.
authors.forename: Sicong
authors.initials: S
authors.lastname: He
authors.affiliation: Biophotonics Research Laboratory, Department of Electronic and Computer Engineering, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong SAR, P.R. China.
authors.forename: Jun
authors.initials: J
authors.lastname: Jiang
authors.affiliation: State Key Laboratory of Molecular Neuroscience, Division of Life Science, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong SAR, P.R. China; Center of Systems Biology and Human Health, School of Science and Institute for Advanced Study, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong SAR, P.R. China. Electronic address: zilong@ust.hk.
authors.forename: Zilong
authors.initials: Z
authors.lastname: Wen
authors.affiliation: Biophotonics Research Laboratory, Department of Electronic and Computer Engineering, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong SAR, P.R. China; Center of Systems Biology and Human Health, School of Science and Institute for Advanced Study, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong SAR, P.R. China. Electronic address: eequ@ust.hk.
authors.forename: Jianan Y
authors.initials: JY
authors.lastname: Qu
mesh_terms: D000818:Animals; D004625:Embryo, Nonmammalian / Q000166:cytology / Q000276:immunology / Q000382:microbiology; D004734:Energy Metabolism / Q000276:immunology; D004926:Escherichia coli / Q000302:isolation & purification / Q000502:physiology; D007113:Immunity, Innate; D036641:Microscopy, Fluorescence, Multiphoton / Q000379:methods*; D018482:Muscle, Skeletal / Q000166:cytology / Q000276:immunology; D009218:Myosins / Q000378:metabolism; D009243:NAD / Q000378:metabolism; D009504:Neutrophils / Q000276:immunology / Q000378:metabolism; D015398:Signal Transduction / Q000276:immunology; D015027:Zebrafish / Q000276:immunology / Q000382:microbiology
chemical_list: D009243:NAD; D009218:Myosins