Dataset Information

In Vivo Visualization of Stromal Macrophages via label-free FLIM-based metabolite imaging.

ABSTRACT: Macrophage infiltration and recruitment in breast tumors has been correlated with poor prognosis in breast cancer patients and has been linked to tumor cell dissemination. Much of our understanding comes from animal models in which macrophages are labeled by expression of an extrinsic fluorophore. However, conventional extrinsic fluorescence labeling approaches are not readily applied to human tissue and clinical use. We report a novel strategy that exploits endogenous fluorescence from the metabolic co-factors NADH and FAD with quantitation from Fluorescence Lifetime Imaging Microscopy (FLIM) as a means to non-invasively identify tumor-associated macrophages in the intact mammary tumor microenvironment. Macrophages were FAD(HI) and demonstrated a glycolytic-like NADH-FLIM signature that was readily separated from the intrinsic fluorescence signature of tumor cells. This non-invasive quantitative technique provides a unique ability to discern specific cell types based upon their metabolic signatures without the use of exogenous fluorescent labels. Not only does this provide high resolution temporal and spatial views of macrophages in live animal breast cancer models, this approach can be extended to other animal disease models where macrophages are implicated and has potential for clinical applications.

SUBMITTER: Szulczewski JM

PROVIDER: S-EPMC4879594 | biostudies-literature | 2016 May

REPOSITORIES: biostudies-literature

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In Vivo Visualization of Stromal Macrophages via label-free FLIM-based metabolite imaging.

Szulczewski Joseph M JM Inman David R DR Entenberg David D Ponik Suzanne M SM Aguirre-Ghiso Julio J Castracane James J Condeelis John J Eliceiri Kevin W KW Keely Patricia J PJ

Scientific reports 20160525

Macrophage infiltration and recruitment in breast tumors has been correlated with poor prognosis in breast cancer patients and has been linked to tumor cell dissemination. Much of our understanding comes from animal models in which macrophages are labeled by expression of an extrinsic fluorophore. However, conventional extrinsic fluorescence labeling approaches are not readily applied to human tissue and clinical use. We report a novel strategy that exploits endogenous fluorescence from the meta ...[more]

PMID: 27220760

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Project description:BackgroundFlesh lignification, leading to increased fruit firmness, has been reported in several kinds of fruit. Understanding the mechanisms underlying fruit lignification is important to optimize the postharvest storage strategies and reduce the quality deterioration of postharvest fruit. Especially cellular level investigation of lignin deposition in fruits provides novel insight for deciphering the mechanisms underlying fruit lignification. The primary objective of this study was to establish a procedure of using Raman microspectroscopy technique to depict fruit lignification at the cell level.ResultsLignified cells, a special kind of cells contained high lignin content, were found abundantly scattered in red-fleshed 'Luoyangqing' loquat. Whereas these special lignified cells were barely detected in 'Baisha' loquat flesh. Dominant Raman bands of lignified cells were found primarily attributed to lignin (1664, 1628, 1603, 1467, and 1272 cm-1), cellulose (1383, 1124 and 1098 cm-1) and pectin (852 and 1740 cm-1). The band intensity correlation analysis indicated the peak at 1335 cm-1 assigned to either lignin or cellulose in previous works was related to lignin for the lignified cells. Multi-peaks Gaussian fitting successfully resolved the overlapped fingerprint peaks of lignin in 1550-1700 cm-1 into three independent peaks, which were assigned to different functional groups of lignin. Furthermore, the spatially resolved Raman images of lignified cells were generated, indicating that lignin and cellulose saturated the whole lignified cells, pectin mainly located in the cell corner, and the parenchyma cells contained little lignin. In addition, both phloroglucinol-HCl staining and autofluorescence analysis confirmed the results of lignin distribution of Raman microscopic analysis.ConclusionsA procedure for the simultaneous visualization of the main components of the flesh cells without labeling by high-resolution Raman microspectroscopy has been established. With Raman microscopic imaging technique, we can add a microscopic level to cell compositions, essential for a detailed molecular understanding of loquat lignification. Such method can be further used to chemically monitor the textural changes during the ripening process or postharvest storage of other fruits and vegetables.

Project description:The detection of cancer micrometastasis for early diagnosis and treatment poses a great challenge for conventional imaging techniques. The aim of our study was to evaluate the performance of photoacoustic imaging (PAI) in detecting hepatic micrometastases from melanoma at a very early stage and in aiding tumor resection by intraoperative guidance. Methods: In vivo studies were performed by following protocols approved by the Ethical Committee for Animal Research at Xiamen University. First, a mouse model of B16 melanoma metastatic to the liver (n = 10) was established to study the development of micrometastases in vivo. Next, the mice were imaged by a scalable PAI instrument, ultrasound, 9.4-T high-resolution MRI, PET/CT, and bioluminescence imaging. PAI scans acquired with optical wavelengths of 680-850 nm were kept spectrally unmixed by using a linear least-squares method to differentiate various components. Differences in signal-to-background ratios among different modalities were determined with the 2-tailed paired t test. The diagnostic results were assessed with histologic examination. Excised liver samples from patients diagnosed with hepatic cancer were also examined to identify the tumor boundaries. Surgical removal of metastatic melanoma was precisely guided in vivo by the portable PAI system. Results: PAI was able to detect metastases as small as approximately 400 ?m at a depth of up to 7 mm in vivo-a size that is smaller than can be detected with ultrasound and MRI. The tumor-to-liver ratio for PAI at 8 d (4.2 ± 0.2, n = 6) and 14 d (9.2 ± 0.4, n = 5) was significantly higher than for PET/CT (1.8 ± 0.1, n = 5, and 4.5 ± 0.2, n = 5, respectively; P < 0.001 for both). Functional PAI revealed dynamic oxygen saturation changes during tumor growth. The limit of detection was approximately 219 cells/?L in vitro. We successfully performed intraoperative PAI-guided surgery in vivo using the portable PAI system. Conclusion: Our findings offer a rapid and effective complementary clinical imaging application to noninvasively detect micrometastases and guide intraoperative resection.

Dataset Information

In Vivo Visualization of Stromal Macrophages via label-free FLIM-based metabolite imaging.

Publications

In Vivo Visualization of Stromal Macrophages via label-free FLIM-based metabolite imaging.

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