In Vivo Flow Cytometry Combined with Confocal Microscopy to Study Cancer Metastasis

Circulating tumor cells (CTCs) are of great interest in cancer research, but methods for their enumeration remain far from optimal. We developed a new small animal research tool called Diffuse in vivo Flow Cytometry (DiFC) for detecting extremely rare fluorescently- labeled circulating cells directly in the bloodstream. The technique exploits near-infrared diffuse photons to detect and count cells flowing in large superficial arteries and veins without drawing blood samples. DiFC uses custom-designed, dual fiber optic probes that are placed in contact with the skin surface approximately above a major vascular bundle. In combination with a novel signal processing, algorithm DiFC allows counting of individual cells moving in arterial or venous directions, as well as measurement of their speed and depth. We show that DiFC allows sampling of the entire circulating blood volume of a mouse in under 10 minutes, while maintaining a false alarm rate of 0.014 per minute. Hence, the unique capabilities of DiFC are highly suited to biological applications involving very rare cell types such as the study of hematogenic cancer metastasis.

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In vivo flow cytometry reveals a circadian rhythm of circulating tumor cells

Light Science & Applications ◽

10.1038/s41377-021-00542-5 ◽

2021 ◽

Vol 10 (1) ◽

Author(s):

Xi Zhu ◽

Yuanzhen Suo ◽

Yuting Fu ◽

Fuli Zhang ◽

Nan Ding ◽

...

Keyword(s):

Flow Cytometry ◽

Circadian Rhythm ◽

Tumor Cells ◽

Circulating Tumor Cells ◽

Cancer Progression ◽

Cancer Metastasis ◽

Detection Methods ◽

Orthotopic Mouse Model ◽

In Vivo Flow Cytometry

AbstractCirculating tumor cells (CTCs) is an established biomarker of cancer metastasis. The circulation dynamics of CTCs are important for understanding the mechanisms underlying tumor cell dissemination. Although studies have revealed that the circadian rhythm may disrupt the growth of tumors, it is generally unclear whether the circadian rhythm controls the release of CTCs. In clinical examinations, the current in vitro methods for detecting CTCs in blood samples are based on a fundamental assumption that CTC counts in the peripheral blood do not change significantly over time, which is being challenged by recent studies. Since it is not practical to draw blood from patients repeatedly, a feasible strategy to investigate the circadian rhythm of CTCs is to monitor them by in vivo detection methods. Fluorescence in vivo flow cytometry (IVFC) is a powerful optical technique that is able to detect fluorescent circulating cells directly in living animals in a noninvasive manner over a long period of time. In this study, we applied fluorescence IVFC to monitor CTCs noninvasively in an orthotopic mouse model of human prostate cancer. We observed that CTCs exhibited stochastic bursts over cancer progression. The probability of the bursting activity was higher at early stages than at late stages. We longitudinally monitored CTCs over a 24-h period, and our results revealed striking daily oscillations in CTC counts that peaked at the onset of the night (active phase for rodents), suggesting that the release of CTCs might be regulated by the circadian rhythm.

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Imaging Cardiovascular and Lung Macrophages With the Positron Emission Tomography Sensor 64 Cu-Macrin in Mice, Rabbits, and Pigs

Circulation Cardiovascular Imaging ◽

10.1161/circimaging.120.010586 ◽

2020 ◽

Vol 13 (10) ◽

Cited By ~ 1

Author(s):

Matthias Nahrendorf ◽

Friedrich Felix Hoyer ◽

Anu E. Meerwaldt ◽

Mandy M.T. van Leent ◽

Max L. Senders ◽

...

Keyword(s):

Positron Emission Tomography ◽

Flow Cytometry ◽

Confocal Microscopy ◽

Pet Imaging ◽

Near Infrared ◽

Emission Tomography ◽

Imaging Biomarker ◽

Positron Emission ◽

Pulmonary Macrophages

Background: Macrophages, innate immune cells that reside in all organs, defend the host against infection and injury. In the heart and vasculature, inflammatory macrophages also enhance tissue damage and propel cardiovascular diseases. Methods: We here use in vivo positron emission tomography (PET) imaging, flow cytometry, and confocal microscopy to evaluate quantitative noninvasive assessment of cardiac, arterial, and pulmonary macrophages using the nanotracer 64 Cu-Macrin—a 20-nm spherical dextran nanoparticle assembled from nontoxic polyglucose. Results: PET imaging using 64 Cu-Macrin faithfully reported accumulation of macrophages in the heart and lung of mice with myocardial infarction, sepsis, or pneumonia. Flow cytometry and confocal microscopy detected the near-infrared fluorescent version of the nanoparticle ( VT680 Macrin) primarily in tissue macrophages. In 5-day-old mice, 64 Cu-Macrin PET imaging quantified physiologically more numerous cardiac macrophages. Upon intravenous administration of 64 Cu-Macrin in rabbits and pigs, we detected heightened macrophage numbers in the infarcted myocardium, inflamed lung regions, and atherosclerotic plaques using a clinical PET/magnetic resonance imaging scanner. Toxicity studies in rats and human dosimetry estimates suggest that 64 Cu-Macrin is safe for use in humans. Conclusions: Taken together, these results indicate 64 Cu-Macrin could serve as a facile PET nanotracer to survey spatiotemporal macrophage dynamics during various physiological and pathological conditions. 64 Cu-Macrin PET imaging could stage inflammatory cardiovascular disease activity, assist disease management, and serve as an imaging biomarker for emerging macrophage-targeted therapeutics.

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