Abstract
Genetically encoded calcium indicators (GECIs) and high-resolution confocal microscopy enable dynamic visualization of calcium signals in cells and tissues. Two-dimensional and 3D biocompatible materials mimic the mechanical microenvironments of tumor and healthy tissues in a programmable manner. Cancer xenograft models and ex vivo functional imaging of tumor slices reveal physiologically relevant functions of calcium dynamics in tumors at different progression stages. Integration of these powerful techniques allows us to quantify, diagnose, model, and understand cancer pathobiology. Here, we describe detailed materials and methods used to establish this integrated interrogation platform, from generating transduced cancer cell lines that stably express CaViar (GCaMP5G + QuasAr2) to in vitro and ex vivo calcium imaging of the cells in 2D/3D hydrogels and tumor tissues. These tools open the possibility for detailed explorations of mechano-electro-chemical network dynamics in living systems.
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Acknowledgments
This project is funded by UF Gatorade Award Start-up Package (X. T.), UFHCC Cancer Pilot Award (X. T. and D. S.), UF Opportunity Seed Fund (X. T., H. K., R. C., and C. M.), NIH R35 GM 128753 (M. X.), and National Natural Science Foundation of China #81974093 (B. Z.). We genuinely appreciate the invaluable discussions with and technical support from Dr. Jonathan Licht (UFHCC), Dr. John Wingard (UFHCC), Dr. Rolf Renne (UFHCC), Dr. Thomas George (UFHCC), Dr. Adam Cohen (Harvard University), Dr. Tian He (Harvard University), Dr. Hongkang Zhang (Q-State), Dr. Christopher Werley (Vertex Pharmaceuticals), Dr. Mark Sheplak (MAE & ECE), Dr. Warren Dixon (MAE), Dr. Hugh Fan (MAE), Dr. Scott Banks (MAE), Dr. Ting Dong (MAE), Dr. Yong Huang (MAE), Dr. David Hahn (University of Arizona), Dr. Mike Stapleton (ECE), Dr. Walter Lee Murfee (BME), Dr. Habibeh Khoshbouei (Neuroscience), Dr. Min Lin (Neuroscience), Dr. Todd Golde (Neuroscience), Dr. Daniel Ryu (Neuroscience), Dr. Robert Caudle (Oral and Maxillofacial Surgery), Dr. John Neubert (Orthodontics), Dr. Justin Hilliard (Neurosurgery), Dr. Lance McMahon (Texas Tech University), Dr. Chritopher McCurdy (Pharmacology), Dr. Yenisel Cruz-Almeida (Pain Research and Intervention), Dr. Roger Fillingim (Pain Research and Intervention), Dr. Lizi Wu (Molecular Genetics and Microbiology), Dr. Yuqing Li (Neuroscience), Dr. Alice Harmon (Biology), Dr. Christopher Batich (MSE), Dr. Richard Dickson (ChE), Dr. Claudio Grosman (University of Illinois-UC), Dr. Ji-Hyun Lee (UFHCC), and Dr. Jeffrey Thinschmidt (Pharmacodynamics). We are sincerely grateful for the generous supports from all members of Tang’s, Zeng’s, Siemann’s, Sharma’s, Xie’s, Wu’s, and Phelps’ laboratories and all staff members of the MAE, BME, ECE, CHE, MSE, and UFHCC at UF..
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Liang, C. et al. (2023). Functional Interrogation of Ca2+ Signals in Human Cancer Cells In Vitro and Ex Vivo by Fluorescent Microscopy and Molecular Tools. In: Garcia-Cordero, J.L., Revzin, A. (eds) Microfluidic Systems for Cancer Diagnosis . Methods in Molecular Biology, vol 2679. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3271-0_7
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DOI: https://doi.org/10.1007/978-1-0716-3271-0_7
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