A Novel Ex Vivo Method for Visualizing Live-Cell Calcium Response Behavior in Intact Human Tumors.

Koh, James

Hogue, Joyce A

Sosa, Julie A

United States

2017-08-28T16:39:45Z

2017-08-28T16:39:45Z

2016

The functional impact of intratumoral heterogeneity has been difficult to assess in the absence of a means to interrogate dynamic, live-cell biochemical events in the native tissue context of a human tumor. Conventional histological methods can reveal morphology and static biomarker expression patterns but do not provide a means to probe and evaluate tumor functional behavior and live-cell responsiveness to experimentally controlled stimuli. Here, we describe an approach that couples vibratome-mediated viable tissue sectioning with live-cell confocal microscopy imaging to visualize human parathyroid adenoma tumor cell responsiveness to extracellular calcium challenge. Tumor sections prepared as 300 micron-thick tissue slices retain viability throughout a >24 hour observation period and retain the native architecture of the parental tumor. Live-cell observation of biochemical signaling in response to extracellular calcium challenge in the intact tissue slices reveals discrete, heterogeneous kinetic waveform categories of calcium agonist reactivity within each tumor. Plotting the proportion of maximally responsive tumor cells as a function of calcium concentration yields a sigmoid dose-response curve with a calculated calcium EC50 value significantly elevated above published reference values for wild-type calcium-sensing receptor (CASR) sensitivity. Subsequent fixation and immunofluorescence analysis of the functionally evaluated tissue specimens allows alignment and mapping of the physical characteristics of individual cells within the tumor to specific calcium response behaviors. Evaluation of the relative abundance of intracellular PTH in tissue slices challenged with variable calcium concentrations demonstrates that production of the hormone can be dynamically manipulated ex vivo. The capability of visualizing live human tumor tissue behavior in response to experimentally controlled conditions opens a wide range of possibilities for personalized ex vivo therapeutic testing. This highly adaptable system provides a unique platform for live-cell ex vivo provocative testing of human tumor responsiveness to a range of physiological agonists or candidate therapeutic compounds.

https://www.ncbi.nlm.nih.gov/pubmed/27537691

PONE-D-16-16622

1932-6203

https://hdl.handle.net/10161/15385

eng

Public Library of Science (PLoS)

PLoS One

10.1371/journal.pone.0161134

Adenoma

Calcium

Humans

Microscopy, Confocal

Neoplasms

Parathyroid Glands

Parathyroid Hormone

Parathyroid Neoplasms

A Novel Ex Vivo Method for Visualizing Live-Cell Calcium Response Behavior in Intact Human Tumors.

Journal article

Sosa, Julie A|0000-0002-3356-7316

https://www.ncbi.nlm.nih.gov/pubmed/27537691

e0161134

8

Clinical Science Departments

Duke

Duke Cancer Institute

Duke Clinical Research Institute

Institutes and Centers

Medicine

Medicine, Medical Oncology

School of Medicine

Surgery

Surgery, Advanced Oncologic and Gastrointestinal Surgery

Surgery, Surgical Sciences

Published online

11