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A genetic reporter of thermal stress defines physiologic zones over a defined temperature range

CAITLIN E. O’CONNELL-RODWELL, DAVID SHRIVER, DMITRI M. SIMANOVSKII^*, CAMERON MCCLURE, YU-AN CAO, WEISHENG ZHANG, MICHAEL H. BACHMANN, JOSHUA T. BECKHAM, E. DUCO JANSEN, DANIEL PALANKER^*, H. ALAN SCHWETTMAN^* and CHRISTOPHER H. CONTAG¹

Department of Pediatrics, Microbiology & Immunology and Radiology, Stanford School of Medicine, Stanford, California, USA;
^* Picosecond Free Electron Laser Center, W. W. Hansen Experimental Physics Laboratory, Stanford University, Stanford, California, USA;
Xenogen Corporation, Alameda, California, USA; and
Department of Biomedical Engineering, Vanderbilt University, Nashville, Tennessee, USA

¹ Correspondence: Department of Pediatrics, Stanford School of Medicine, 300 Pasteur Dr., Stanford, CA 94035-5208, USA. E-mail:ccontag{at}cmgm.stanford.edu

We define five unique cellular responses to thermal stress using a reporter construct generated using the stress-inducible promoter from the gene encoding a murine 70 kDa heat shock protein (Hsp70A.1) to express luciferase (luc). Thermal stress was delivered over a range of temperatures (42–68°C) for 5 s to 20 min and luciferase activity was measured in live cells using a cooled CCD camera as a measure of reporter gene transcription. Reporter gene expression was assessed every 2 h for 10 h, and at 24 h post-stress. Expression patterns were validated for selected temperatures. A transition zone where cells lose the ability to produce light and beyond which >50% of cells die was observed to occur within a narrow (2.5°C) temperature window. Although luc and hsp70 mRNA levels in this transition zone were high, there were reduced levels of Luc and Hsp70 protein and ATP levels. Cells treated at these temperatures recovered the ability to produce light in response to a secondary stress at 30 h. This Hsp70-luc reporter gene construct may be useful for defining zones of physiologic responses and assessing collateral thermal damage generated during treatment of biological tissue with lasers and other sources of heat.—O’Connell-Rodwell, C. E., Shriver, D., Simanovskii, D. M., McClure, C., Cao, Y-a., Zhang, W., Bachmann, M. H., Beckham, J. T., Jansen, E. D., Palanker, D., Schwettman, H. A., Contag, C. H. A genetic reporter of thermal stress defines physiologic zones over a defined temperature range.

Key Words: laser–tissue interactions • luciferase • Hsp70 • thermal stress

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