A genetic reporter of thermal stress defines physiologic zones over a defined temperature range.
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 degrees 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 degrees 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
Caitlin E
CE
Department of Pediatrics, Stanford School of Medicine, Stanford, California 94035-5208, USA.
Shriver
David
D
Simanovskii
Dmitri M
DM
McClure
Cameron
C
Cao
Yu-An
YA
Zhang
Weisheng
W
Bachmann
Michael H
MH
Beckham
Joshua T
JT
Jansen
E Duco
ED
Palanker
Daniel
D
Schwettman
H Alan
HA
Contag
Christopher H
CH
D000818:Animals; D015153:Blotting, Western; D002470:Cell Survival; D005434:Flow Cytometry; D005786:Gene Expression Regulation / Q000528:radiation effects*; D017930:Genes, Reporter / Q000235:genetics*; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics; D006358:Hot Temperature; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D051379:Mice; D041681:NIH 3T3 Cells; D011401:Promoter Regions, Genetic / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013696:Temperature; D013997:Time Factors
D018840:HSP70 Heat-Shock Proteins; D012333:RNA, Messenger; D008156:Luciferases