Activation of heat shock protein 70 promoter with meso-tetrahydroxyphenyl chlorin photodynamic therapy reported by green fluorescent protein in vitro and in vivo.
Cellular responses to photodynamic therapy (PDT) include induction of heat shock proteins (HSP). We examined meso-tetrahydroxyphenyl chlorin (mTHPC) PDT-mediated HSP activation in EMT6 cells stably transfected with a plasmid containing the gene for green fluorescent protein (GFP) driven by an hsp70 promoter. mTHPC incubation induced concentration-dependent GFP expression. Irradiation of cells exposed to a sensitizer concentration that induced a slight increase in GFP and no loss of cell viability resulted in fluence-dependent GFP accumulation. In response to drug only and to PDT, GFP levels increased to a maximum of four- to five-fold above control levels with increasing drug or fluence and then decreased at higher doses. A trypan blue-exclusion assay confirmed that decreased GFP levels in both cases were due to a loss of cell viability. For initial evaluation in vivo, HSP70/ GFP-transfected EMT6 tumors were grown in BALB/c mice and subjected to mTHPC-PDT with a fluence of 1 J/cm2. Six hours after PDT, GFP fluorescence was imaged in these tumors through the intact skin in vivo. These results indicate that sublethal doses of mTHPC-PDT stimulate GFP expression under the control of an hsp70 promoter and illustrate the potential of noninvasively monitoring reporter protein fluorescence as a measure of molecular response to PDT.
Mitra
Soumya
S
Department of Biochemistry and Biophysics, University of Rochester, Rochester, NY 14642, USA.
Goren
Evan M
EM
Frelinger
John G
JG
Foster
Thomas H
TH
D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects / Q000528:radiation effects; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation / Q000187:drug effects; D049452:Green Fluorescent Proteins; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis / Q000235:genetics*; D008027:Light; D008164:Luminescent Proteins; D008652:Mesoporphyrins / Q000494:pharmacology*; D051379:Mice; D010778:Photochemotherapy / Q000379:methods*; D011401:Promoter Regions, Genetic / Q000187:drug effects
D018840:HSP70 Heat-Shock Proteins; D008164:Luminescent Proteins; D008652:Mesoporphyrins; D049452:Green Fluorescent Proteins; C072269:temoporfin