Differential susceptibilities of murine hepatoma 1c1c7 and Tao cells to the lysosomal photosensitizer NPe6: influence of aryl hydrocarbon receptor on lysosomal fragility and protease contents.
Irradiation of murine hepatoma 1c1c7 cultures presensitized with N-aspartyl chlorin e6 (NPe6) caused lysosomal disruption and apoptosis. Tao cells, a variant of the 1c1c7 line having lower aryl hydrocarbon receptor (AhR) contents, were resistant to the pro-apoptotic effects of NPe6 in the same photodynamic therapy protocol. Colony-forming assays were used to establish light dose-dependent and NPe6 concentration-dependent cytotoxicity curves. Lysosomal breakage and cell survival paralleled one another in both cell types. When analyzed at comparable lethal dose conditions, the onset of apoptosis was delayed, and the magnitude of the apoptotic response was muted in Tao cells, as assessed by morphology, annexin V binding, caspase-3 activities, and analyses of Bid, procaspase-9, and pro-caspase-3 cleavage. In contrast, the kinetics/magnitude of pro-caspase-3 activation in the two cell lines were identical after exposure to HA14 -1 or Jo2 antibody, inducers of the intrinsic and extrinsic apoptotic pathways, respectively. Tao endosomal/lysosomal extracts contained approximately 50%, 35%, and 55% of the Bid cleavage and cathepsin B and D activities of 1c1c7 endosomes/lysosomes, respectively. Western blot analyses confirmed reduced cathepsin B/D contents in Tao cells. Analyses of 1c1c7/Tao variants engineered to express antisense/sense AhR constructs suggested that endosomal/lysosomal cathepsin B and D content, but not whole cell content, correlated with AhR expression. These studies provide a mechanism for the resistance of Tao cultures to the proapoptotic effects of a protocol causing targeted disruption of lysosomes. They also suggest that the AhR, in the absence of exogenous ligand, may affect the trafficking/processing of proteases normally found in endosomes/lysosomes.
Caruso
Joseph A
JA
Institute of Environmental Health Sciences, Wayne State University, Detroit, MI 48201, USA.
Mathieu
Patricia A
PA
Joiakim
Aby
A
Leeson
Brendan
B
Kessel
David
D
Sloane
Bonnie F
BF
Reiners
John J
JJ
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis; D051023:BH3 Interacting Domain Death Agonist Protein; D001578:Benzopyrans / Q000494:pharmacology; D006528:Carcinoma, Hepatocellular / Q000473:pathology; D002352:Carrier Proteins / Q000378:metabolism; D002403:Cathepsins / Q000378:metabolism; D008113:Liver Neoplasms / Q000473:pathology; D008247:Lysosomes / Q000187:drug effects* / Q000378:metabolism; D051379:Mice; D009570:Nitriles / Q000494:pharmacology; D017319:Photosensitizing Agents / Q000494:pharmacology*; D011166:Porphyrins / Q000494:pharmacology*; D018336:Receptors, Aryl Hydrocarbon / Q000502:physiology*; D014226:Triazenes / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D019014:fas Receptor / Q000494:pharmacology
D000970:Antineoplastic Agents; D051023:BH3 Interacting Domain Death Agonist Protein; D001578:Benzopyrans; C494815:Bid protein, mouse; D002352:Carrier Proteins; D009570:Nitriles; D017319:Photosensitizing Agents; D011166:Porphyrins; D018336:Receptors, Aryl Hydrocarbon; D014226:Triazenes; C456800:ethyl 2-amino-6-bromo-4-(1-cyano-2-ethoxy-2-oxoethyl)-4H-chromene-3-carboxylate; D019014:fas Receptor; C008848:1-phenyl-3,3-dimethyltriazene; D002403:Cathepsins; C053434:Talaporfin