title: Photodynamic treatment of Chaoborus crystallinus larvae with chlorophyllin induces necrosis and apoptosis.
abstract: Chlorophyllin kills mosquito larvae (Culex, Aedes) in the aquatic habitat at low concentrations via photodynamic reactions under irradiation. The effects of chlorophyllin were investigated at the cellular level using the transparent larvae of Chaoborus crystallinus as a model system. Their transparency enabled in situ fluorescence investigation, showing that chlorophyllin accumulates in the intestine of the larvae. Uptake of chlorophyllin at room temperature took about 2 h. The fluorescence signal peaked after 5 h of incubation. Chlorophyllin accumulates up to about 15 ng per larvae. The intestine of treated larvae was dissected and stained with several dyes (acridine orange, Hoechst 33342 and propidium iodide). Apoptosis and necrosis increased with higher concentrations of chlorophyllin (to a smaller extent in dark controls) and were elevated in irradiated samples. Single cells from treated larvae were isolated and subjected to Annexin V flow cytometry. The fraction of apoptotic and necrotic cells increased significantly at a high chlorophyllin concentration (21.4 mg L(-1)) and under intensive irradiation. The activity of caspases-3, -8 and -9 as well as Bcl-2 and cytochrome c was investigated by means of western blot analysis. The data suggest a possible chlorophyllin concentration-dependent shift of the apoptotic pathway.
authors.affiliation: Bavarian State Research Centre for Agriculture, Institute for Fisheries, Starnberg, Germany.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Wohllebe
authors.forename: Claudia
authors.initials: C
authors.lastname: Ulbrich
authors.forename: Daniela
authors.initials: D
authors.lastname: Grimm
authors.forename: Jessica
authors.initials: J
authors.lastname: Pietsch
authors.forename: Gilmar
authors.initials: G
authors.lastname: Erzinger
authors.forename: Roland
authors.initials: R
authors.lastname: Richter
authors.forename: Michael
authors.initials: M
authors.lastname: Lebert
authors.forename: Peter Rolf
authors.initials: PR
authors.lastname: Richter
authors.forename: Donat-Peter
authors.initials: DP
authors.lastname: Häder
mesh_terms: D000330:Aedes; D000818:Animals; D017304:Annexin A5 / Q000032:analysis; D017209:Apoptosis; D015153:Blotting, Western; D020169:Caspases / Q000235:genetics / Q000378:metabolism; D002735:Chlorophyllides / Q000494:pharmacology*; D003465:Culex; D045304:Cytochromes c / Q000235:genetics / Q000378:metabolism; D004175:Diptera / Q000187:drug effects / Q000528:radiation effects; D005434:Flow Cytometry; D005456:Fluorescent Dyes / Q000032:analysis; D015870:Gene Expression / Q000187:drug effects / Q000528:radiation effects; D020287:In Situ Nick-End Labeling; D007422:Intestines / Q000187:drug effects / Q000528:radiation effects; D007814:Larva / Q000187:drug effects / Q000528:radiation effects; D009336:Necrosis; D017319:Photosensitizing Agents / Q000494:pharmacology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D014466:Ultraviolet Rays
chemical_list: D017304:Annexin A5; D002735:Chlorophyllides; D005456:Fluorescent Dyes; D017319:Photosensitizing Agents; D019253:Proto-Oncogene Proteins c-bcl-2; D045304:Cytochromes c; D020169:Caspases; C007020:chlorophyllin