title: Staining and in vitro toxicity of dithizone with canine, porcine, and bovine islets.
abstract: Dithizone (DTZ) is a recognized diabetogenic agent in vivo, and a supravital stain commonly used for identification of islets to be used for transplantation. In the present studies, we compared DTZ staining of freshly isolated and cultured canine, bovine, and porcine islets, and the effect of DTZ on the function and viability of islets. Incubation with DTZ resulted in staining of canine and porcine islets, but no discernible staining with bovine islets. Insulin content of porcine, canine, and bovine islet was 2.0 +/- 0.2, 2.2 +/- 0.3, and 1.9 +/- 0.2 mU/EIN, indicating a lack of correspondence of DTZ staining and insulin content. Seven days of culture with canine islets resulted in > or = 50% reduction of DTZ stained cells. Exposure to DTZ at 50 micrograms/mL resulted in a maximal number of stained cells in preparations of purified islets (80-85%; counted after dispersion), a lower percentage of cells stained faintly at 20 micrograms/mL (50-55%), with no discernible staining at 10 micrograms/mL. Prolonged exposure of islets (4-48 h) to 20 micrograms/mL DTZ led to reduced insulin secretion and islet cell death. Incubation of canine or porcine islets with 100 micrograms/mL of DTZ for 0.5 h resulted in a dramatic loss of viability and diminished insulin secretory function, which was not reversed with continued culture. The concentration dependence of toxic effects paralleled the concentration dependence of cellular staining. The minimally effective staining concentration (20 micrograms/mL) also resulted in a loss of viability. An additional assessment of DTZ toxicity was made using the RIN-38 beta-cell line, which shows no discernible staining with DTZ.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: BioHybrid Technologies, Shrewsbury, MA 01545.
authors.forename: S A
authors.initials: SA
authors.lastname: Clark
authors.forename: K M
authors.initials: KM
authors.lastname: Borland
authors.forename: S D
authors.initials: SD
authors.lastname: Sherman
authors.forename: T C
authors.initials: TC
authors.lastname: Rusack
authors.forename: W L
authors.initials: WL
authors.lastname: Chick
mesh_terms: D000818:Animals; D000968:Antimycin A / Q000031:analogs & derivatives / Q000494:pharmacology; D002417:Cattle; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D004230:Dithizone / Q000633:toxicity*; D004285:Dogs; D004305:Dose-Response Relationship, Drug; D005947:Glucose / Q000494:pharmacology; D066298:In Vitro Techniques; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000187:drug effects* / Q000503:physiopathology; D013194:Staining and Labeling; D013552:Swine
chemical_list: D007328:Insulin; C032456:antimycin; D004230:Dithizone; D000968:Antimycin A; D005947:Glucose