title: Induction of apoptosis by cationic amphiphilic drugs amiodarone and imipramine.
abstract: Phospholipidosis is the excessive accumulation of intracellular phospholipids in cell lysosomes. Drugs that induce this disease often share common physiochemical properties and are collectively classified as cationic amphiphilic drugs (CADs). Although the cause of phospholipidosis and morphologic appearance of affected lysosomes have been studied extensively, less is known about the physiologic effects of the condition. In the current study, U-937 cells were incubated with the CADs amiodarone (2.5-10 microg/mL) and imipramine (2.5-20 microg/mL). Treatment of U-937 cells with these compounds for 96 h resulted in concentration-related increases in phospholipids, as assessed by flow cytometry using the fluorophore nile red. These results were verified by measuring the concentrations of choline-derived phospholipids, which were significantly increased in drug-treated cells. Cell number in amiodarone (10 microg/mL) and imipramine (20 microg/mL) cultures following the 96-h incubation period were markedly reduced compared to control cultures. These observations suggested that accumulation of cellular phospholipids could inhibit cell proliferation. Flow cytometric analysis revealed a decrease in the percentage of cells in the S-phase of the cell cycle following drug treatment, yet DNA replication still occurred in a significant portion of cells. Interestingly, amiodarone and imipramine induced apoptosis in U-937 cells as shown by annexin V-FITC staining and DNA fragmentation. Enzymatic assays demonstrated that amiodarone and imipramine induced the activity of caspases 2 and 3. These results suggest that disruption of cell lysosomes in U-937 cells following accumulation of phospholipids does not cause a cell cycle arrest but instead induces apoptosis by activation of caspase pathways.
authors.affiliation: Worldwide Safety Sciences, Pfizer Global Research and Development, Ann Arbor, Michigan 48105, USA.
authors.forename: Joseph R
authors.initials: JR
authors.lastname: Piccotti
authors.forename: Mark S
authors.initials: MS
authors.lastname: LaGattuta
authors.forename: Stephanie A
authors.initials: SA
authors.lastname: Knight
authors.forename: Andrea J
authors.initials: AJ
authors.lastname: Gonzales
authors.forename: Michael R
authors.initials: MR
authors.lastname: Bleavins
mesh_terms: D000638:Amiodarone / Q000633:toxicity*; D017304:Annexin A5 / Q000378:metabolism; D000889:Anti-Arrhythmia Agents / Q000633:toxicity*; D017209:Apoptosis / Q000187:drug effects*; D020169:Caspases / Q000096:biosynthesis; D002470:Cell Survival / Q000187:drug effects; D053938:DNA Fragmentation; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate; D006801:Humans; D007099:Imipramine / Q000633:toxicity*; D010743:Phospholipids / Q000378:metabolism; D013194:Staining and Labeling; D013501:Surface-Active Agents / Q000633:toxicity*; D020298:U937 Cells / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology
chemical_list: D017304:Annexin A5; D000889:Anti-Arrhythmia Agents; D010743:Phospholipids; D013501:Surface-Active Agents; D020169:Caspases; D016650:Fluorescein-5-isothiocyanate; D000638:Amiodarone; D007099:Imipramine