title: 1-amino-4-phenyl-1,2,3,6-tetrahydropyridine and 1-amino-4-phenylpyridinium salt, the 1-amino analogues of neurotoxins, MPTP and MPP+, induce apoptosis in PC12 cells: detection of apoptotic cells by Comet assay and flow cytometric analysis.
abstract: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is known to induce parkinsonism in humans when it is oxidized to the 1-methyl-4-phenylpyridinium salt (MPP+). We previously reported the syntheses of 1-amino-4-phenyl-1,2,3,6-tetrahydropyridine (APTP) and 1-amino-4-phenyl-pyridinium salt (APP+), the 1-amino analogues of the dopaminergic neurotoxins, MPTP and MPP+, respectively, and demonstrated that both APTP and APP+ are cytotoxic to PC12 cells. In this study, we found that both APTP and APP+ induce apoptotic cell death in PC12 cells. Apoptosis was determined by the Comet assay and flow cytometric analysis. Prior to using the Comet assay for detection of apoptotic PC12 cells, Comet images of apoptotic and necrotic cells were first distinguished by using several standards. Comet images were classified into four groups (A to D) according to their shapes. Class D consisted of the apoptotic cells and was easily distinguished. We also demonstrated that apoptotic and necrotic PC12 cells can be easily differentiated and quantified using the convenient Comet assay.
authors.affiliation: Graduate School of Pharmaceutical Sciences, Nagoya City University, Tanabedori, Mizuho-ku, Nagoya 467-8603, Japan.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Noda
authors.forename: Tomoe
authors.initials: T
authors.lastname: Sumino
authors.forename: Yuki
authors.initials: Y
authors.lastname: Fujisawa
authors.forename: Naoki
authors.initials: N
authors.lastname: Miyata
authors.forename: Toyo
authors.initials: T
authors.lastname: Kaiya
authors.forename: Kohfuku
authors.initials: K
authors.lastname: Kohda
mesh_terms: D015632:1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D015655:1-Methyl-4-phenylpyridinium / Q000031:analogs & derivatives / Q000494:pharmacology*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D020552:Comet Assay; D004249:DNA Damage; D053938:DNA Fragmentation; D004273:DNA, Neoplasm / Q000032:analysis / Q000187:drug effects; D005434:Flow Cytometry; D016716:PC12 Cells / Q000187:drug effects* / Q000473:pathology; D011726:Pyridinium Compounds; D051381:Rats
chemical_list: C118284:1-amino-4-phenyl-1,2,3,6-tetrahydropyridine; D004273:DNA, Neoplasm; D011726:Pyridinium Compounds; D015632:1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; D015655:1-Methyl-4-phenylpyridinium