title: A potential role of calcium in apoptosis and aberrant chromatin forms in porcine kidney PK15 cells induced by individual and combined ochratoxin A and citrinin.
abstract: The aim of this study was to establish the involvement of calcium signalling in genotoxicity, apoptosis and necrosis evoked by ochratoxin A (OTA) and citrinin (CTN) alone or in combination in porcine kidney PK15 cells. Cell proliferation test (MTT) and trypan blue assays (24 h) demonstrated that CTN (IC(50) = 73.5 ± 1.0, 75.4 ± 1.4 μM, respectively) was less toxic than OTA (IC(50) = 14.0 ± 2.4, 20.5 ± 1.0 μM, respectively). To test their cytotoxic interactions, two doses of single OTA (6 and 10 μM) and CTN (30 and 50 μM) and their combinations were applied. Combined treatment showed additive cytotoxic effects. OTA and CTN induced dose-dependent increase in cytosolic calcium level (assessed with Fura-2 AM). However, combined treatment did not provoke additional increase in calcium signal. The rate of apoptosis and necrosis (DAPI-antifade staining) was significantly higher after 12 h than 24 h, while the frequencies of micronuclei (MNs) and nuclear buds (NBs) were higher after 24 h than 12 h treatment. Combined exposure resulted in apoptotic and necrotic synergism, while genotoxic effects of OTA + CTN were noted as antagonistic or additive. Co-exposure of cells to calcium chelator BAPTA-AM significantly reduced CTN and OTA + CTN-evoked apoptosis. Twenty-four hour after co-exposure to BAPTA-AM and a single OTA and CTN, MNs significantly decreased while NBs dropped significantly after co-treatment with BAPTA-AM and OTA + CTN. In conclusion, disturbance of Ca(2+) homeostasis caused by OTA and CTN plays a significant role in cell genotoxicity and death.
authors: Department of Microbiology, Faculty of Pharmacy and Biochemistry, University of Zagreb, Croatia. msegvic@pharma.hr
authors: Maja Segvić
authors: MS
authors: Klarić
authors: Davor
authors: D
authors: Zelježić
authors: Lada
authors: L
authors: Rumora
authors: Maja
authors: M
authors: Peraica
authors: Stjepan
authors: S
authors: Pepeljnjak
authors: Ana-Marija
authors: AM
authors: Domijan
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D002843:Chromatin / Q000187:drug effects / Q000378:metabolism; D002953:Citrinin / Q000008:administration & dosage / Q000633:toxicity*; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D020128:Inhibitory Concentration 50; D007668:Kidney / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D009152:Mutagenicity Tests / Q000379:methods; D009336:Necrosis / Q000139:chemically induced; D009793:Ochratoxins / Q000008:administration & dosage / Q000633:toxicity*; D013552:Swine; D013997:Time Factors
chemical_list: D002843:Chromatin; D009793:Ochratoxins; C025589:ochratoxin A; D002953:Citrinin; D002118:Calcium