Ochratoxin A activates neutrophils and kills these cells through necrosis, an effect eliminated through its conversion into ochratoxin α.
Ochratoxin A (OTA) is a mycotoxin produced by several species of fungi from the Aspergillus and Penicillium genera that frequently grow in improperly stored food products. OTA has carcinogenic, teratogenic and nephrotoxic potential and sustains a high half-life in human blood. Despite the recently efforts to decontaminate OTA through its conversion into its metabolite ochratoxin alpha (OTα), there are just a few reports in literature comparing the toxic effects of these toxins. Thus, herein we studied and compared the proinflammatory and toxicological effects of OTA and its metabolite OTα in human neutrophils in vitro. The effect of OTA and OTα on human neutrophils viability was evaluated by trypan blue, annexin-V and propidium iodide methods as well as by the analysis of cytomorphological alterations. The ATP levels were also evaluated using the luciferin-luciferase bioluminescence assay. The alteration on mitochondrial potential was assessed by a mitoscreen flow cytometry mitochondrial membrane potential detection kit and the intracellular calcium levels through the probe FLUO-4/AM. To study the human neutrophils' oxidative burst, the fluorescent probe dichlorodihydrofluorescein diacetate was used. OTA induced an increase on the intracellular calcium, human neutrophils' oxidative burst followed by depletion of ATP levels and alterations on mitochondrial potential leading to cell death by necrosis, while OTα did not induce significant toxic effects. Our results strongly suggest that the toxicity in human neutrophils induced by OTA started with the release of calcium from internal stores triggering several neutrophils' activities that culminate in cell death by necrosis.
UCIBIO, REQUIMTE, Laboratório de Química Aplicada, Departamento de Ciências Químicas, Faculdade de Farmácia, Universidade do Porto, 4050-313 Porto, Portugal; Laboratório de Ciência de Alimentos, Escola de Química e Alimentos, Universidade Federal do Rio Grande - FURG, 96201-900 Rio Grande, RS, Brazil.
Larine
L
Kupski
UCIBIO, REQUIMTE, Laboratório de Química Aplicada, Departamento de Ciências Químicas, Faculdade de Farmácia, Universidade do Porto, 4050-313 Porto, Portugal. Electronic address: marisafreitas@ff.up.pt.
Marisa
M
Freitas
UCIBIO, REQUIMTE, Laboratório de Química Aplicada, Departamento de Ciências Químicas, Faculdade de Farmácia, Universidade do Porto, 4050-313 Porto, Portugal.
Daniela
D
Ribeiro
Laboratório de Ciência de Alimentos, Escola de Química e Alimentos, Universidade Federal do Rio Grande - FURG, 96201-900 Rio Grande, RS, Brazil.
Eliana Badiale
EB
Furlong
UCIBIO, REQUIMTE, Laboratório de Química Aplicada, Departamento de Ciências Químicas, Faculdade de Farmácia, Universidade do Porto, 4050-313 Porto, Portugal. Electronic address: egracas@ff.up.pt.
Eduarda
E
Fernandes
D002273:Carcinogens / Q000633:toxicity; D002470:Cell Survival / Q000187:drug effects; D005506:Food Contamination; D005516:Food Microbiology; D006801:Humans; D053078:Membrane Potential, Mitochondrial; D009336:Necrosis / Q000139:chemically induced / Q000473:pathology*; D009504:Neutrophils / Q000166:cytology / Q000187:drug effects*; D009793:Ochratoxins / Q000737:chemistry* / Q000633:toxicity*; D013723:Teratogens / Q000633:toxicity
D002273:Carcinogens; D009793:Ochratoxins; D013723:Teratogens; C000426:5-chloro-8-hydroxy-3,4-dihydro-3-methylisocoumarin-7-carboxylic acid; C025589:ochratoxin A
ATP depletion; Intracellular Ca(2+); Necrosis; Ochratoxin A; Ochratoxin α; Oxidative burst