Involvement of autophagy in ochratoxin-a (OTA) -mediated toxicity in HK-2 cell line

Abstract

Ochratoxin A (OTA) is a secondary metabolite produced by fungi and found in a variety of food and feed. It is classi ed as a possible human carcinogen and immense amount of work has revealed to its nephrotoxic and carcinogenic properties. Autophagy is a cellular degradation process in which long-lived or defective organelles and proteins are cleared in order to maintain cellular homeostasis. In this study, we investigated the involvement of autophagy and related signaling pathways MAPK/ERK and PI3K/AKT during progression of OTA-induced toxicity in human proximal tubule epithelial HK-2 cell lines. Time- and dose-dependent autophagic activity in response to OTA exposure was tested by examining the alterations of the amount of autophagyrelated proteins, the conversion of microtubule-associated protein 1 light chain 3 (LC3)- 1 to LC3-II, and by visualization of autophagasomes and acidic compartments. The results showed that OTA at 10 M concentration induces autophagy at very early hours of treatment. After 6 hours, autophagasome formation decreases while AKT phosphorylation is up-regulated. Concordantly, inhibition of PI3K/AKT pathway increases the autophagasome formation while inhibition of MAPK/ERK had no e ect on the regulation of autophagy in response to OTA. OTA has also decreased global steadystate levels of protein ubiquitinylation. In addition, the decrease in p62/SQSTM and increase in acidic vacuoles at later time points when the autophagic activity decreased could be the consequence of OTA-induced fast degradation by ubiquitin proteasome system (UPS) rather than autophagy. Taken together, this study suggest that OTA up-regulates autophagy at early hours of treatment and then down-regulates by the OTA-mediated PI3K/AKT pathway activation at later time points in HK-2 cell lines.