Abstract
This study describes the investigation of the efficiency of conjugated linoleic acid (CLA) isomers in reducing cancer cells viability exploring the role of the oxidative stress and acylpeptide hydrolase (APEH)/proteasome mediated pathways on pro-apoptotic activity of the isomer trans10,cis12 (t10,c12)-CLA. The basal activity/expression levels of APEH and proteasome (β-5 subunit) were preliminarily measured in eight cancer cell lines and the functional relationship between these enzymes was clearly demonstrated through their strong positive correlation. t10,c12-CLA efficiently inhibited the activity of APEH and proteasome isoforms in cell-free assays and the negative correlation between cell viability and caspase 3 activity confirmed the pro-apoptotic role of this isomer. Finally, modulatory effects of t10,c12-CLA on cellular redox status (intracellular glutathione, mRNA levels of antioxidant/detoxifying enzymes activated through NF-E2-related factor 2, Nrf2, pathway) and on APEH/β-5 activity/expression levels, were investigated in A375 melanoma cells. Dose- and time-dependent variations of the considered parameters were established and the resulting pro-apoptotic effects were shown to be associated with an alteration of the redox status and a down-regulation of APEH/proteasome pathway. Therefore, our results support the idea that these events are involved in ROS-dependent apoptosis of t10,c12-CLA-treated A375 cells. The combined inhibition, triggered by t10,c12-CLA, via the modulation of APEH/proteasome and Nrf2 pathway for treating melanoma, is suggested as a subject for further in vivo studies.
Highlights
Oxidative stress is a dynamic status characterized by an imbalance between the production of reactive oxygen species (ROS) and the activity and availability of antioxidants
A delicate balance between cellular redox status and proteasome activity is clearly indicated by ROS production during the initiation of apoptotic signalling triggered by bortezomib (BTZ, Velcade), a widely used proteasome inhibitor [7,8,9,10] and the impairment of proteasome activity by oxidative stress [11,12]
We examined the basal expression/activity level of proteasome and acylpeptide hydrolase (APEH) and the anti-proliferative activities elicited by three different conjugated linoleic acid (CLA) isomers (c9,t11, t9,t11- or t10,c12-CLA) in eight randomly selected cancer cell lines
Summary
Oxidative stress is a dynamic status characterized by an imbalance between the production of reactive oxygen species (ROS) and the activity and availability of antioxidants. Among intracellular antioxidants, reduced glutathione (GSH) plays a central role in the maintenance of the thiol-disulfide redox state in mammalian cells and its deregulation is responsible for apoptosis evasion [2], colonizing ability [3] and multidrug resistance of cancer cells [4]. Alterations in redox status are known to lead the induction of apoptosis in cancer cells and its decrease represents a molecular mechanism whereby anti-cancer agents reduces malignant cell survival [5]. Of note, during oxidative stress, the higher activity of 20S proteasome core enzymes aimed at counteracting the accumulation of oxidatively damaged proteins, greatly contributes to secondary anti-oxidative defences [1]. A delicate balance between cellular redox status and proteasome activity is clearly indicated by ROS production during the initiation of apoptotic signalling triggered by bortezomib (BTZ, Velcade), a widely used proteasome inhibitor [7,8,9,10] and the impairment of proteasome activity by oxidative stress [11,12]
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