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Title: The effects of mitochondria-targeted antioxidant mitoquinone on pancreatic acinar cells
Author: Cash, N. J.
ISNI:       0000 0004 6058 7261
Awarding Body: University of Liverpool
Current Institution: University of Liverpool
Date of Award: 2016
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While oxidative stress has been repeatedly implicated in the pathogenesis of acute pancreatitis (AP), recent clinical evidence has found little benefit of general antioxidant therapy. The actions, however, of newer mitochondria-targeted antioxidants such as MitoQ, have yet to be determined in AP. A recently undertaken in vivo study by our group has highlighted mixed effects of MitoQ treatment in caerulein-induced AP (CER-AP) and no protection against bile acid-induced AP (TLCS-AP). Therefore, the in vitro effects of MitoQ against toxin-induced pathophysiological effects of bile acid taurolithocholic acid-3-sulphate (TLCS) and caerulein analogue CCK hyperstimulation were determined. Furthermore, the effects of MitoQ on pancreatic acinar cell physiology and cell death were evaluated and compared to non-antioxidant analogue DecylTPP (dTPP). Additional investigations assessed the potential protective capabilities of MitoQ against non-oxidative ethanol metabolite palmitoleic acid ethyl ester (POAEE)-, ethanol- and H2O2-induced effects. These experiments were carried out alongside a more detailed assessment of H2O2-induced effects in isolated pancreatic acinar cells. Key Findings  MitoQ and non-antioxidant analogue dTPP caused predominantly adverse effects on pancreatic acinar cell responses, in a concentration-dependent manner. These effects are likely due to the targeting component of both TPP+ derivatives.  In vitro, MitoQ did not protect against bile acid TLCS- and CCK hyperstimulation-induced effects on mitochondrial membrane potential (ΔΨm), NAD(P)H levels, cytosolic Ca2+ concentration ([Ca2+]c), cellular apoptosis or necrosis.  CCK and TLCS treatment in vitro, led to differing levels and mode of cell death. CCK induced substantial apoptosis and necrosis, the latter in a biphasic pattern. In contrast, the predominant mode of cell death with TLCS treatment was necrosis. These results mirror in vivo results in AP models demonstrating a more severe AP pathophysiology with TLCS-induced AP than CCK/caerulein-induced AP.  MitoQ had no effects on POAEE- and ethanol-induced cellular necrosis. In contrast, MitoQ provided a mild inhibition of apoptosis in a concentration-dependent manner, consistent with a proposed role of reactive oxygen species (ROS) to promote apoptosis in these cells.  H2O2 induced concentration-dependent effects on levels of ROS, NAD(P)H/FAD+, [Ca2+]c, ΔΨm, cellular apoptosis and necrosis. Low micromolar concentrations favoured apoptosis and high millimolar concentrations necrosis.  MitoQ effectively inhibited H2O2-induced ROS increases. However, MitoQ exacerbated H2O2-induced effects on NAD(P)H/FAD+ levels and provided no protection against high micromolar and millimolar H2O2-induced cell death. These results enhance our understanding of the ROS balance in pancreatic acinar cells. The findings of this study emphasize the unsuitability of the use of targeted antioxidant therapy in the treatment of AP. The novel effects of H2O2 on mitochondrial metabolism, observed at low micromolar concentrations, highlight our incomplete understanding of the role of ROS in cellular function and warrant further investigation.
Supervisor: Criddle, D. Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral