Hypertension alters the function and expression profile of the peptide cotransporters PEPT1 and PEPT2 in the rodent renal proximal tubule

Hypertension is a major risk factor for kidney and cardiovascular disease. The treatment of hypertensive individuals by selected ACE inhibitors and certain di-and tripeptides halts the progression of renal deterioration and extends life-span. Renal reabsorption of these low molecular weight substrates are mediated by the PEPT1 and PEPT2 cotransporters. This study aims to investigate whether hypertension and ageing affects renal PEPT cotransporters at gene, protein expression and distribution as well as function in the superficial cortex and the outer medulla of the kidney. Membrane vesicles from the brush border (BBMV) and outer medulla (OMMV) were isolated from the kidneys of young Wistar Kyoto (Y-WKY), young spontaneously hypertensive (Y-SHR), and middle aged SHR (M-SHR) rats. Transport activity was measured using the substrate, β-Ala-Lys (AMCA). Gene expression levels of PEPT genes were assessed with qRT-PCR while renal localisation of PEPT cotransporters was examined by immunohistochemistry with Western Blot validation. The Km and Vmax of renal PEPT1 were decreased significantly in SHR compared to WKY BBMV, whilst the Vmax of PEPT2 showed differences between SHR and WKY. By contrast to the reported cortical distribution of PEPT1, PEPT1-staining was detected in the outer medulla, whilst PEPT2 was expressed primarily in the cortex of all SHR; PEPT1 was significantly upregulated in the cortex of Y-SHR. These outcomes are indicative of a redistribution of PEPT1 and PEPT2 in the kidney proximal tubule under hypertensive conditions that has potential repercussions for nutrient handling and the therapeutic use of ACE inhibitors in hypertensive individuals

Modulation of the expression of components of the stress response by dietary arachidonic acid in European sea bass (Dicentrarchus labrax) larvae

This study reports for the first time in European sea bass, Dicentrarchus labrax (L.), larvae, the effect of different levels of dietary arachidonic acid (ARA; 20:4n-6) on the expression of genes related to the fish stress response. Copies of mRNA from genes related to steroidogenesis (StAR (steroidogenic acute regulatory protein), c-Fos, and CYP11β (11β- hydroxylase gene)), glucocorticoid receptor complex (GR (glucorticoid receptor) and HSP (heat shock proteins) 70 and 90) and antioxidative stress (catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPX)) were quantified. Eighteen day-old larvae were fed for 14 days with three experimental diets with increasing levels of ARA (0.3, 0.6 and 1.2% d.w.) and similar levels of docosahexaenoic (DHA; 22:6n-3) and eicosapentaenoic (EPA; 20:5n-3) acids (5 and 3%, respectively). The quantification of stress-related genes transcripts was conducted by One-Step TaqMan real time RT-PCR with the standard curve method (absolute quantification). Increase dietary levels of ARA induced a significantly (p<0.05) down-regulation of genes related to cortisol synthesis, such as StAR and CYP11β and up-regulated genes related to glucocorticoid receptor complex, such as HSP70 and GR. No effects were observed on antioxidant enzymes gene expression. These results revealed the regulatory role of dietary ARA on the expression of stress-related genes in European sea bass larvae

Synthetic Cathinones Induce Cell Death in Dopaminergic SH-SY5Y Cells via Stimulating Mitochondrial Dysfunction.

Increasing reports of neurological and psychiatric complications due to psychostimulant synthetic cathinones (SCs) have recently raised public concern. However, the precise mechanism of SC toxicity is unclear. This paucity of understanding highlights the need to investigate the in-vitro toxicity and mechanistic pathways of three SCs: butylone, pentylone, and 3,4-Methylenedioxypyrovalerone (MDPV). Human neuronal cells of SH-SY5Y were cultured in supplemented DMEM/F12 media and differentiated to a neuronal phenotype using retinoic acid (10 μM) and 12-O-tetradecanoylphorbol-13-acetate (81 nM). Trypan blue and lactate dehydrogenase assays were utilized to assess the neurotoxicity potential and potency of these three SCs. To investigate the underlying neurotoxicity mechanisms, measurements included markers of oxidative stress, mitochondrial bioenergetics, and intracellular calcium (Ca2+), and cell death pathways were evaluated at two doses (EC15 and EC40), for each drug tested. Following 24 h of treatment, all three SCs exhibited a dose-dependent neurotoxicity, characterized by a significant (p < 0.0001 vs. control) production of reactive oxygen species, decreased mitochondrial bioenergetics, and increased intracellular Ca2+ concentrations. The activation of caspases 3 and 7 implicated the orchestration of mitochondrial-mediated neurotoxicity mechanisms for these SCs. Identifying novel therapeutic agents to enhance an altered mitochondrial function may help in the treatment of acute-neurological complications arising from the illicit use of these SCs

Evidence supporting oxidative stress in a moderately affected area of the brain in Alzheimer’s disease

The pathogenesis of Alzheimer's disease (AD) remains to be elucidated. Oxidative damage and excessive beta-amyloid oligomers are components of disease progression but it is unclear how these factors are temporally related. At post mortem, the superior temporal gyrus (STG) of AD cases contains plaques, but displays few tangles and only moderate neuronal loss. The STG at post mortem may represent a brain region that is in the early stages of AD or alternately a region resistant to AD pathogenesis. We evaluated expression profiles and activity of endogenous anti-oxidants, oxidative damage and caspase activity in the STG of apolipoprotein ε4-matched human AD cases and controls. Total superoxide dismutase (SOD) activity was increased, whereas total glutathione peroxidase (GPX), catalase (CAT) and peroxiredoxin (Prx) activities, were decreased in the AD-STG, suggesting that hydrogen peroxide accumulates in this brain region. Transcripts of the transcription factor NFE2L2 and inducible HMOX1, were also increased in the AD-STG, and this corresponded to increased Nuclear factor erythroid 2-related factor (NRF-2) and total heme-oxygenase (HO) activity. The protein oxidation marker 4-hydroxynonenal (4-HNE), remained unchanged in the AD-STG. Similarly, caspase activity was unaltered, suggesting that subtle redox imbalances in early to moderate stages of AD do not impact STG viability

The cyclic nitroxide antioxidant 4-methoxy-TEMPO decreases mycobacterial burden in vivo through host and bacterial targets.

Tuberculosis is a chronic inflammatory disease caused by persistent infection with Mycobacterium tuberculosis. The rise of antibiotic resistant strains necessitates the design of novel treatments. Recent evidence shows that not only is M. tuberculosis highly resistant to oxidative killing, it also co-opts host oxidant production to induce phagocyte death facilitating bacterial dissemination. We have targeted this redox environment with the cyclic nitroxide derivative 4-methoxy-TEMPO (MetT) in the zebrafish-M. marinum infection model. MetT inhibited the production of mitochondrial ROS and decreased infection-induced cell death to aid containment of infection. We identify a second mechanism of action whereby stress conditions, including hypoxia, found in the infection microenvironment appear to sensitise M. marinum to killing by MetT both in vitro and in vivo. Together, our study demonstrates MetT inhibited the growth and dissemination of M. marinum through host and bacterial targets