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We have previously shown in a murine model of Non-alcoholic Fatty Liver Disease (NAFLD) that chronic, low-dose exposure to the Harmful Algal Bloom cyanotoxin microcystin-LR (MC-LR), resulted in significant hepatotoxicity including micro-vesicular lipid accumulation, impaired toxin metabolism as well as dysregulation of the key signaling pathways involved in inflammation, immune response and oxidative stress. On this background we hypothesized that augmentation of hepatic drug metabolism pathways with targeted antioxidant therapies would improve MC-LR metabolism and reduce hepatic injury in NAFLD mice exposed to MC-LR. We chose N-acetylcysteine (NAC, 40 mM), a known antioxidant that augments the glutathione detoxification pathway and a novel peptide (pNaKtide, 25 mg/kg) which is targeted to interrupting a specific Src-kinase mediated pro-oxidant amplification mechanism. Histological analysis showed significant increase in hepatic inflammation in NAFLD mice exposed to MC-LR which was attenuated on treatment with both NAC and pNaKtide (both p ≤ 0.05). Oxidative stress, as measured by 8-OHDG levels in urine and protein carbonylation in liver sections, was also significantly downregulated upon treatment with both antioxidants after MC-LR exposure. Genetic analysis of key drug transporters including Abcb1a, Phase I enzyme-Cyp3a11 and Phase II metabolic enzymes-Pkm (Pyruvate kinase, muscle), Pklr (Pyruvate kinase, liver, and red blood cell) and Gad1 (Glutamic acid decarboxylase) was significantly altered by MC-LR exposure as compared to the non-exposed control group (all p ≤ 0.05). These changes were significantly attenuated with both pNaKtide and NAC treatment. These results suggest that MC-LR metabolism and detoxification is significantly impaired in the setting of NAFLD, and that these pathways can potentially be reversed with targeted antioxidant treatment.

Papraoxonase-1 (PON1) is a hydrolytic lactonase enzyme that is synthesized in the liver and circulates attached to high-density lipoproteins (HDL). Clinical studies have demonstrated an association between diminished PON-1 and the progression of chronic kidney disease (CKD). However, whether decreased PON-1 is mechanistically linked to renal injury is unknown. We tested the hypothesis that the absence of PON-1 is mechanistically linked to the progression of renal inflammation and injury in CKD. Experiments were performed on control Dahl salt-sensitive rats (SSMcwi, hereafter designated SS rats) and Pon1 knock-out rats (designated SS-Pon1em1Mcwi, hereafter designated SS-PON-1 KO rats) generated by injecting a CRISPR targeting the sequence into SSMcwi rat embryos. The resulting mutation is a 7 bp frameshift insertion in exon 4 of the PON-1 gene. First, to examine the renal protective role of PON-1 in settings of CKD, ten-week-old, age-matched male rats were maintained on a high-salt diet (8% NaCl) for up to 5 weeks to initiate the salt-sensitive hypertensive renal disease characteristic of this model. We found that SS-PON-1 KO rats demonstrated several hallmarks of increased renal injury vs. SS rats including increased renal fibrosis, sclerosis, and tubular injury. SS-PON-1 KO also demonstrated increased recruitment of immune cells in the renal interstitium, as well as increased expression of inflammatory genes compared to SS rats (all p < 0.05). SS-PON-1 KO rats also showed a significant (p < 0.05) decline in renal function and increased renal oxidative stress compared to SS rats, despite no differences in blood pressure between the two groups. These findings suggest a new role for PON-1 in regulating renal inflammation and fibrosis in the setting of chronic renal disease independent of blood pressure.

Paraoxonases (PONs) are a family of hydrolytic enzymes consisting of three members, PON1, PON2, and PON3, located on human chromosome 7. Identifying the physiological substrates of these enzymes is necessary for the elucidation of their biological roles and to establish their applications in the biomedical field. PON substrates are classified as organophosphates, aryl esters, and lactones based on their structure. While the established native physiological activity of PONs is its lactonase activity, the enzymes’ exact physiological substrates continue to be elucidated. All three PONs have antioxidant potential and play an important anti-atherosclerotic role in several diseases including cardiovascular diseases. PON3 is the last member of the family to be discovered and is also the least studied of the three genes. Unlike the other isoforms that have been reviewed extensively, there is a paucity of knowledge regarding PON3. Thus, the current review focuses on PON3 and summarizes the PON substrates, specific activities, kinetic parameters, and their association with cardiovascular as well as other diseases such as HIV and cancer.