Novel Model of Oxalate Diet-Induced Chronic Kidney Disease in Dahl-Salt-Sensitive Rats.

Diet-induced models of chronic kidney disease (CKD) offer several advantages, including clinical relevance and animal welfare, compared with surgical models. Oxalate is a plant-based, terminal toxic metabolite that is eliminated by the kidneys through glomerular filtration and tubular secretion. An increased load of dietary oxalate leads to supersaturation, calcium oxalate crystal formation, renal tubular obstruction, and eventually CKD. Dahl-Salt-Sensitive (SS) rats are a common strain used to study hypertensive renal disease; however, the characterization of other diet-induced models on this background would allow for comparative studies of CKD within the same strain. In the present study, we hypothesized that SS rats on a low-salt, oxalate rich diet would have increased renal injury and serve as novel, clinically relevant and reproducible CKD rat models. Ten-week-old male SS rats were fed either 0.2% salt normal chow (SS-NC) or a 0.2% salt diet containing 0.67% sodium oxalate (SS-OX) for five weeks.Real-time PCR demonstrated an increased expression of inflammatory marker interleukin-6 (IL-6) (p < 0.0001) and fibrotic marker Timp-1 metalloproteinase (p < 0.0001) in the renal cortex of SS-OX rat kidneys compared with SS-NC. The immunohistochemistry of kidney tissue demonstrated an increase in CD-68 levels, a marker of macrophage infiltration in SS-OX rats (p < 0.001). In addition, SS-OX rats displayed increased 24 h urinary protein excretion (UPE) (p < 0.01) as well as significant elevations in plasma Cystatin C (p < 0.01). Furthermore, the oxalate diet induced hypertension (p < 0.05). A renin-angiotensin-aldosterone system (RAAS) profiling (via liquid chromatography-mass spectrometry; LC-MS) in the SS-OX plasma showed significant (p < 0.05) increases in multiple RAAS metabolites including angiotensin (1-5), angiotensin (1-7), and aldosterone. The oxalate diet induces significant renal inflammation, fibrosis, and renal dysfunction as well as RAAS activation and hypertension in SS rats compared with a normal chow diet. This study introduces a novel diet-induced model to study hypertension and CKD that is more clinically translatable and reproducible than the currently available models.

Cardioprotective Role for Paraoxonase-1 in Chronic Kidney Disease.

Paraoxonase-1 (PON-1) is a hydrolytic enzyme associated with HDL, contributing to its anti-inflammatory, antioxidant, and anti-atherogenic properties. Deficiencies in PON-1 activity result in oxidative stress and detrimental clinical outcomes in the context of chronic kidney disease (CKD). However, it is unclear if a decrease in PON-1 activity is mechanistically linked to adverse cardiovascular events in CKD. We investigated the hypothesis that PON-1 is cardioprotective in a Dahl salt-sensitive model of hypertensive renal disease. Experiments were performed on control Dahl salt-sensitive rats (SSMcwi, hereafter designated SS-WT rats) and mutant PON-1 rats (SS-Pon1em1Mcwi, hereafter designated SS-PON-1 KO rats) generated using CRISPR gene editing technology. Age-matched 10-week-old SS and SS-PON-1 KO male rats were maintained on high-salt diets (8% NaCl) for five weeks to induce hypertensive renal disease. Echocardiography showed that SS-PON-1 KO rats but not SS-WT rats developed compensated left ventricular hypertrophy after only 4 weeks on the high-salt diet. RT-PCR analysis demonstrated a significant increase in the expression of genes linked to cardiac hypertrophy, inflammation, and fibrosis, as well as a significant decrease in genes essential to left ventricular function in SS-PON-1 KO rats compared to SS-WT rats. A histological examination also revealed a significant increase in cardiac fibrosis and immune cell infiltration in SS-PON-1 KO rats, consistent with their cardiac hypertrophy phenotype. Our data suggest that a loss of PON-1 in the salt-sensitive hypertensive model of CKD leads to increased cardiac inflammation and fibrosis as well as a molecular and functional cardiac phenotype consistent with compensated left ventricular hypertrophy.

Paraoxonase-1 Regulation of Renal Inflammation and Fibrosis in Chronic Kidney Disease.

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.

A PON for All Seasons: Comparing Paraoxonase Enzyme Substrates, Activity and Action including the Role of PON3 in Health and Disease.

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.