Combination exposure of melamine and cyanuric acid is associated with polyuria and activation of NLRP3 inflammasome in rats

Abstract

The molecular mechanisms of melamine-induced renal toxicity have not been fully understood. The purpose of the study aimed to investigate whether melamine and cyanuric acid induced NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome activation in the kidney, which may contribute to abnormal water and sodium handling in a rat model. Wistar rats received melamine (Mel; 200 mg·kg body wt·day), cyanuric acid (CA; 200 mg·kg body wt·day), or Mel plus CA (Mel + CA; 100 mg·kg body wt·day, each) for 2 wk. Mel + CA caused damaged tubular epithelial structure and organelles, dilated tubular lumen, and inflammatory responses. Crystals were observed in urine and serum specimen, also in the lumen of dilated distal renal tubules. The combined ingestion of Mel and CA in rats caused a markedly impaired urinary concentration, which was associated with reduced protein expression of aquaporin (AQP)1, 2, and 3 in inner medulla and α-Na-K-ATPase and Na-K-2Cl transporters in cortex and outer medulla. Mel + CA treatment was associated with increased protein expression of CD3 and mRNA levels of CD68 and F4/80 as well as phosphorylation of NF-κB in the kidney. Mel + CA treatment increased protein and mRNA expression of NLRP3 inflammasome components apoptosis-associated speck-like protein containing a caspase recruitment domain, caspase-1, and IL-1β in the inner medulla of rats. NF-κB inhibitor Bay 11-7082 reduced IL-1β expression induced by Mel + CA and prevented downregulation of AQP2 in inner medullary collecting duct cell suspensions. In conclusion, Mel + CA treatment caused urinary-concentrating defects and reduced expression of renal AQPs and key sodium transporters, which is likely due to the inflammatory responses and activation of NLRP3 inflammasome induced by crystals formed in the kidney.

Statins ameliorate cholesterol-induced inflammation and improve AQP2 expression by inhibiting NLRP3 activation in the kidney

“…Our recent studies 36 , 37 have revealed an inhibitory role of NLRP3 inflammasome in the regulation of aquaporins in the kidneys. In order to unravel the potential mechanism by which cholesterol may affect the protein expression of AQP2, the protein expression profiles of ASC, an adaptor of NLRP3 inflammasome, were examined in the kidney specimens of patients with or without atorvastatin treatment.…”

“…Next we examined a potential mechanism by which cholesterol may impact the protein expression of AQP2. Our recent studies 36 , 37 have revealed an inhibitory role of NLRP3 inflammasome in the regulation of aquaporins of the kidneys. As expected, the protein expression of ASC monomer, cleaved Caspase-1 (p20) and cleaved IL-1β in the kidneys was found to be upregulated in wild type mice on a HFD; while in nlrp3 -/- mice, the HFD seemed to have failed to induce significant changes in the protein expression of ASC, Caspase-1 (p20) and cleaved IL-1β expression (Figure 3 A and B).…”

Inhibition of IL-1β by Aliskiren Improved Renal AQP2 Expression and Urinary Concentration Defect in Ureteral Obstruction and Release

“…For semiquantitative immunoblotting and immunocyto chemistry, previously characterized affinity-purified polyclonal antibodies to AQP2 and AQP3 were used (Wang et al, 2018). Antibodies to AQP1 were obtained from BOSTER (Wuhan, China); antibodies to IL-1β, p-NF-κB p65, and NF-κB p65 from Cell Signaling Technology; caspase-1 from Abcam; antibodies to ASC from Santa Cruz; and NLRP3 from Novus.…”

“…The left kidney blocks containing all kidney zones were dehydrated and embedded in paraffin. For immunoperoxidase microscopy, sections (4 μm thick) cut from paraffin-embedded kidney samples were used for immunostaining of AQP2 (Wang et al, 2018). Briefly, after dewaxing and rehydration, a microwave pretreatment in citrate buffer (pH 6.2) was performed to unmask antigens present in the renal tissue.…”

“…NOD-, LRR-, and pyrin domain-containing 3 (NLRP3) inflammasome is a cytosolic signaling macromolecular complex comprising a sensor molecule [NOD-like receptors (NLRs)], the adaptor apoptosis-associated speck-like protein containing a CARD (ASC), and the effector protease caspase 1, which processes pro-interleukin (IL-1β) into its mature form IL-1β that causes inflammation and tissue damage (Mangan et al, 2018). Recent studies have suggested that NLRP3 inflammasome and downstream cytokines contribute to several types of kidney disease, including crystalline nephropathy (Mulay et al, 2013; Wang et al, 2018), obstructive nephropathy (Wang W. et al, 2015), and obesity-related kidney diseases (Ke et al, 2018). We previously demonstrated that IL-1β directly inhibits AQP2 expression in the collecting duct principal cells of the kidneys (Wang W. et al, 2015), suggesting that inflammatory cytokines downregulate the expression of water channels and sodium transporters, resulting in altered water and sodium regulation in the kidneys.…”