Cyclophosphamide-induced cystitis results in NLRP3-mediated inflammation in the hippocampus and symptoms of depression in rats.

Recent breakthroughs demonstrate that peripheral diseases can trigger inflammation in the brain, causing psychosocial maladies, including depression. While few direct studies have been made, anecdotal reports associate urological disorders with mental dysfunction. Thus, we investigated if insults targeted at the bladder might elicit behavioral alterations. Moreover, the mechanism of neuroinflammation elicited by other peripheral diseases involves the NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome, which is present in microglia in the brain and cleaves and activates proinflammatory cytokines such as IL-1ß. Thus, we further explored the importance of NLRP3 in behavioral and neuroinflammatory changes. Here, we used the well-studied cyclophosphamide (CP)-treated rat model. Importantly, CP and its metabolites do not cross the blood-brain barrier or trigger inflammation in the gut, so that any neuroinflammation is likely secondary to bladder injury. We found that CP triggered an increase in inflammasome activity (caspase-1 activity) in the hippocampus but not in the pons. Evans blue extravasation demonstrated breakdown of the blood-brain barrier in the hippocampal region and activated microglia were present in the fascia dentata. Both changes were dependent on NLRP3 activation and prevented with 2-mercaptoethane sulfonate sodium (Mesna), which masks the effects of the CP metabolite acrolein in the urine. Finally, CP-treated rats displayed depressive symptoms that were prevented by NLRP3 inhibition or treatment with Mesna or an antidepressant. Thus, we conclude that CP-induced cystitis causes NLRP3-dependent hippocampal inflammation leading to depression symptoms in rats. This study proposes the first-ever causative explanation of the previously anecdotal link between benign bladder disorders and mood disorders.

Calcium Pyrophosphate And Monosodium Urate Activate The NLRP3 Inflammasome Within Bladder Urothelium Via Reactive Oxygen Species And TXNIP.

OBJECTIVE: To investigate the in vitro activation of the NLRP3 inflammasome within bladder urothelium by stone-forming components. Further, to describe the contributions of reactive oxygen species (ROS) and thioredoxin-interacting protein (TXNIP), an important structural component of the inflammasome, to this activation. METHODS: Urothelial cells were harvested and incubated overnight. For agonist studies, cells were treated with varying concentrations of calcium pyrophosphate (CPPD) and monosodium urate (MSU). For inhibitor studies, cells were treated with either N-acetylcysteine (NAC) (1 hr) or Verapamil (4 hrs) prior to incubation with either CPPD (62.5 ug/mL) or MSU (1.25 ug/mL) for 24 hrs. Untreated controls were incubated with ATP (1.25 mM) for 1 hr to maximally stimulate NLRP3 inflammasome activity (measured as caspase-1 cleavage of the fluorogenic substrate Ac-YVAD-AFC). Results are reported as a percentage of maximum ATP response. RESULTS: CPPD and MSU activate caspase-1 in urothelial cells in a dose-dependent manner, reaching ~50% and ~25% of the ATP response, respectively. Pre-treatment with the general ROS scavenger NAC reduces this activation in a dose-dependent manner. Additionally, activation was suppressed through treatment with Verapamil, a known downregulator of TXNIP expression. CONCLUSION: The stone components CPPD and MSU activate NLRP3 in an ROS and TXNIP-dependent manner in bladder urothelium. These findings demonstrate the importance of ROS and TXNIP, and suggest that targeting either may be a way to decrease stone-dependent NLRP3 inflammation within the bladder.