<p>Calcium Pyrophosphate And Monosodium Urate Activate The NLRP3 Inflammasome Within Bladder Urothelium Via Reactive Oxygen Species And TXNIP</p>

Abstract

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 stonedependent NLRP3 inflammation within the bladder.

Fluoxetine increases astrocytic glucose uptake and glycolysis in corticosterone-induced depression through restricting GR-TXNIP-GLUT1 Pathway

“…Fluoxetine significantly inhibited corticosteroneinduced TXNIP transcription in A9433 cells (IC 50 = 13.6 μM) (Figure 5G). In this experiment, a reported TXNIP transcriptional inhibitor verapamil (Xu et al, 2012;Harper et al, 2019), presented IC 50 value of 125.7 μM (Figure 5H).…”

“…RU486 (a GR antagonist, 1 μM) was used to inhibit the activity of GR. Verapamil, a reported TXNIP transcription inhibitor ( Xu et al, 2012 ; Harper et al, 2019 ), was used as positive control in TXNIP transcription-related experiments. Different indicators were detected after 72 h treatment of corticosterone, fluoxetine or RU486.…”

Inflammation triggered by the NLRP3 inflammasome is a critical driver of diabetic bladder dysfunction

“…Recent studies have begun to explore a possible role of NLRP3 in a number of different inflammatory bladder pathologies. The NLRP3 inflammasome was first implicated in bladder inflammation in a cyclophosphamideinduced hemorrhagic cystitis model (Hughes et al, 2014) but, since that time, it has been implicated in other pathologies including bladder outlet obstruction (Hughes et al, 2016a;Lutolf et al, 2018;Chen et al, 2021), urinary tract infections (Hughes et al, 2016b;Demirel et al, 2018;Lindblad et al, 2019;Mao et al, 2021), chemotherapy (Hughes et al, 2014), stone damage (Harper et al, 2019), interstitial cystitis/painful bladder syndrome (Tudrej et al, 2019;Shih et al, 2021;Wang et al, 2021), cigarette smoke-induced bladder dysfunction (Wu et al, 2020;Ma et al, 2021), ageing (Chen et al, 2019) and, the focus of this review, diabetes (Hughes et al, 2019a;Hughes et al, 2022a;Odom et al, 2022).…”

“…In the early stages, hyperglycemia creates oxidative stress and this would be particularly prevalent in insulin-independent tissues like urothelia. Oxidative stress is a well-known activator of NLRP3 (Vanaja et al, 2015;Malik and Kanneganti, 2017;Swanson et al, 2019;Yang et al, 2019b;Mamun et al, 2021) and is known to do so in urothelia (Harper et al, 2019). In addition, classic diabetic DAMPs such as MSU, HMGB-1, C6-ceramide, and AGEs would become more prevalent systemically while increasing in concentration in the urine as they are stored for excretion.…”

Dihydrolipoic acid protects against lipopolysaccharide-induced behavioral deficits and neuroinflammation via regulation of Nrf2/HO-1/NLRP3 signaling in rat

“…The excessive level of ROS would damage the integrity of cells and result in dysfunction of the tissues via peroxidation of lipids, proteins, mitochondria, and DNA of cells [7,8]. Despite that the specific regulatory mechanism of NLRP3 inflammasome activation is unclear, ROS has been frequently reported to be correlated with NLRP3 inflammasome activation [9,10]. Also, it regulates the expression and/ or activation of NLRP3 inflammasome in several diseases, including intestinal inflammation and cardiovascular disease.…”