Evaluation of Urothelial Stretch-Induced Cyclooxygenase-2 Expression in Novel Human Cell Culture and Porcine in Vivo Ureteral Obstruction Models

Abstract

Obstruction and stretch induce cyclooxygenase (COX)-2 expression and prostanoid synthesis in urinary tissues, causing pain, inflammation, hypercontractility, and cell proliferation. Our objective was to characterize acute COX-2 induction during in vivo ureteral obstruction, establish a cell culture model of urothelial stretch-induced COX-2 expression, and evaluate whether mechanotransduction could alter transcriptional and post-transcriptional regulation of COX-2. We performed laparoscopic unilateral ureteral ligation in pigs and allowed progression for 1, 2, 6, 24, or 48 h. We evaluated COX-2 expression with reverse transcriptase (RT)-polymerase chain reaction (PCR) and immunoblotting. We cultured primary human urothelial cells on stretch plates, applied stretch for up to 48 h, and measured COX-2 expression by RT-PCR and immunoblotting, transcription with run-on assays, and mRNA stability with actinomycin mRNA decay assays. In vivo ureteral obstruction induced COX-2 expression 4-fold within 6 h, maintaining induction for 24 h. In cell culture, stretch induced COX-2 steady-state mRNA and protein within the first 3 h of stretch, maintaining this induction for over 6 h. Three hours of stretch doubled COX-2 transcription relative to unstretched controls and increased COX-2 mRNA half-life 3-fold. This is the first report to characterize in vivo temporal stretch-induced COX-2 expression in the urothelium and establish a primary urothelial cell culture model for the study of stretch-induced COX-2 mechanisms. This is also the first report to identify alterations in steady-state COX-2 mRNA having components of both transcriptional and posttranscriptional regulation of stretch-regulated COX-2. Future elucidation of COX-2 signaling may identify novel therapeutic targets for treating stretch and distension of urinary tissues.

Ureteral Access Sheath Influence on the Ureteral Wall Evaluated by Cyclooxygenase-2 and Tumor Necrosis Factor-α in a Porcine Model

“…The same study showed a four-fold induction in COX-2 expression after 6 hours of acute ureteral obstruction in swine ureters harvested ex vivo. 11 In a similar study using mice subjected to unilateral ureteral obstruction, COX-2 levels in whole ureteral tissue extracts were increased nine-fold after 6 hours compared to unobstructed controls. 10 …”

“…The elevated expression levels were comparable to what has previously been reported after several hours of ureteral obstruction. 8,9,11 Furthermore, this effect persisted after 2 hours of rest and reperfusion before tissue sampling was performed. To our knowledge this is the first study showing such a substantial impact on COX-2 and TNF-α expression in the ureter by what may be characterized as a minimal surgical intervention.…”

Nicotine Augments Glomerular Injury in a Rat Model of Acute Nephritis

“…Clinical and experimental studies have demonstrated that COX-2 is upregulated in conditions associated with glomerular inflammation including acute glomerulonephritis and ureteral obstruction [29,30,31]. In addition, recent experimental studies have also demonstrated that COX-2 expression is upregulated in experimental models of diabetes [32, 33] and chronic kidney disease [34, 35] and that COX-2 inhibition in a model of renal ablation reduces mesangial matrix expansion, glomerular sclerosis and fibronectin and collagen IV expression, suggesting that COX-2-derived prostaglandins play an important role in the pathogenesis of renal injury [36].…”

PI3K Mediates Stretch-Induced COX-2 Expression During Urinary Tract Obstruction

“…Our in vivo findings are therefore physiologically comparable to leukocyteindependent mechanisms of COX-2 expression observed during human UTO. 16 We further showed that PI3K signaling is upstream of both stretch-induced PKCζ activation and stretchinduced COX-2 expression during urothelial cell stretch.…”

“…The final published version may differ from this proof. 16 16 stretch-induced PKCζ activation ( Figure 5C). These results indicate that both stretch-induced PKCζ activation and COX-2 expression are PI3K-dependent.…”