Cholinergic regulation of proliferation of the urothelium in response to E. coli lipopolysaccharide exposition

Abstract

How the proliferation of the urothelium is regulated is known to a little degree. E. coli lipopolysaccharide (LPS) activates the innate immune response of the urinary bladder via the Toll-like receptor 4 (TLR4) on the urothelium but induces also urothelial proliferation. We wanted to assess whether muscarinic receptors are involved in the regulation of urothelial proliferation triggered by LPS stimulation. Female Fischer 344 rats were instilled with LPS or saline (control) in the urinary bladder in the absence or presence of muscarinic receptor blockade with atropine and regeneration of the urothelium was assessed 4h and 24h later. In the Fischer 344 bladder, urothelial thinning and urothelial caspase 3 up-regulation occurred at 4h after LPS urinary bladder instillation, which were totally blocked in rats pre-treated with atropine. TLR4 was only expressed in blood vessels in the Fischer 344 bladder, while it was also expressed in umbrella cells in the Sprague-Dawley bladder. Proliferation (Ki67 incorporation) of the human urothelial cell line UROtsa was reduced in the presence of the muscarinic receptor antagonists methoctramine (M2/M4-selective) and pirenzepine (M1/M4-selective), while proliferation instead was enhanced in the presence of atropine. In UROtsa cells exposed to LPS for 24h, 4-DAMP (M3/M1/M5-selective) inhibited instead proliferation. In conclusion, muscarinic receptors regulate urothelial proliferation and LPS may induce urothelial apoptosis via muscarinic receptor-dependent pathways. Our findings also suggest that species differences exist in the expressional pattern of TLR4 in the urothelium.