Nobuhiro Suzuki1, Hiroshi Mihara2, Hirofumi Nishizono3, Makoto Tominaga4, Toshiro Sugiyama5. 1. Department of Gastroenterology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, 2630 Sugitani, Toyama, Toyama, 930-0194, Japan. nobuhiro@joetsu-hp.jp. 2. Department of Gastroenterology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, 2630 Sugitani, Toyama, Toyama, 930-0194, Japan. m164-tym@umin.net. 3. Division of Animal Experimental Laboratory, Life Science Research Center, University of Toyama, 2630 Sugitani, Toyama, Toyama, 930-0194, Japan. hirofumi@cts.u-toyama.ac.jp. 4. Division of Cell Signaling, Okazaki Institute for Integrative Bioscience (National Institute for Physiological Sciences), National Institutes of Natural Sciences, Nishigonaka 38, Myodaiji, Okazaki, Aichi, 444-8585, Japan. tominaga@nips.ac.jp. 5. Department of Gastroenterology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, 2630 Sugitani, Toyama, Toyama, 930-0194, Japan. tsugi@med.u-toyama.ac.jp.
Abstract
BACKGROUND: The reflux of pancreatic-duodenal fluids is implicated in the pathophysiology of proton-pump inhibitor-resistant gastroesophageal reflux disease (GERD). Protease-activated receptor-2 (PAR-2) is activated by proteases, the pancreatic enzyme, trypsin, and the activated PAR-2 enhances transient receptor potential vanilloid 4 (TRPV4) function in neurons. TRPV4 stimulates ATP exocytosis in conjunction with the vesicular nucleotide transporter, which mediates mechano-transduction and vagal stimulation. The aim of the present study was to verify whether the activated PAR-2 up-regulates TRPV4 function in mouse esophageal keratinocytes, which may link to the pathophysiology in PPI-resistant GERD. METHODS: TRPV4 and PAR-2 expressions were detected by RT-PCR, immunostaining, and western blotting in mouse esophageal keratinocytes. The functional response of TRPV4 to esophageal keratinocytes was analyzed using a Ca(2+) imaging system. Cellular ATP release was examined by luciferase-luciferin reaction. TRPV4 phosphorylation was studied by immunoprecipitation and western blotting. RESULTS: PAR-2 and TRPV4 mRNAs and proteins were expressed in esophageal keratinocytes. Pre-treatment with trypsin significantly increased the responses to TRPV4 activator in esophageal keratinocytes, probably via the phosphorylation of serine residue of TRPV4 by protein kinase C and resulted in cellular ATP release from the cells. CONCLUSIONS: Activated PAR-2 with trypsin exposure up-regulated TRPV4 function and increased ATP release in mouse esophageal keratinocytes. This mechanism might be related to the pathophysiology of GERD, especially non-erosive GERD.
BACKGROUND: The reflux of pancreatic-duodenal fluids is implicated in the pathophysiology of proton-pump inhibitor-resistant gastroesophageal reflux disease (GERD). Protease-activated receptor-2 (PAR-2) is activated by proteases, the pancreatic enzyme, trypsin, and the activated PAR-2 enhances transient receptor potential vanilloid 4 (TRPV4) function in neurons. TRPV4 stimulates ATP exocytosis in conjunction with the vesicular nucleotide transporter, which mediates mechano-transduction and vagal stimulation. The aim of the present study was to verify whether the activated PAR-2 up-regulates TRPV4 function in mouse esophageal keratinocytes, which may link to the pathophysiology in PPI-resistant GERD. METHODS:TRPV4 and PAR-2 expressions were detected by RT-PCR, immunostaining, and western blotting in mouse esophageal keratinocytes. The functional response of TRPV4 to esophageal keratinocytes was analyzed using a Ca(2+) imaging system. Cellular ATP release was examined by luciferase-luciferin reaction. TRPV4 phosphorylation was studied by immunoprecipitation and western blotting. RESULTS:PAR-2 and TRPV4 mRNAs and proteins were expressed in esophageal keratinocytes. Pre-treatment with trypsin significantly increased the responses to TRPV4 activator in esophageal keratinocytes, probably via the phosphorylation of serine residue of TRPV4 by protein kinase C and resulted in cellular ATP release from the cells. CONCLUSIONS: Activated PAR-2 with trypsin exposure up-regulated TRPV4 function and increased ATP release in mouse esophageal keratinocytes. This mechanism might be related to the pathophysiology of GERD, especially non-erosive GERD.
Authors: Rink-Jan Lohman; Adam J Cotterell; Jacky Suen; Ligong Liu; Anh T Do; David A Vesey; David P Fairlie Journal: J Pharmacol Exp Ther Date: 2011-10-25 Impact factor: 4.030
Authors: Kevin S Thorneloe; Mui Cheung; Weike Bao; Hasan Alsaid; Stephen Lenhard; Ming-Yuan Jian; Melissa Costell; Kristeen Maniscalco-Hauk; John A Krawiec; Alan Olzinski; Earl Gordon; Irina Lozinskaya; Lou Elefante; Pu Qin; Daniel S Matasic; Chris James; James Tunstead; Brian Donovan; Lorena Kallal; Anna Waszkiewicz; Kalindi Vaidya; Elizabeth A Davenport; Jonathan Larkin; Mark Burgert; Linda N Casillas; Robert W Marquis; Guosen Ye; Hilary S Eidam; Krista B Goodman; John R Toomey; Theresa J Roethke; Beat M Jucker; Christine G Schnackenberg; Mary I Townsley; John J Lepore; Robert N Willette Journal: Sci Transl Med Date: 2012-11-07 Impact factor: 17.956