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01 May 2004

Expression and functional role of serine/threonine phosphatases in rat esophagus

Ivo R. van der Voort, Jörg Knapp, Jan W. Konturek, Peter Boknik, Wolfram Domschke, Wilhelm Schmitz, Joachim Neumann

Med Sci Monit 2004; 10(5): BR123-129 :: ID: 11665

Abstract

Background:Smooth muscle contraction is due to phosphorylation, while relaxation results from dephosphorylation of regulatory proteins such as the light chains of myosin (MLC,sub>20[/sub]). Dephosphorylation of MLC[sub]20[/sub] is catalyzed by protein phosphatases (PP) types 1 and 2A. While some data are available on the expression and functional importance of PP1 and PP2A in vascular smooth muscle preparations, little is known of the importance of PP for force regulation in the gastrointestinal tract.Material/Methods: Here we address the expression and functional importance of PP1 and PP2A in the rat esophagus. We measured both the isometric force of contraction and PP activity and also performed Western and Northern blotting for PPs in the tunica muscularis mucosae (TMM) and the muscularis mucosae propria (MM).Results: Cantharidin (0.01–100 µM) inhibited PP activity in homogenates from TMM (IC[sub]50[/sub]=20 µM) in a concentration-dependent manner. The mRNAs of catalytic and regulatory subunits (PP1a, PP1b, PP2Aa and M110) were detectable at 1.8 kb, 3.2 kb, 2.0 kb and 5.7 kb in both TMM and MM. Western blot analysis revealed PP1 in TMM and MM. PP2A (protein) was more abundant in TMM than in TMM. Cantharidin increased muscular tone in isolated preparations of TMM (starting at 100 µM) to 164.6 (14.8 % of predrug force at 300 µM (the highest concentration studied). This increase in force of contraction was comparable to that of 10 µM carbachol.Conclusions: The results suggest that PP activity can play an important role in the regulation of esophageal tone.

Keywords: Blotting, Northern, Blotting, Western, Cantharidin - chemistry, Catalysis, Catalytic Domain, Digestive System, Dose-Response Relationship, Drug, Esophagus - enzymology, Esophagus - metabolism, Esophagus - pathology, Inhibitory Concentration 50, Muscle, Smooth - cytology, Muscle, Smooth, Vascular - cytology, Myocytes, Smooth Muscle - cytology, Myosins - chemistry, Phosphoprotein Phosphatase - biosynthesis, Phosphoprotein Phosphatase - metabolism, Phosphoprotein Phosphatase - physiology, Phosphoric Monoester Hydrolases - metabolism, Phosphorylation, Protein Isoforms, RNA, Messenger - metabolism, Rats, Wistar, Time Factors, Blotting, Northern, Blotting, Western, Cantharidin - chemistry, Catalysis, Catalytic Domain, Digestive System, Dose-Response Relationship, Drug, Esophagus - pathology, Inhibitory Concentration 50, Muscle, Smooth - cytology, Muscle, Smooth, Vascular - cytology, Myocytes, Smooth Muscle - cytology, Myosins - chemistry, Phosphoprotein Phosphatases - physiology, Phosphoric Monoester Hydrolases - metabolism, Phosphorylation, Protein Isoforms, RNA, Messenger - metabolism, Rats, Wistar, Time Factors

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Medical Science Monitor eISSN: 1643-3750
Medical Science Monitor eISSN: 1643-3750