Changes in the Expression of Smooth Muscle Contractile Proteins in TNBS- and DSS-Induced Colitis in Mice
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Changes in the Expression of Smooth Muscle Contractile Proteins in TNBS- and DSS-Induced Colitis in Mice Reem Alkahtani,1 Sunila Mahavadi,1 Othman Al-Shboul,1 Shakir Alsharari,2 John R. Grider,1 and Karnam S. Murthy1,3
Abstract—Thin filament-associated proteins such as calponin, caldesmon, tropomyosin, and smoothelin are thought to regulate acto-myosin interaction and thus, muscle contraction. However, the effect of inflammation on the expression of thin filament-associated proteins is not known. The aim of the present study is to determine the changes in the expression of calponin, caldesmon, tropomyosin, and smoothelin in colonic smooth muscle from trinitrobenzene sulphonic acid (TNBS)- and dextran sodium sulphate (DSS)-induced colitis in mice. Expression of h-caldesmon, h2-calponin, α-tropomyosin, and smoothelin-A was measured by qRT-PCR and Western blot. Contraction in response to acetylcholine in dispersed muscle cells was measured by scanning micrometry. mRNA and protein expression of α-actin, h2-calponin, h-caldesmon, smoothelin, and α-tropomyosin in colonic muscle strips from mice with TNBS- or DSS-induced colitis was significantly increased compared to control animals. Contraction in response to acetylcholine was significantly decreased in muscle cells isolated from inflamed regions of TNBS- or DSS-treated mice compared to control mice. Our results show that increase in the expression of thin filament-associated contractile proteins, which inhibit acto-myosin interaction, could contribute to decrease in smooth muscle contraction in inflammation. KEY WORDS: inflammation; contractile proteins; TNBS.
INTRODUCTION The smooth muscle cells of the gastrointestinal tract are the final effectors of force development and work. The main contractile apparatus in the smooth muscle consists of two types of filaments: thin filaments and thick filaments [1–6]. Thin filaments consist of actin, a ∼42-kDa protein which exists in vivo as filamentous actin (F-actin), and associated proteins such as caldesmon, calponin, tropomyosin, and smoothelin. Thick filaments are aggregates of myosin molecules. The interaction of actin with myosin and subsequent hydrolysis of ATP is the fundamental reaction whereby chemical energy is converted into mechanical 1
Department of Physiology, VCU Program in Enteric Neuromuscular Sciences, Virginia Commonwealth University, Richmond, VA 232980551, USA 2 Department of Pharmacology, Virginia Commonwealth University, Richmond, VA 23298-0551, USA 3 To whom correspondence should be addressed at Department of Physiology, VCU Program in Enteric Neuromuscular Sciences, Virginia Commonwealth University, Richmond, VA 23298-0551, USA. E-mail: [email protected]
energy. An essential step in smooth muscle contraction is phosphorylation of the 20-kDa regulatory light chains (MLC20) at Ser19, which increases significantly the actinactivated myosin ATPase activity [1, 4]. Phosphorylation and dephosphorylation of MLC20 are directly correlated to smooth muscle contraction and relaxation, respectively, and MLC20 phosphorylation level
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