title: Contraction of gut smooth muscle cells assessed by fluorescence imaging.
abstract: Here we discuss the development of a novel cell imaging system for the evaluation of smooth muscle cell (SMC) contraction. SMCs were isolated from the circular and longitudinal muscular layers of mouse small intestine by enzymatic digestion. SMCs were stimulated by test agents, thereafter fixed in acrolein. Actin in fixed SMCs was stained with phalloidin and cell length was determined by measuring diameter at the large end of phalloidin-stained strings within the cells. The contractile response was taken as the decrease in the average length of a population of stimulated-SMCs. Various mediators and chemically identified compounds of daikenchuto (DKT), pharmaceutical-grade traditional Japanese prokinetics, were examined. Verification of the integrity of SMC morphology by phalloidin and DAPI staining and semi-automatic measurement of cell length using an imaging analyzer was a reliable method by which to quantify the contractile response. Serotonin, substance P, prostaglandin E2 and histamine induced SMC contraction in concentration-dependent manner. Two components of DKT, hydroxy-α-sanshool and hydroxy-β-sanshool, induced contraction of SMCs. We established a novel cell imaging technique to evaluate SMC contractility. This method may facilitate investigation into SMC activity and its role in gastrointestinal motility, and may assist in the discovery of new prokinetic agents.
authors.affiliation: Comprehensive Kampo Research Planning Dept, Kampo Scientific Strategies Division, Tsumura & Co., 2-17-11 Akasaka, Minato-ku, Tokyo 107-8521, Japan; Tsumura Research Laboratories, Kampo Scientific Strategies Division, Tsumura & Co., 3586 Yoshiwara, Ami-machi, Inashiki-gun, Ibaraki 300-1192, Japan. Electronic address: tokita_youhei@mail.tsumura.co.jp.
authors.forename: Yohei
authors.initials: Y
authors.lastname: Tokita
authors.affiliation: Department of Gastroenterology, Kitakyushu Municipal Medical Center, 2-1-1 Bashaku, Kokurakita-ku, Kitakyushu-shi, Fukuoka 802-0077, Japan; Department of Medicine and Bioregulatory Science, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
authors.forename: Hirotada
authors.initials: H
authors.lastname: Akiho
authors.affiliation: Department of Medicine and Bioregulatory Science, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Nakamura
authors.affiliation: Department of Medicine and Bioregulatory Science, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
authors.forename: Eikichi
authors.initials: E
authors.lastname: Ihara
authors.affiliation: Tsumura Research Laboratories, Kampo Scientific Strategies Division, Tsumura & Co., 3586 Yoshiwara, Ami-machi, Inashiki-gun, Ibaraki 300-1192, Japan.
authors.forename: Masahiro
authors.initials: M
authors.lastname: Yamamoto
mesh_terms: D000818:Animals; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D015232:Dinoprostone / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D055808:Drug Discovery; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D005769:Gastrointestinal Motility; D006632:Histamine / Q000494:pharmacology; D007421:Intestine, Small / Q000166:cytology*; D008297:Male; D008807:Mice, Inbred BALB C; D009119:Muscle Contraction / Q000187:drug effects*; D009130:Muscle, Smooth / Q000166:cytology* / Q000187:drug effects*; D061848:Optical Imaging / Q000379:methods*; D005894:Panax; D010936:Plant Extracts / Q000494:pharmacology; D053284:Polyunsaturated Alkamides / Q000494:pharmacology; D012701:Serotonin / Q000494:pharmacology; D013268:Stimulation, Chemical; D013373:Substance P / Q000494:pharmacology; D032106:Zanthoxylum; D030521:Zingiberaceae
chemical_list: D005231:Fatty Acids, Unsaturated; D010936:Plant Extracts; D053284:Polyunsaturated Alkamides; C121414:dai-kenchu-to; C000598329:hydroxy-alpha sanshool; D012701:Serotonin; D013373:Substance P; D006632:Histamine; D015232:Dinoprostone
keywords: Daikenchuto; Hydroxy-sanshool; Kampo; Neurotransmitter; Smooth muscle cell contraction