TNFalpha suppresses human colonic circular smooth muscle cell contractility by SP1- and NF-kappaB-mediated induction of ICAM-1.
OBJECTIVE
Intercellular adhesion molecule 1 (ICAM-1) receptors are expressed at low levels on human colonic circular smooth muscle cells (HCCSMCs) and their expression is increased in patients with Crohn's disease. We investigated the roles of transcription factors Sp1 and nuclear factor kappa B (NF-kappaB) in the regulation of ICAM-1 expression on HCCSMCs and examined whether ICAM-1 expression mediates the suppression of contractility in response to TNFalpha.


METHODS
Experiments were performed on primary cultures of HCCSMCs and fresh human colonic circular muscle strips.


RESULTS
TNFalpha treatment of HCCSMCs induced rapid and prolonged accumulation of ICAM-1 messenger RNA (mRNA) and protein. NF-kappaB inhibition before, but not after, 1 hour of TNFalpha-stimulation blocked the expression of ICAM-1. TNFalpha significantly enhanced Sp1/DNA binding. Sp1 bound to the 3' flanking region of a variant kappaB site in the -192/-172 region of ICAM-1 promoter. Mutation of this region abolished the response to TNFalpha. The treatment of HCCSMCs with Sp1 antisense oligonucleotides (ODNs) blocked the expression of ICAM-1, but sense ODNs had no effect. Protein kinase C zeta (PKCzeta) inhibition before or 3 hours after stimulation with TNFalpha also blocked the expression of ICAM-1. TNFalpha treatment of circular muscle strips pretreated with ICAM-1 sense ODNs or control medium significantly reduced their response to acetylcholine, whereas pretreatment with antisense ODNs blocked this effect.


CONCLUSIONS
The expression of ICAM-1 on HCCSMCs in response to TNFalpha is regulated by transcription factors Sp1 and NF-kappaB binding independently to the -192/-172 region of the ICAM-1 promoter. The expression of ICAM-1 plays a critical role in the suppression of cell contractility in response to TNFalpha.
Department of Internal Medicine, Enteric Neuromuscular Disorders and Visceral Pain Center, Division of Gastroenterology, The University of Texas Medical Branch at Galveston, 77555-1064, USA.
Konrad
K
Pazdrak
Xuan-Zheng
XZ
Shi
Sushil K
SK
Sarna
D001483:Base Sequence; D001665:Binding Sites; D002478:Cells, Cultured; D003106:Colon / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000235:genetics*; D008969:Molecular Sequence Data; D009119:Muscle Contraction / Q000187:drug effects*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects* / Q000502:physiology; D016328:NF-kappa B / Q000502:physiology*; D011401:Promoter Regions, Genetic; D011493:Protein Kinase C / Q000502:physiology; D012333:RNA, Messenger / Q000032:analysis; D016329:Sp1 Transcription Factor / Q000502:physiology*; D051996:Transcription Factor RelA; D014158:Transcription, Genetic; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D000099221:Protein Kinase C zeta
D018799:Intercellular Adhesion Molecule-1; D016328:NF-kappa B; D011493:Protein Kinase C; D012333:RNA, Messenger; D016329:Sp1 Transcription Factor; D051996:Transcription Factor RelA; D014409:Tumor Necrosis Factor-alpha; D000099221:Protein Kinase C zeta