title: Intercellular communication in the immune system: differential expression of connexin40 and 43, and perturbation of gap junction channel functions in peripheral blood and tonsil human lymphocyte subpopulations.
abstract: The distribution and function of connexins (integral membrane proteins assembled into gap junction intercellular communication channels) were studied in human lymphocyte subpopulations. The expression of mRNA encoding connexins in peripheral blood and tonsil-derived T, B and natural killer (NK) lymphocytes was examined. Connexin43 (Cx43) mRNA was expressed in peripheral blood and tonsil lymphocytes, but Cx40 mRNA expression was confined to tonsil-derived T and B lymphocytes; Cx26, Cx32, Cx37 and Cx45 were not detected by reverse transcription-polymerase chain reaction (RT-PCR). Western blot analysis also demonstrated the presence of Cx40 and Cx43 proteins in T and B lymphocytes in a manner coincidental to the mRNA detection. Stimulation in vitro of T and B lymphocytes with phytohaemagglutinin (PHA) and lipopolysaccharide (LPS), respectively, increased Cx40 and Cx43 protein expression. Flow cytometric analysis, using antibodies to extracellular loop amino acid sequences of connexins, confirmed the surface expression of connexins in all lymphocyte subpopulations. Assembly of connexins into gap junctions providing direct intercellular channels linking attached lymphocytes was demonstrated by using a dye transfer technique. The exchange of dye between lymphocytes was inhibited by a connexin extracellular loop mimetic peptide and alpha-glycyrrhetinic acid, two reagents that restrict intercellular communication across gap junctions. Dye coupling occurred between homologous and heterologous co-cultures of T and B lymphocytes, and was not influenced by their stimulation with PHA and LPS. The connexin mimetic peptide caused a significant decrease in the in vitro synthesis of immunoglobulin M (IgM) by T- and B-lymphocyte co-cultured populations in the presence or absence of stimulation by PHA. The results identify connexins as important cell surface components that modulate immune processes.
authors.affiliation: Department of Medical Biochemistry and Wales Heart Research Institute, University of Wales College of Medicine, Cardiff, UK.
authors.forename: E
authors.initials: E
authors.lastname: Oviedo-Orta
authors.forename: T
authors.initials: T
authors.lastname: Hoy
authors.forename: W H
authors.initials: WH
authors.lastname: Evans
mesh_terms: D001402:B-Lymphocytes / Q000378:metabolism; D015153:Blotting, Western; D002450:Cell Communication / Q000502:physiology*; D002478:Cells, Cultured; D002648:Child; D018920:Coculture Techniques; D000072259:Connexin 26; D018031:Connexin 43 / Q000032:analysis / Q000235:genetics; D017630:Connexins / Q000032:analysis / Q000235:genetics*; D005434:Flow Cytometry; D017629:Gap Junctions / Q000378:metabolism*; D015870:Gene Expression; D006034:Glycyrrhetinic Acid / Q000494:pharmacology; D006801:Humans; D007107:Immune System / Q000502:physiology*; D007075:Immunoglobulin M / Q000378:metabolism; D007694:Killer Cells, Natural / Q000378:metabolism; D008213:Lymphocyte Activation; D016131:Lymphocyte Subsets / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D014066:Palatine Tonsil / Q000276:immunology; D012333:RNA, Messenger / Q000032:analysis*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013601:T-Lymphocytes / Q000378:metabolism; D000097003:Gap Junction alpha-5 Protein
chemical_list: D000072259:Connexin 26; D018031:Connexin 43; D017630:Connexins; D006034:Glycyrrhetinic Acid; D007075:Immunoglobulin M; D012333:RNA, Messenger; D000097003:Gap Junction alpha-5 Protein; C000607042:GJB2 protein, human