Summary
We studied the change in gap junctional intercellular communication (GJIC) on human umbilical vein endothelial cells (HUVECs) under hypoxia/reoxygenation (H/R) conditions and the influence of leukocytes on GJIC by the fluorescence redistribution after photobleaching (FRAP) method. Contrast microscopic observations showed no significant changes in the morphology of the HUVECs after H/R. Reoxygenation following hypoxia (12 h) caused a time-dependent decrease in GJIC; that is, GJIC reduction was induced after 2 h and reached maximum levels at 4-6 h, recovering to normal levels after 18 h. The oxidant-sensitive fluorescence dye assay revealed that generation of reactive oxygen species (ROS) increased during the first 2 h after reoxygenation. Hydroxyl radical scavengers abolished reduction of GJIC by H/R. H/R led to significant release of membrane and supernatant tumor necrosis factor-α (TNFα). Anti-human TNFa antibody abolished the H/R-induced suppression of GJIC. GJIC of HUVECs that had adherent neutrophils was suppressed. Treatment with anti-human intercellular adhesion molecule-1 (ICAM-1) antibody abolished this suppression. Assays using intercell chambers showed that no suppression of GJIC was observed. This suppression was abolished by pretreatment with tyrosine kinase inhibitors. These data suggest that (1) ischemia/reperfusion injury may be involved in suppression of GJIC induced by TNFa release via reactive oxygen species; (2) suppression of GJIC by adherent neutrophils is mediated by ROS in HUVECs, not by proteases, and this suppression is attributed to tyrosine phosphorylation of gap junction protein caused by neutrophil adhesion through integrin and ICAM-1 binding.
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References
McCord JM (1985) Oxygen derived free radicals in post-ischemic tissue injury. N Engl J Med 312:159–163
Grace PA (1994) Ischemic-reperfusion injury. Br J Surg 81:637–647
Trosko JE, Madhukar BV, Chang CC (1993) Endogenous and exogenous modulating of gap junctional intercellular communication. Life Sci 53:1–19
Dejana E, Carada M, Lampugnari MG (1995) Endothelial cell-cell junctions. FASEB J 9:910–918
Jaffe EA, Nachman RL, Becker CG (1973) Culture of human endothelial cells derived from umbilical veins. J Clin Invest 52:2745–2756
Yoshimoto S, Ishizaki Y, Murota S (1986) Effect of carbon dioxide and oxygen on endothelin production by cultured porcine cerebral endothelial cells. Stroke 22:378–383
Wade MH, Trosko JE, Schindler M (1986) A fluorescence photobleaching assay of gap junction-mediated communication between human cells. Science 232:525–528
Fujita H, Morita I, Murota S (1993) A possible mechanism for vascular endothelial cell injury elicited by activated leukocytes. Arch Biochem Biophys 309:62–69
Barchowsky A, Williams ME, Benz CC (1994) Oxidant-sensitive protein phosphorylation in endothelial cells. Free Radic Biol Med 16:771–777
Oh SY, Grupen CG, Murray AW (1991) Phorbol ester induces phosphorylation and down-regulation of connexin 43 in WB cells. Biochim Biophys Acta 1094:243–245
Hu VW, Xie HQ (1994) Interleukin-1α suppresses gap junction-mediated intercellular communication in human endothelial cells. Exp Cell Res 213:218–223
Mensink A, Haan LH, Lakemond CM (1995) Inhibition of gap junctional intercellular communication between primary human smooth muscle cells by TNFα. Carcinogenesis 16:2063–2067
Murota S, Fujita H, Morita I (1996) Cell adhesion molecules mediates endothelial cell injury caused by activated neutrophils. Keio J Med 45:207–212
Jansen LA, Mesnil M, Jongen WM (1996) Inhibition gap junctional intercellular communication and delocalization of the cell adhesion molecule E-cadherin by tumor promoters. Carcinogenesis 17:1527–1531
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© 2001 Springer Japan
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Nishida, M., Morita, I., Murota, SI. (2001). Mechanism of Inhibitory Effect of Hypoxia/Reoxygenation on Gap Junctional Intercellular Communication of Vascular Endothelial Cells in Culture. In: Fukuuchi, Y., Tomita, M., Koto, A. (eds) Ischemic Blood Flow in the Brain. Keio University Symposia for Life Science and Medicine, vol 6. Springer, Tokyo. https://doi.org/10.1007/978-4-431-67899-1_7
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DOI: https://doi.org/10.1007/978-4-431-67899-1_7
Publisher Name: Springer, Tokyo
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