Endothelial Cell Products as a Key Player in Hypoxia-Induced Nerve Cell Injury after Stroke

  • Cho, Chul-Min (Departments of Neurosurgery, College of Medicine, Dong-A University) ;
  • Ha, Se-Un (Departments of Physiology, College of Medicine, Dong-A University) ;
  • Bae, Hae-Rahn (Departments of Physiology, College of Medicine, Dong-A University) ;
  • Huh, Jae-Taeck (Departments of Neurosurgery, College of Medicine, Dong-A University)
  • 발행 : 2006.08.30

초록

Objective : Activated endothelial cells mediate the cascade of reactions in response to hypoxia for adaptation to the stress. It has been suggested that hypoxia, by itself, without reperfusion, can activate the endothelial cells and initiate complex responses. In this study, we investigated whether hypoxia-induced endothelial products alter the endothelial permeability and have a direct cytotoxic effect on nerve cells. Methods : Hypoxic condition of primary human umbilical vein endothelial cells[HUVEC] was induced by $CoCl_2$ treatment in culture medium. Cell growth was evaluated by 3,4,5-dimethyl thiazole-3,5-diphenyl tetrazolium bromide [MTT] assay Hypoxia-induced products [$IL-1{\beta},\;TGF-{\beta}1,\;IFN-{\gamma},\;TNF-{\alpha}$, IL-10, IL-6, IL-8, MCP-l and VEGF] were assessed by enzyme-linked immunosorbent assay. Endothelial permeability was evaluated by Western blotting. Results : Prolonged hypoxia caused endothelial cells to secrete IL -6, IL -8, MCP-1 and VEGF. However, the levels of IL -1, IL -10, $TNF-{\alpha},\;TGF-{\beta},\;IFN-{\gamma}$ and nitric oxide remained unchanged over 48 h hypoxia. Hypoxic exposure to endothelial cells induced the time-dependent down regulation of the expression of cadherin and catenin protein. The conditioned medium taken from hypoxic HUVECs had the cytotoxic effect selectively on neuroblastoma cells, but not on astroglioma cells. Conclusion : These results suggest the possibility that endothelial cell derived cytokines or other secreted products with the increased endothelial permeability might directly contribute to nerve cell injury followed by hypoxia.

키워드

참고문헌

  1. Ala Y, Palluy O, Favero J, Bonne C, Modat G, Dornand J : Hypoxia/ reoxygenationstimulates endothelial cells to promote interleukin-1 and interleukin-6 production. Effects of free radical scavengers. Agents Actions 37 : 134-139, 1992 https://doi.org/10.1007/BF01987902
  2. Ali MH, Schlidt SA, Chandel NS, Hynes KL, Schumacker PT, Gewertz BL : Endothelial permeability and IL-6 production during hypoxia : role of ROS in signal transduction. Am J Physiol 277 : L1057-L1065, 1999
  3. Ali MH, Schlidt SA, Hynes KL, Marcus BC, Gewertz BL : Prolonged hypoxia alters endothelial barrier function. Surgery 124 : 491-497, 1998 https://doi.org/10.1016/S0039-6060(98)70094-7
  4. Arnould T, Michiels C, Remacle J : Increased PMN adherence on endothelial cells after hypoxia : involvement of PAF, CD18/CD11b, and ICAM-1. Am J Physiol 264 : C1102-C1110, 1993 https://doi.org/10.1152/ajpcell.1993.264.5.C1102
  5. Barth AI, Nathke IS, Nelson WJ : Cadherins, catenins and APC protein : Interplay between cytoskeletal complexes and signaling pathways. Curr Opin Cell Biol 9 : 683-690, 1997 https://doi.org/10.1016/S0955-0674(97)80122-6
  6. BradburyMW : The blood-brain barrier. Exp Physiol 78 : 453-472, 1993 https://doi.org/10.1113/expphysiol.1993.sp003698
  7. Chandel NS, Maltepe E, Goldwasser E, Mathieu CE, Simon MC, Schumacker PT : Mitochondrial reactive oxygen species trigger hypoxia-induced transcription. Proc Natl Acad Sci USA 95 : 11715-11720, 1998
  8. Clark ET, Desai TR, Hynes KL, Gewertz BL : Endothelial cell response to hypoxia-reoxygenation is mediated by IL-1. J Surg Res 58 : 675-681, 1995 https://doi.org/10.1006/jsre.1995.1106
  9. Corada M, Mariotti M, Thurston G, Smith K, Kunkel R, Brockhaus M, et al : Vascular endothelial-cadherin is an important determinant of microvascular integrity in vivo. Proc Natl Acad Sci USA 96 : 9815- 9820, 1999
  10. Dejana E, Valiron O, Navarro P, Lampugnani MG : Intercellular junctions in the endothelium and the control of vascular permeability. Ann N Y Acad Sci 811 : 36-43, 1997 https://doi.org/10.1111/j.1749-6632.1997.tb51986.x
  11. del Zoppo G, Ginis I, Hallenbeck JM, Iadecola C, Wang X, Feuerstein GZ : Inflammation and stroke : putative role for cytokines, adhesion molecules and iNOS in brain response to ischemia. Brain Pathol 10 : 95-112, 2000
  12. Dore-Duffy P, Balabanov R, Beaumont T, Hritz MA, Harik SI, LaManna JC : Endothelial activation following prolonged hypobaric hypoxia. Microvascular Res 57 : 75-85, 1999 https://doi.org/10.1006/mvre.1998.2112
  13. Faller DV : Endothelial cell responses to hypoxic stress. Clin Exp Pharmacol Physiol 26 : 74-84, 1999 https://doi.org/10.1046/j.1440-1681.1999.02992.x
  14. Goldberg MA, Dunning SP, Bunn HF : Regulation of the erythropoietin gene : evidence that the oxygen sensor is a heme protein. Science 242 : 1412-1415, 1988 https://doi.org/10.1126/science.2849206
  15. Jin KL, Mao XO, Greenberg DA : Vascular endothelial growth factor : direct neuroprotective effect in in vitro ischemia. Proc Natl Acad Sci USA 97 : 10242-10247, 2000
  16. Lampugnani MG, Resnati M, Raiteri M, Pigott R, Pisacane A, Houen G, et al : A novel endothelial-specific membrane protein is a marker of cell-cell contacts. J Cell Biol 118 : 1511-1522, 1992 https://doi.org/10.1083/jcb.118.6.1511
  17. Levy AP, LevyNS, Wegner S, Goldberg MA : Transcriptional regulation of the rat vascular endothelial growth factor gene by hypoxia. J Biol Chem 270 : 13333-13340, 1995 https://doi.org/10.1074/jbc.270.22.13333
  18. Liu T, Clark RK, Mcdonnell PC, Young PR, White RF, Barone FC, et al : Tumor necrosis factor $\alpha$expression in ischemic neurons. Stroke 25 : 1481-1488, 1994 https://doi.org/10.1161/01.STR.25.7.1481
  19. Liu T, Mcdonnell PC, Young PR, White RF, Siren AL, Barone FC, et al : Interleukin-1${\beta}$ mRNA expression in ischemic rat cortex. Stroke 24 : 1746-1750, 1993 https://doi.org/10.1161/01.STR.24.11.1746
  20. Liu Y, Cox SR, Morita T, Kourembanas S : Hypoxia regulates vascular endothelial growth factor gene expression in endothelial cells. Identification of a 5'enhancer. Circ Res 77 : 638-643, 1995 https://doi.org/10.1161/01.RES.77.3.638
  21. Michiels C, Arnould T, Remacle J : Endothelial cell responses to hypoxia : initiation of a cascade of cellular interactions. Biochim Biophys Acta 2 : 1-10, 2000 https://doi.org/10.1016/0006-3002(48)90002-X
  22. Namiki A, Brogi E, Kearney M, KimEA, Wu T, Couffinhal T, et al : Hypoxia induces vascular endothelial growth factor in cultured human endothelial cells. J Biol Chem 270 : 31189-31195, 1995 https://doi.org/10.1074/jbc.270.52.31189
  23. Neufeld G, Cohen T, Gengrinovitch S, Poltorak Z : Vascular endothelial growth factor(VEGF) and its receptors. FASEB J 13 : 9-22, 1999 https://doi.org/10.1096/fasebj.13.1.9
  24. Palmer LA, Semenza GL, Stoler MH, Johns RA : Hypoxia induces type II NOS gene expression in pulmonary artery endothelial cells via HIF-1. Am J Physiol 274 : L212-L219, 1998
  25. Palmer RM, Ferrige AG, Moncada S : Nitric oxide release accounts for the biological activity of endothelium-derived relaxing factor. Nature 327 : 524-526, 1987 https://doi.org/10.1038/327524a0
  26. Park JH, Okayama N, Gute D, Krsmanovic A, Battarbee H, Alexander JS : Hypoxia/aglycemia increases endothelial permeability : role of second messengers and cytoskeleton. Am J Physiol 277 : C1066-C1074, 1999 https://doi.org/10.1152/ajpcell.1999.277.6.C1066
  27. Phelan MW, Faller DV : Hypoxia decreases constitutive nitric oxide synthetase transcript and protein in cultured endothelial cells. J Cell Physiol 167 : 469-476, 1996 https://doi.org/10.1002/(SICI)1097-4652(199606)167:3<469::AID-JCP11>3.0.CO;2-#
  28. Plateel M, Dehouck MP, Torpier G, Cecchelli R, Teissier E : Hypoxia increases the permeability of the blood-brain barrier endothelial cell monolayer. J Neurochem 65 : 2138-2145, 1995 https://doi.org/10.1046/j.1471-4159.1995.65052138.x
  29. Plateel M, Teissier E, Cecchelli R : Hypoxia dramatically increases the nonspecific transport of blood-borne proteins to the brain. J Neurochem 68 : 874-877, 1997 https://doi.org/10.1046/j.1471-4159.1997.68020874.x
  30. Reyes TM, Fabry Z, Coe CL : Brain endothelial cell production of a neuroprotective cytokine, interleukin-6, in response to noxious stimuli. Brain Res 851 : 215-220, 1999 https://doi.org/10.1016/S0006-8993(99)02189-7
  31. Sawa Y, Ichikawa H, Kagisaki K, Ohata T, Matsuda H : Interleukin-6 derived from hypoxic myocytes promotes neutrophil-mediated reperfusion injury in myocardium. J Thorac Cardiovasc Surg 116 : 511-517, 1998 https://doi.org/10.1016/S0022-5223(98)70018-2
  32. Schweitzer CM, van de Loosdrecht AA, Jonkhoff AR, Ossenkoppele GL, Huijgens PC, Drager AM, et al : Spectrophotometric determination of clonogenic capacity of leukemic cells in a semisolid microtiter culture system. Exp Hematol 21 : 573-578, 1993
  33. Semenza GL : HIF-1 : mediator of physiological and pathophysiological responses to hypoxia. J Appl Physiol 88 : 1474-1480, 2000 https://doi.org/10.1152/jappl.2000.88.4.1474
  34. Semenza GL, Roth PH, Fang HM, Wang GL : Transcriptional regulation of genes encoding glycolytic enzymes by hypoxia-inducible factor 1. J Biol Chem 269 : 23757-23763, 1994
  35. Senger DR, Connolly DT, Van de Water L, Feder J, Dvorak HF : Purification and NH2-terminal amino acid sequence of guinea pig tumor-secreted vascular permeability. Cancer Res 50 : 1774-1778, 1990
  36. Shreeniwas R, Koga S, Karakurum M, Pinsky D, Kaiser E, Brett J, et al : Hypoxia-mediated induction of endothelial cell interleukin-1 alpha. An autocrine mechanism promoting expression of leukocyte adhesion molecules on the vessel surface. J Clin Invest 90 : 2333-2339, 1992 https://doi.org/10.1172/JCI116122
  37. Stavri GT, Hong Y, Zachary IC, Breier G, Baskerville PA, Yla-Herttuala S, et al : Hypoxia and platelet-derived growth factor-BB synergistically upregulate the expression of vascular endothelial growth factor in vascular smooth muscle cells. FEBS Lett 358 : 311-315, 1995 https://doi.org/10.1016/0014-5793(94)01458-D
  38. Terui Y, Ikeda M, Tomizuka H, Kasahara T, Ohtsuki T, Uwai M, et al : Activated endothelial cells induce apoptosis in leukemic cells by endothelial interleukin-8. Blood 92 : 2672-2680, 1998
  39. Ukropec JA, Hollinger MK, Salva SM, Woolkalis MJ : SHP2 association with VE-cadherin complexs in human endothelial cells is regulated by thrombin. J Biol Chem 275 : 5983-5986, 2000 https://doi.org/10.1074/jbc.275.8.5983
  40. Wang GL, Semenza GL : Molecular basis of hypoxia-induced erythropoietin expression. Curr Opin Hematol 3 : 156-162, 1996 https://doi.org/10.1097/00062752-199603020-00009