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Human microvascular endothelial synthesis of interleukin‐8 during in vitro ischemia and reperfusion
Author(s) -
Albadawi Hassan,
Patton George M.,
Bratton Charles F.,
Peterson Brian G.,
Watkins Michael T.
Publication year - 2006
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.21002
Subject(s) - ischemia , intracellular , secretion , extracellular , in vitro , endothelial stem cell , interleukin 8 , pharmacology , chemokine , chemistry , biology , cytokine , immunology , medicine , endocrinology , biochemistry , inflammation
These studies were undertaken to evaluate human microvascular endothelial cell (MEC) synthesis of interleukin‐8 (IL‐8), a potent neutrophil chemoattractant, under in vitro conditions of ischemia and reperfusion. IL‐8 and other related CXC chemokines are believed to mediate tissue injury in a variety of pathologic conditions in humans. MEC grown on microcarrier beads were exposed to 3 or 6 h of in vitro ischemia followed by 2 h of reperfusion. Conditioned medium, MEC protein, and total RNA extracts were assayed for IL‐8 using an ELISA. During ischemia alone, MEC increased intracellular, but not extracellular levels of IL‐8 secretion. In contrast, reperfusion markedly stimulated both intracellular and extracellular IL‐8 secretion. Neither 3 h of ischemia alone or followed by reperfusion altered steady‐state levels of IL‐8 mRNA when compared to pre‐ischemic levels. In contrast, after 6 h of ischemia alone and ischemia followed by reperfusion, IL‐8 mRNA was increased eight‐ and sixfold, respectively, when compared to pre‐ischemic levels. These studies demonstrate an inverse relationship between the rate of IL‐8 protein secretion and the steady‐state levels of IL‐8 mRNA during ischemia and reperfusion. During ischemia and reperfusion both the increase in cell‐associated IL‐8 protein and the release of IL‐8 into the medium is dependent on de novo protein synthesis rather than the intracellular accumulation of IL‐8. These experiments indicate that post‐ischemic modulation of IL‐8 release and synthesis following ischemia reperfusion will require strategies directed towards inhibition of IL‐8 transcription and in depth knowledge of the mechanisms regulating IL‐8 secretion. J. Cell. Biochem. 100: 412–420, 2007. © 2006 Wiley‐Liss, Inc.