title: Hypertonic saline activates protein tyrosine kinases and mitogen-activated protein kinase p38 in T-cells.
abstract: OBJECTIVE
  In previous in vitro studies, we have found that hypertonic saline (HTS) can augment T-cell proliferation and restore the function of suppressed T-cells. Our animal models have shown that HTS resuscitation reverses immunosuppression after hemorrhage and reduces mortality from sepsis. In the present study, we investigated if and how HTS may influence T-cell signaling and function on a subcellular level.
  
  
  METHODS
  Human peripheral blood mononuclear cells (PBMC) were used to determine the effect of HTS on T-cell interleukin 2 (IL-2) production and proliferation. Human Jurkat T-cells were used to study the effects of HTS on T-cell signal transduction, IL-2 mRNA transcription, and IL-2 expression.
  
  
  METHODS
  The effect of HTS on T-cell proliferation and IL-2 production was measured with PBMC and Jurkat T-cells. IL-2 mRNA transcription in HTS-treated Jurkat cells was measured by reverse transcriptase polymerase chain reaction. HTS-induced protein tyrosine phosphorylation in Jurkat T-cells was determined by immunoblotting with anti-phosphotyrosine antibodies. Expression in Jurkat cells of the mitogen-activated protein kinase p38 (MAPK p38), a signal transduction protein that is activated by osmotic stress, was determined by immunoblotting with anti-MAPK p38 antibodies. HTS-induced MAPK p38 activation in Jurkat cells was measured with an immune-complex kinase assay using ATF-2 as a substrate.
  
  
  RESULTS
  Proliferation of activated human PBMC increased significantly upon addition of HTS to the culture medium. This effect of HTS was paralleled by enhanced IL-2 production of activated PBMC and Jurkat cells and IL-2 mRNA transcription of Jurkat cells. HTS exposure of Jurkat cells caused tyrosine phosphorylation of a number of cellular proteins. We found that Jurkat T-cells expressed MAPK p38 and that it was activated in the presence of HTS. All these effects of HTS on T-cell signaling and function were observed at NaCl concentrations that were within physiologically relevant levels (20-100 mmol/L hypertonicity).
  
  
  CONCLUSIONS
  In T-cells, HTS triggers a signaling pathway that includes increased tyrosine phosphorylation of several cellular proteins and activation of MAPK p38. HTS alone does not result in IL-2 mRNA transcription, IL-2 expression, or T-cell proliferation. However, in combination with other stimuli, HTS augments T-cell IL-2 expression and proliferation. We speculate that HTS could "resuscitate" suppressed T-cells in trauma patients by circumvention of, or substituting for, blocked signaling pathways.
authors.affiliation: Department of Surgery, University of California San Diego, USA.
authors.forename: W G
authors.initials: WG
authors.lastname: Junger
authors.forename: D B
authors.initials: DB
authors.lastname: Hoyt
authors.forename: M
authors.initials: M
authors.lastname: Hamreus
authors.forename: F C
authors.initials: FC
authors.lastname: Liu
authors.forename: C
authors.initials: C
authors.lastname: Herdon-Remelius
authors.forename: W
authors.initials: W
authors.lastname: Junger
authors.forename: A
authors.initials: A
authors.lastname: Altman
mesh_terms: D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000096:biosynthesis*; D002455:Cell Division / Q000187:drug effects; D004789:Enzyme Activation; D006801:Humans; D007376:Interleukin-2 / Q000096:biosynthesis; D019169:Jurkat Cells / Q000187:drug effects / Q000201:enzymology*; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000201:enzymology*; D020928:Mitogen-Activated Protein Kinases; D010766:Phosphorylation; D016133:Polymerase Chain Reaction; D011505:Protein-Tyrosine Kinases / Q000096:biosynthesis*; D012462:Saline Solution, Hypertonic / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects; D014443:Tyrosine / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases
chemical_list: D007376:Interleukin-2; D012462:Saline Solution, Hypertonic; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases