title: Novel Approach in Monocyte Intracellular TNF Measurement: Application to Sepsis-Induced Immune Alterations.
abstract: The monitoring of septic shock induced immunosuppression has been proposed to identify patients who could benefit from specific immunoadjuvant therapies. Among potential biomarkers to monitor immunological status, functional testing remains the gold standard because it directly measures the capacity of a cell population to respond to an immune challenge. We investigated a new approach in intracellular staining for flow cytometry to measure tumor necrosis factor (iTNF) produced in vitro by monocytes in response to lipopolysaccharide.
  
  
  
  Observational study in intensive care unit and immunology laboratory of a university medical center. Sixteen septic shock patients and eight control subjects were included.
  
  
  
  Monocyte iTNF was determined by flow cytometry in whole blood and completed in 2.5 h according to a no-wash, no centrifuge procedure. Lipopolysaccharide challenge induced a tremendous expression of iTNF that was statistically more pronounced in controls than in patients. This was observed when results were expressed as medians of fluorescence intensity (median: 16.1 [IQR: 14.5-19.1] vs. 5 [4.0-8.0], P = 0.0001) or as percentages of positive cells (99.7 [99.6-99.8] vs. 85 [74-97], P = 0.0001). iTNF expression was correlated to monocyte HLA-DR expression in patients and controls.
  
  
  
  These preliminary results illustrate the feasibility of immune functional testing on a routine manner in septic shock patients. They now deserve to be widely assessed and validated in various intensive care unit conditions. This could be a major step to characterize the rapidly changing immune response overtime and thus permit personalized medicine.
authors: *Immunology Laboratory, Hospices Civils de Lyon, Hôpital Edouard Herriot, Lyon, France †EA 7426 PI3 "Pathophysiology of Injury-induced Immunosuppression" (Université Claude Bernard Lyon I-Hospices Civils de Lyon-bioMérieux), Lyon, France ‡TRIGGERSEP (TRIal Group for Global Evaluation and Research in SEPsis), F-CRIN Network, Limoges, France §Beckman Coulter Immunotech, Life Sciences Global Assay and Applications Development, Marseille, France ||Department of Anesthesiology, Hôpital Edouard Herriot, Hospices Civils de Lyon, Lyon, France.
authors: Guillaume
authors: G
authors: Monneret
authors: Julie
authors: J
authors: Demaret
authors: Morgane
authors: M
authors: Gossez
authors: Elsa
authors: E
authors: Reverdiau
authors: Fabrice
authors: F
authors: Malergue
authors: Thomas
authors: T
authors: Rimmelé
authors: Fabienne
authors: F
authors: Venet
mesh_terms: D000276:Adjuvants, Immunologic; D000368:Aged; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D007108:Immune Tolerance; D008070:Lipopolysaccharides / Q000633:toxicity; D008297:Male; D008875:Middle Aged; D009000:Monocytes / Q000378:metabolism*; D018805:Sepsis / Q000097:blood* / Q000276:immunology*; D012772:Shock, Septic / Q000097:blood / Q000276:immunology; D014409:Tumor Necrosis Factor-alpha / Q000097:blood / Q000378:metabolism*
chemical_list: D000276:Adjuvants, Immunologic; D008070:Lipopolysaccharides; D014409:Tumor Necrosis Factor-alpha