Detection of Intracellular Tumor Necrosis Factor α in Stimulated Fetal Cells

Autor: John T. Johnson, Dorothea J. Eicher, James A. Cook, Andre Hebra, Carol L. Wagner, Ramona Kramer
Rok vydání: 1999
Předmět:
Zdroj: Journal of Surgical Research. 82:300-304
ISSN: 0022-4804
Popis: Background. It is unknown if immature fetal cells produce tumor necrosis factor (TNF) α in the same manner that adult cells do. The aim of this study was to determine the feasibility of early detection of intracellular TNF produced by circulating human monocytes (Mo) and lymphocytes (Ly) using flow cytometry and to compare the stimulation profiles of mature and fetal cells. Material and methods. Fetal umbilical cord blood ( n = 10) and adult volunteer blood ( n = 10) were obtained. In vitro stimulation with endotoxin (LPS) and ionomycin-PMA was performed. Brefeldin A was added to prevent extracellular transport of TNF. Cell type was determined by using CD-14 marker separating monocyte and lymphocyte populations. Anti-human TNF monoclonal antibody was used to detect intracellular TNF by flow cytometry analysis. Results. Thirty to sixty thousand cells were analyzed per sample. Average TNF expression of stimulated fetal Mo was 28.2%, and that of fetal Ly was only 1.1%. Adult stimulated Ly had an average TNF expression of 31.9%, and adult Mo, 29.6% ( P Conclusion. TNF flow cytometry analysis allows assessment of individual cell types and their ability to produce that cytokine. Fetal cells are able to produce TNF when stimulated, but the stimulation profile of Ly differs from that of adult samples. This observation may be of clinical importance in evaluating the response of immature cells to a septic stimulus. Flow cytometry is reliable, reproducible, quick, and easily obtained from a small sample of peripheral blood. Clinical use will be applicable once appropriate controls are developed, as reported in this study.
Databáze: OpenAIRE
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