In vitro cytokine mRNA expression in normal human peripheral blood mononuclear cells.
OBJECTIVE
Normal human peripheral blood mononuclear cells (PBMC) were analyzed for basic profiles of mRNA expression of distinct genes during incubation in a standard cell culture system.


METHODS
Human PBMC from healthy adult blood donors.


METHODS
Steady-state mRNA expression was measured using a semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) assay.


RESULTS
Shortly after isolation and cell seeding, mRNA levels of monocyte-derived cytokines (IL-1alpha, IL-6, and TNF-alpha) were significantly increased, whereas lymphocyte-derived cytokines (IL-4, IFN-gamma) were not affected. Expression levels of monokines returned to basal within 24 h. At later stages of culture, the mRNA levels of all genes studied gradually increased and were significantly elevated after 96 h of incubation.


CONCLUSIONS
Monokine and lymphokine mRNAs respond differently to cell culture even under control conditions. With regard to the enhanced mRNA expression of distinct cytokines in the early stages of culture, human PBMC should not be used for gene expression studies in vitro within 24 h of isolation.
Institute of Hygiene, University of Graz, Austria. sonja.barth@kfunigraz.ac.at
S
S
Barth
B
B
Kleinhappl
A
A
Gutschi
S
S
Jelovcan
E
E
Marth
D000199:Actins / Q000235:genetics; D000328:Adult; D002478:Cells, Cultured; D016207:Cytokines / Q000235:genetics*; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000235:genetics; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D007963:Leukocytes, Mononuclear / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012269:Ribosomal Proteins / Q000235:genetics
D000199:Actins; D016207:Cytokines; D012333:RNA, Messenger; D012269:Ribosomal Proteins; C035201:ribosomal protein L30; D018799:Intercellular Adhesion Molecule-1; D007770:L-Lactate Dehydrogenase