Expression and regulation of procalcitonin in different human cells
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KeywordsReverse Transcriptase Polymerase Chain Reaction Procalcitonin Peripheral Blood Monocyte Liver Parenchymal Cell Primary Human Cell
Procalcitonin (PCT) was recently forwarded as a diagnostic marker of systemic bacterial infection and sepsis. The biological function(s) and biochemical properties of this protein are poorly defined and the cellular sources of plasma PCT remains yet to be established.
Primary human cells (peripheral blood monocytes, umbilical vein endothelial cells) and cell lines (liver, renal parenchymal and lung fibroblastic lines) were cultivated under standard conditions. Basal and stimulated mRNA expression of PCT was investigated using a semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR). Intracellular PCT protein expression was verified by Western blotting and surface-enhanced laser desorption/ionization (SELDI). Experiments elucidating the intracellular location of PCT were performed after protein fragmentation in different fractions by secondary immunofluorescence and laser scan confocal microscopy.
(1 )A basal and inducible mRNA expression of PCT was found only in human peripheral blood monocytes. (2) In these cells, a distinct influence of various proinflammatory mediators was observed. (3) Western blotting of monocyte lysates using various primary antibodies directed against PCT showed a strong intracellular protein expression. (4) Experiments with SELDI revealed a molecular weight for PCT in monocytes of 12.1 kDa. (5) Human monocytes express PCT protein in association with cytoskeleton. No PCT was found in cytoplasmic fractions.
Since human peripheral blood monocytes produce PCT and its expression depends strongly from sepsis-related mediators, we conclude, that this cell population is one important source of elevated PCT serum levels during sepsis. Further experiments analyzing the role of Kupffer cells and liver parenchymal cells are in progress.