ABSTRACT
OBJECTIVE: The aim of the present study was to investigate the capacity of normal immune blood cells from non-Hodgkin's lymphoma patients to produce tumor necrosis factor (TNF) after lipopolysaccharide (LPS) stimulation and the influence of the TNF (-308) polymorphism in this production. MATERIALS AND METHODS: A whole peripheral blood cell assay was utilized following LPS stimulation. At selected incubation times, supernatants were harvested for protein dosage, while mRNA was extracted and reverse-transcribed. The amount of TNF mRNA was quantified using real-time quantitative polymerase chain reaction (PCR) and genomic DNA was typed for TNF (-308) polymorphism. RESULTS: Upon LPS stimulation, TNF-secreted protein was slightly but not significantly increased in lymphoma patients when compared to controls. In contrast, the relative TNF mRNA amounts were significantly higher in lymphoma patients at 30 minutes (median 27.75 vs. 16.00; Mann-Whitney U-test p < 0.05), at 4 hours (52.00 vs. 31.00; p < 0.05), and at 24 hours (19.50 vs. 9.00; p < 0.05). In addition, patients carrying the variant TNF2 allele had higher relative TNF mRNA levels than TNF1 homozygotes (p = 0.02). CONCLUSION: The LPS-induced TNF mRNA levels are higher in peripheral blood cells (PBC) from lymphoma patients than from controls, while TNF protein secretion is not strikingly different. Altered regulation of TNF mRNA translation or TNF protein secretion may contribute to these observations. Taken together, an increased susceptibility for TNF gene transcription after LPS stimulation was observed in PBC (mainly in monocytes) from lymphoma patients, and especially those carrying the TNF2 allele.
Subject(s)
Gene Expression Regulation, Neoplastic/drug effects , Leukocytes, Mononuclear/chemistry , Lipopolysaccharides/pharmacology , Lymphoma, Non-Hodgkin/blood , Neoplasm Proteins/genetics , RNA, Messenger/blood , RNA, Neoplasm/blood , Tumor Necrosis Factor-alpha/genetics , Adult , Aged , Alleles , DNA, Complementary/genetics , Dactinomycin/pharmacology , Genotype , Humans , Leukocytes, Mononuclear/metabolism , Lymphocyte Activation/drug effects , Lymphoma, Non-Hodgkin/genetics , Middle Aged , Neoplasm Proteins/biosynthesis , Neoplasm Proteins/metabolism , Nucleic Acid Synthesis Inhibitors/pharmacology , Polymerase Chain Reaction , Tumor Necrosis Factor-alpha/biosynthesis , Tumor Necrosis Factor-alpha/metabolismABSTRACT
The pathogenesis of septic shock is mainly due to unregulated tumour necrosis factor-alpha (TNF-alpha) production. Procalcitonin (PCT) and calcitonin gene-related peptide (CGRP) are alternative transcription products of the calcitonin gene. Since high PCT levels have been described in human sepsis, and since CGRP inhibits TNF synthesis in rats, we examined the role of these peptides in the regulation of the inflammatory response during septic shock. LPS-induced TNF production was assessed using a human whole blood model. In this model, PCT (10(-7) M) and CGRP (10(-6) M) significantly inhibit TNF production by 27 and 24 % respectively. The effect of CGRP was reversed by CGRP 8-37 (10 microM), an antagonist of CGRP receptor. No effect on interleukin (IL)-1, IL-6 and IL-8 was found. This is the first description of an anti-inflammatory role for PCT and CGRP in humans.
