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Published on: April 29, 2015
Differential regulation of TSG-14 expression in murine fibroblasts and peritoneal macrophages
A R Goodman1, D E Levy, L F Reis
1Department of Microbiology, New York University School of Medicine, NY 10016, USA.
Abstract:
Tumor necrosis factor (TNF)-stimulated gene 14 (TSG-14, also termed PTX3) encodes a secreted glycoprotein whose carboxy-terminal half shares sequence similarity with the pentraxin family of acute phase proteins (C-reactive protein and serum amyloid P component). We compared TSG-14 mRNA expression in cultures of murine BALB/c 3T3 fibroblasts and thioglycollate-elicited peritoneal macrophages. TNF and interleukin-1 (IL-1) potently induced TSG-14 expression in 3T3 fibroblasts but not in peritoneal macrophages. Lipopolysaccharide (LPS) elicited TSG-14 expression in both cell types, but induction in 3T3 cells and macrophages showed several distinct characteristics. Whereas in 3T3 fibroblasts TSG-14 mRNA was rapidly up-regulated by LPS, expression in macrophages was substantially delayed. Furthermore, cycloheximide greatly reduced LPS-induced TSG-14 mRNA up-regulation in macrophages but not in 3T3 cells. Finally, interferon-gamma (IFN-gamma; but not IFN-alpha/beta) inhibited LPS-induced TSG-14 expression in macrophages and not in 3T3 fibroblasts. The antioxidant pyrrolidine dithiocarbamate inhibited LPS-induced nuclear factor-kappaB (NF-kappaB) activation and TSG-14 expression in macrophages. In contrast, IFN-gamma did not inhibit NF-kappaB function as measured by IkappaB-alpha and IkappaB-beta degradation, IkappaB-alpha resynthesis, or electrophoretic mobility shift analysis. Inhibition of LPS-induced TSG-14 mRNA expression by IFN-gamma in macrophages was also observed in the presence of cycloheximide and in cells from STAT1 null mice, suggesting that IFN-gamma inhibits TSG-14 expression through an unconventional mechanism.
Insights
Tumor necrosis factor (TNF)-stimulated gene 14 (TSG-14) expression differs between fibroblasts and macrophages. Interferon-gamma (IFN-gamma) uniquely inhibits TSG-14 in macrophages via an unconventional pathway.
Area of Science:
- Immunology
- Molecular Biology
- Cell Biology
Background:
- TSG-14 (PTX3) is a secreted glycoprotein with sequence similarity to pentraxin acute phase proteins.
- Understanding differential gene expression in immune and non-immune cells is crucial for deciphering inflammatory responses.
- Cytokine and microbial stimuli differentially regulate gene expression in various cell types.
Purpose of the Study:
- To compare TSG-14 mRNA expression patterns in murine 3T3 fibroblasts and peritoneal macrophages.
- To investigate the distinct regulatory mechanisms of TSG-14 induction by TNF, IL-1, LPS, and IFN-gamma in these cell types.
- To elucidate the signaling pathways involved in LPS-induced TSG-14 expression and its modulation by IFN-gamma.
Main Methods:
- Quantitative analysis of TSG-14 mRNA expression using cell cultures (3T3 fibroblasts, peritoneal macrophages).
- Stimulation with cytokines (TNF, IL-1, IFN-gamma) and lipopolysaccharide (LPS).
- Assessment of protein synthesis inhibition (cycloheximide) and nuclear factor-kappaB (NF-kappaB) pathway activation (electrophoretic mobility shift assays, IkappaB degradation).
Main Results:
- TNF and IL-1 induced TSG-14 in fibroblasts but not macrophages.
- LPS induced TSG-14 in both cell types, with delayed kinetics and cycloheximide sensitivity in macrophages.
- IFN-gamma inhibited LPS-induced TSG-14 in macrophages via a mechanism independent of NF-kappaB activation and STAT1.
Conclusions:
- TSG-14 regulation is cell-type specific, with distinct responses in fibroblasts versus macrophages.
- IFN-gamma exerts an unconventional inhibitory effect on LPS-induced TSG-14 expression in macrophages.
- These findings highlight complex regulatory networks governing acute phase protein expression in innate immunity.
