Human TNF-Luc reporter mouse: A new model to quantify inflammatory responses.

Area of Science:

• Immunology
• Molecular Biology
• Genetics

Background:

• Tumour necrosis factor (TNF) is a critical cytokine in inflammatory processes.
• Dysregulation of TNF contributes to inflammatory diseases like rheumatoid arthritis and inflammatory bowel disease.
• Novel tools are needed to study TNF regulation and develop targeted therapies.

Purpose of the Study:

• To generate and validate a novel transgenic mouse model (hTNF.LucBAC) for studying human TNF (hTNF) gene regulation.
• To assess the utility of the hTNF.LucBAC strain as a reporter system for inflammatory responses.
• To investigate the regulation of TNF synthesis in response to various stimuli.

Main Methods:

• Generation of a transgenic mouse strain (hTNF.LucBAC) using a bacterial artificial chromosome (BAC) construct linking luciferase reporter to the human TNF locus.
• In vitro stimulation of hTNF.LucBAC cells (macrophages, intestinal organoids) with lipopolysaccharide (LPS) and TNF.
• Measurement of luciferase activity as a proxy for TNF gene expression.
• Assessment of regulatory effects using NF-κB pathway inhibitors and Interleukin-10 (IL-10).
• Correlation analysis between transgene-dependent luciferase activity and endogenous soluble TNF levels.

Main Results:

• The hTNF.LucBAC strain demonstrated cell-specific responses to stimuli, validating its use as an inflammatory gene response proxy.
• Lipopolysaccharide (LPS) was a potent inducer in macrophages; TNF strongly activated intestinal organoids.
• Lipopolysaccharide-induced luciferase activity in macrophages was suppressed by NF-κB inhibitors and IL-10.
• Transgene-dependent luciferase activity positively correlated with secreted murine soluble TNF.

Conclusions:

• The hTNF.LucBAC transgenic mouse is a valuable tool for studying TNF synthesis and screening therapeutic molecules.
• This model facilitates functional studies of the regulatory elements within the TNF locus.
• The findings highlight the utility of reporter gene systems for investigating inflammatory pathways.