Optimizing THP-1 Macrophage Culture for an Immune-Responsive Human Intestinal Model

Area of Science:

• Immunology
• Gastroenterology
• Cell Biology

Background:

• Existing macrophage co-culture models face limitations due to macrophage dedifferentiation in long-term cultures.
• Developing stable, long-term immune-responsive models is crucial for studying chronic intestinal conditions.

Purpose of the Study:

• To establish and validate a novel, long-term (21-day) triple co-culture model involving THP-1 macrophages (THP-1m), Caco-2 intestinal epithelial cells, and HT-29-methotrexate goblet cells.
• To assess the stability and immune responsiveness of differentiated THP-1m within this co-culture system under inflammatory conditions.

Main Methods:

• High-density THP-1 cells were differentiated into THP-1m using phorbol 12-myristate 13-acetate and cultured for up to 21 days.
• A triple co-culture system was established with Caco-2 and HT-29-MTX cells.
• Lipopolysaccharide was used to induce an inflammatory state, and cytokine levels (TNF-α, IL-6) and transepithelial electrical resistance (TEER) were measured.

Main Results:

• Differentiated THP-1m exhibited stable morphology and lysosome expansion throughout the 21-day culture period.
• The triple co-culture model demonstrated significant increases in Tumor Necrosis Factor-alpha (824.7 ± 130.0 pg/mL) and Interleukin-6 (609.7 ± 139.5 pg/mL) upon lipopolysaccharide stimulation.
• Intestinal membrane integrity was successfully maintained, indicated by a TEER value of 336.4 ± 18.0 Ω·cm².

Conclusions:

• THP-1m can be effectively utilized in a stable, long-term triple co-culture model for studying intestinal immune responses.
• This model is suitable for investigating both normal and chronic inflammatory states of the intestinal epithelium.
• The developed model offers a valuable tool for future research on the interplay between the immune system and gut health.