TNF-α augments RANKL-dependent intestinal M cell differentiation in enteroid cultures.

Area of Science:

• Immunology
• Gastroenterology
• Cell Biology

Background:

• Microfold (M) cells are specialized intestinal epithelial cells crucial for antigen transport in Peyer's patches.
• M cell differentiation from stem cells depends on RANKL and Spi-B.
• Understanding M cell differentiation pathways is key to gut immunity research.

Purpose of the Study:

• To investigate signaling pathways governing M cell differentiation using a 3D enteroid model.
• To explore the influence of cytokines, particularly TNF-α, on RANKL-induced M cell differentiation.

Main Methods:

• Utilized three-dimensional (3D) enteroid cultures derived from mouse small intestinal crypts.
• Administered RANKL and TNF-α to enteroids to assess M cell-associated gene expression.
• Employed enteroids from Map3k14 knockout mice to investigate the role of the noncanonical NF-κB pathway.

Main Results:

• RANKL alone induced M cell-associated genes (Spib, Ccl9, Tnfaip2, Anxa5, Marcksl1) within 24 hours.
• The mature M cell marker Gp2 was significantly induced by day 3, present in 11% of enteroid cells.
• TNF-α synergistically enhanced RANKL-induced M cell differentiation, increasing gene expression 3-6 fold, partly via induction of Relb and Nfkb2.

Conclusions:

• The noncanonical NF-κB pathway is essential for RANKL-mediated M cell differentiation in enteroids.
• TNF-α acts as a supportive cytokine, significantly boosting RANKL-driven M cell differentiation.
• Gut-associated lymphoid tissue microenvironment factors like TNF-α play a crucial role in M cell development.