Background  Enteric glial cells (EGCs) have been recently indicated as key regulators of intestinal inflammation in animals. Whether or not this is true and how these cells participate to inflammatory responses in humans is unknown.

Methods  We isolated primary EGCs from human small bowel and then, we purified and characterized those using specific glial markers, such as S100B and glial fibrillary acidic protein (GFAP). To mimic an inflammatory scenario, we exposed EGCs to exogenous stimuli, such as lipopolysaccharide and interferon‐gamma (LPS and IFN‐γ), alone or in combination, to evaluate glial activation [measuring GFAP, S100B level together with c‐fos, major histocompatibility complex (MHC) class II, inducible nitric oxide (iNOS) proteins expression and nitric oxide (NO) production] and proliferation, respectively.

Key Results  We showed that, when challenged with a combination of LPS and IFN‐γ, EGCs are significantly activated, as indicated by their positivity to c‐fos and MHC class II. Similarly, pro‐inflammatory stimuli significantly increase the cell proliferation rate, the expression of both S100B and GFAP, and the NO production consequent to the induction of EGCs‐derived iNOS protein, with the last being dependent on S100B‐RAGE (receptor for advanced glycation endproducts) interaction.

Conclusions & Inferences  Our data provide the first evidence that human EGCs directly respond to pro‐inflammatory stimuli by changing their expression profile and by proliferating. The finding that stimulated EGCs are able to produce NO points to a role of this cell population in the scenario of intestinal inflammation.