The spatiotemporal development of mesenteric lymphatic changes in the TNF^ΔARE/+ mouse model of terminal ileitis

Crohn's disease (CD) is a chronic inflammatory bowel disease, which also encompasses significant alterations of the mesenteric lymphatic system. Whether these changes are a mere consequence of or directly contribute to the inflammation is unknown. Here, we characterized the spatial and temporal development of these events in the TNF^ΔARE/+ mouse, which develops CD-like ileitis and significant mesenteric lymphatic alterations. At 8, 12, 20, and 28 wk of age, specific pathogen-free (SPF), germ-free (GF) TNF^ΔARE/+ and wild-type (WT) mice were assessed for ileitis via myeloperoxidase (MPO) activity while mesenteric lymphatic alterations were assessed by confocal immunofluorescence imaging. Lymphatic alterations in the SPF TNF^ΔARE/+ occurred in a stepwise manner between 8 and 28 wk of age beginning with the development of mesenteric lymphadenopathy at 8 wk despite no significant ileitis. By 12-wk ileal MPO significantly elevates concomitantly with lymphangiectasia of the mesenteric collecting lymphatic vessels (CLVs) and clustering of CD45⁺ immune cells around them. At 20 wk, significant lymphangiogenesis of the initials (initial lymphatic vessel) and tertiary lymphoid organs aligned along lymphatic collectors (CA-TLOs) had developed. At 28 wk, lymphangiectasia, lymphangiogenesis, and CA-TLOs increased. However, 28-wk-old GF TNF^ΔARE/+, while displaying no ileitis, presented with mesenteric lymphadenopathy, lymphangiectasia, and lymphangiogenesis but no immune cell clustering nor CA-TLOs. The TNF^ΔARE/+ mice develop terminal ileitis and lymphatic alterations in a stepwise manner beginning with mesenteric lymph node lymphadenopathy and ileal inflammation, followed by CLV dilation and lymphangiogenesis. These lymphatic alterations are exacerbated by the gut microbiome, with immune cell clustering and tertiary lymphoid organ formation being entirely dependent of its presence.NEW & NOTEWORTHY The mesenteric lymphatic system displays striking morphological alterations in Crohn's disease. To assess the importance of these changes in the perpetuation of the disease, we established the timeframe of their occurrence with respect to the development of ileitis in a mouse model of Crohn's disease and in the same model derived germ-free where intestinal inflammation does not occur. Although immune-related alterations seem to depend on microbiome, changes specifically affecting lymphatic vessels persist in its absence.