Lymphatic deletion of calcitonin receptor–like receptor exacerbates intestinal inflammation
Reema B. Davis, Daniel O. Kechele, Elizabeth S. Blakeney, John B. Pawlak, Kathleen M. Caron
Reema B. Davis, Daniel O. Kechele, Elizabeth S. Blakeney, John B. Pawlak, Kathleen M. Caron
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Research Article Gastroenterology

Lymphatic deletion of calcitonin receptor–like receptor exacerbates intestinal inflammation

Abstract

Lymphatics play a critical role in maintaining gastrointestinal homeostasis and in the absorption of dietary lipids, yet their roles in intestinal inflammation remain elusive. Given the increasing prevalence of inflammatory bowel disease, we investigated whether lymphatic vessels contribute to, or may be causative of, disease progression. We generated a mouse model with temporal and spatial deletion of the key lymphangiogenic receptor for the adrenomedullin peptide, calcitonin receptor–like receptor (Calcrl), and found that the loss of lymphatic Calcrl was sufficient to induce intestinal lymphangiectasia, characterized by dilated lacteals and protein-losing enteropathy. Upon indomethacin challenge, Calcrlfl/fl/Prox1-CreERT2 mice demonstrated persistent inflammation and failure to recover and thrive. The epithelium and crypts of Calcrlfl/fl/Prox1-CreERT2 mice exhibited exacerbated hallmarks of disease progression, and the lacteals demonstrated an inability to absorb lipids. Furthermore, we identified Calcrl/adrenomedullin signaling as an essential upstream regulator of the Notch pathway, previously shown to be critical for intestinal lacteal maintenance and junctional integrity. In conclusion, lymphatic insufficiency and lymphangiectasia caused by loss of lymphatic Calcrl exacerbates intestinal recovery following mucosal injury and underscores the importance of lymphatic function in promoting recovery from intestinal inflammation.

Authors

Reema B. Davis, Daniel O. Kechele, Elizabeth S. Blakeney, John B. Pawlak, Kathleen M. Caron

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Figure 5

Impaired lipid uptake and junctional barriers in lymphatic Calcrl-deficient mouse intestines.

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Impaired lipid uptake and junctional barriers in lymphatic Calcrl-defici...
(A) Images of tamoxifen-treated (TAM-treated) Calcrlfl/fl and Calcrlfl/fl/Prox1-CreERT2 animals 7 days after indomethacin (INDO) challenge showing lean Calcrlfl/fl/Prox1-CreERT2 animals. (B) Image of TAM-treated Calcrlfl/fl/Prox1-CreERT2 mouse abdomen and mesenteric region 7 days after INDO challenge. Arrow points to chyle leakage. (C) Measurement of weight loss in TAM-treated Calcrlfl/fl and Calcrlfl/fl/Prox1-CreERT2 mice 1 day after INDO and 7 days after INDO. Quantitative data are represented as a box-and-whisker plot, with bounds from 25th to 75th percentile, median line, and whiskers ranging from minimum to maximum percentage weight loss; n = 7–10 animals per group per treatment. Significance was determined by 2-way ANOVA, *P < 0.05. (D) Representative images of Oil red O–stained duodena of TAM-treated Calcrlfl/fl and Calcrlfl/fl/Prox1-CreERT2 mice 7 days after INDO challenge. n = 7–8 animals in each group. Scale bar: 25 μm. Boxed regions are shown as zoomed insets to the right. Magnification: ×25. Arrows highlight lipid droplets inside the villi or gathered at the top of the villi. (E) Representative images of VE-Cadherin– and lymphatic vessel endothelial hyaluronan receptor 1–stained (LYVE1-stained) duodena of TAM-treated Calcrlfl/fl and Calcrlfl/fl/Prox1-CreERT2 mice 1 day after INDO challenge. n = 8–10 animals in each group. Boxed regions highlight lacteals. Scale bar: 50 μm. (F) Representative VE-Cadherin– and zona occludens 1–stained (ZO-1–stained) images from 3 independent experiments in which human neonatal dermal lymphatic endothelial cells were treated with control siRNA or human calcitonin receptor–like receptor (CALCRL) siRNA. Scale bar: 100 μm. Arrows indicate junctions.