Intestinal P-glycoprotein exports endocannabinoids to prevent inflammation and maintain homeostasis
Rose L. Szabady, … , Randall J. Mrsny, Beth A. McCormick
Rose L. Szabady, … , Randall J. Mrsny, Beth A. McCormick
Published August 13, 2018
Citation Information: J Clin Invest. 2018;128(9):4044-4056. https://doi.org/10.1172/JCI96817.
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Research Article Gastroenterology

Intestinal P-glycoprotein exports endocannabinoids to prevent inflammation and maintain homeostasis

Abstract

Neutrophil influx into the intestinal lumen is a critical response to infectious agents, but is also associated with severe intestinal damage observed in idiopathic inflammatory bowel disease. The chemoattractant hepoxilin A3, an eicosanoid secreted from intestinal epithelial cells by the apically restricted efflux pump multidrug resistance protein 2 (MRP2), mediates this neutrophil influx. Information about a possible counterbalance pathway that could signal the lack of or resolution of an apical inflammatory signal, however, has yet to be described. We now report a system with such hallmarks. Specifically, we identify endocannabinoids as the first known endogenous substrates of the apically restricted multidrug resistance transporter P-glycoprotein (P-gp) and reveal a mechanism, which we believe is novel, for endocannabinoid secretion into the intestinal lumen. Knockdown or inhibition of P-gp reduced luminal secretion levels of N-acyl ethanolamine–type endocannabinoids, which correlated with increased neutrophil transmigration in vitro and in vivo. Additionally, loss of CB2, the peripheral cannabinoid receptor, led to increased pathology and neutrophil influx in models of acute intestinal inflammation. These results define a key role for epithelial cells in balancing the constitutive secretion of antiinflammatory lipids with the stimulated secretion of proinflammatory lipids via surface efflux pumps in order to control neutrophil infiltration into the intestinal lumen and maintain homeostasis in the healthy intestine.

Authors

Rose L. Szabady, Christopher Louissaint, Anneke Lubben, Bailu Xie, Shaun Reeksting, Christine Tuohy, Zachary Demma, Sage E. Foley, Christina S. Faherty, Alejandro Llanos-Chea, Andrew J. Olive, Randall J. Mrsny, Beth A. McCormick

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Figure 6

CB2-deficient mice are vulnerable to severe intestinal inflammation with increased neutrophil transmigration.

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CB2-deficient mice are vulnerable to severe intestinal inflammation with...
WT or Cnr2–/– mice were treated with 3% DSS for 7 days and sacrificed at day 9. (A) Weights are shown as percentage of day 0 (D0) weight. n = 25 WT; n = 24 Cnr2–/– mice. **P < 0.01; ***P < 0.001. (B and C) Histopathology of mid and distal colon as in Figure 1. Arrows highlight accumulation of neutrophils in intestinal lumen. *P = 0.021. Original magnification, ×20. (D) Fecal MPO activity. n = 14 WT; n = 12 Cnr2–/– mice. **P = 0.008. (E) Tissue MPO activity. n = 13 WT; n = 11 Cnr2–/– mice. NS, P = 0.471. (F) Number (NS, P = 0.482) and (G) percentage (NS, P = 0.095) of tissue neutrophils by flow cytometry analysis. (H) Representative plots of lamina propria neutrophils. (F–H) n = 13 WT; n = 12 Cnr2–/– mice. (I–K) WT or Cnr2–/– mice were lethally irradiated and reconstituted with homologous or heterologous BM (donor→recipient). Following reconstitution for 7 weeks, mice were treated with DSS as above. (I) Weight curve. (J) Colon length. (K) Histopathology score. n = 7 WT→WT; n = 9 KO→WT; n = 9 WT→KO; n = 7 KO→KO mice. For all experiments, data are shown as mean ± SEM. Statistical analysis was performed with Mann-Whitney 1-tailed nonparametric U test. Histopathology scoring, MPO activity measurement, and flow cytometry analyses were performed as in Figure 1.