Dietary limonene promotes gastrointestinal barrier function via upregulating tight/adherens junction proteins through cannabinoid receptor type-1 antagonistic mechanism and alters cellular metabolism in intestinal epithelial cells

• Published in: BioFactors (Oxford) Vol. 51; no. 1; p. e2106
• Main Authors: Senthil Kumar, K J, Gokila Vani, M, Dakpa, Gyaltsen, Wang, Sheng-Yang
• Format: Journal Article
• Language: English
• Published: Netherlands 01.01.2025
• Subjects: Adherens Junctions - drug effects; Adherens Junctions - metabolism; Caco-2 Cells; Epithelial Cells - drug effects; Epithelial Cells - metabolism; Humans; Intestinal Mucosa - drug effects; Intestinal Mucosa - metabolism; Limonene - metabolism; Limonene - pharmacology; Receptor, Cannabinoid, CB1 - antagonists & inhibitors; Receptor, Cannabinoid, CB1 - genetics; Receptor, Cannabinoid, CB1 - metabolism; Tight Junction Proteins - genetics; Tight Junction Proteins - metabolism; Tight Junctions - drug effects; Tight Junctions - metabolism; Up-Regulation - drug effects; Zonula Occludens-1 Protein - genetics; Zonula Occludens-1 Protein - metabolism; limonene; intestinal epithelial cells; adherence junction; metabolites; tight junction; CB1R
• DOI: 10.1002/biof.2106

Limonene, a dietary monocyclic monoterpene commonly found in citrus fruits and various aromatic plants, has garnered increasing interest as a gastrointestinal protectant. This study aimed to assess the effects of limonene on intestinal epithelial barrier function and investigate the involvement of cannabinoid receptor type-1 (CB1R) in vitro. Additionally, the study focused on examining the metabolomic changes induced by limonene in the intestinal epithelial cells (Caco-2). Initial analysis of transepithelial electrical resistance (TEER) revealed that both l-limonene and d-limonene, isomers of limonene, led to a dose- and time-dependent increase in TEER in normal cells and those inflamed by pro-inflammatory cytokines mixture (CytoMix). Furthermore, both types of limonene reduced CytoMix-induced paracellular permeability, as demonstrated by a decrease in Lucifer yellow flux. Moreover, d-limonene and l-limonene treatment increased the expression of tight junction molecules (TJs) such as occludin, claudin-1, and ZO-1, at both the transcriptional and translational levels. d-Limonene upregulates E-cadherin, a molecule involved in adherens junctions (AJs). Mechanistic investigations demonstrated that d-limonene and l-limonene treatment significantly inhibited CB1R at the protein, while the mRNA level remained unchanged. Notably, the inhibitory effect of d-limonene on CB1R was remarkably similar to that of pharmacological CB1R antagonists, such as rimonabant and ORG27569. d-limonene also alters Caco-2 cell metabolites. A substantial reduction in β-glucose and 2-succinamate was detected, suggesting limonene may impact intestinal epithelial cells' glucose uptake and glutamate metabolism. These findings suggest that d-limonene's CB1R antagonistic property could effectively aid in the recovery of intestinal barrier damage, marking it a promising gastrointestinal protectant.