Lactobacillus plantarum (L. plantarum) is a probiotic that has emerged as novel therapeutic agents for managing various diseases, such as cancer, atopic dermatitis, inflammatory bowel disease, and infections. In this study, we investigated the potential mechanisms underlying the anticancer effect of the metabolites of L. plantarum. We cultured L. plantarum cells to obtain their metabolites, created several dilutions, and used these solutions to treat human colonic Caco-2 cells. Our results showed a 10% dilution of L. plantarum metabolites decreased cell viability and reduced the expression of autophagy-related proteins. Moreover, we found co-treatment with L. plantarum metabolites and chloroquine, a known autophagy inhibitor, had a synergistic effect on cytotoxicity and downregulation of autophagy-related protein expression. In conclusion, we showed the metabolites from the probiotic, L. plantarum, work synergistically with chloroquine in killing Caco-2 cells and downregulating the expression of autophagy-related proteins, suggesting the involvement of autophagy, rather than apoptosis, in their cytotoxic effect. Hence, this study provides new insights into new therapeutic methods via inhibiting autophagy.

Protease-activated receptor 2 (PAR2) alleviates intestinal inflammation by upregulating autophagy. PAR2 also modulates tight junctions through β‑arrestin signaling. Therefore, we investigated the effect of PAR2-induced autophagy on intestinal epithelial tight junctions and permeability. RT-PCR, Western blot analysis, and immunoprecipitation were performed to investigate the underlying molecular mechanisms by which PAR2 regulates autophagy and intestinal epithelial tight junctions. Inhibition of PAR2 by GB83, a PAR2 antagonist, decreased the expression of autophagy-related and tight-junction-related factors in Caco-2 cells. Moreover, inhibition of PAR2 decreased intestinal transepithelial electrical resistance. When PAR2 was activated, intestinal permeability was maintained, but when autophagy was suppressed by chloroquine, intestinal permeability was significantly increased. In addition, the prolongation of ERK1/2 phosphorylation by PAR2–ERK1/2–β-arrestin assembly was reduced under autophagy inhibition conditions. Therefore, PAR2 induces autophagy to regulate intestinal epithelial permeability, suggesting that it is related to the β-arrestin–ERK1/2 pathway. In conclusion, regulating intestinal epithelial permeability through PAR2-induced autophagy can help maintain mucosal barrier integrity. Therefore, these findings suggest that the regulation of PAR2 can be a suitable strategy to treat intestinal diseases caused by permeability dysfunction.

Atractylodin is a major compound in the rhizome of Atractylodes lancea, an oriental herbal medicine used for the treatment of gastrointestinal diseases, including dyspepsia, nausea, and diarrhea. Recent studies have shown that atractylodin exerts anti-inflammatory effects in various inflammatory diseases. Herein, we investigated the anti-colitis effects of atractylodin and its molecular targets. We determined the non-cytotoxic concentration of atractylodin (50 μM) using a cell proliferation assay in colonic epithelial cells. We found that pretreatment with atractylodin significantly inhibits tumor necrosis factor-α-induced phosphorylation of nuclear factor-κ-light-chain-enhancer of activated B in HCT116 cells. Through docking simulation analysis, luciferase assays, and in vitro binding assays, we found that atractylodin has an affinity for peroxisome proliferator-activated receptor alpha (PPARα). Daily administration of atractylodin (40 mg/kg) increased the survival rate of mice in a dextran sodium sulfate-induced colitis mouse model. Thus, atractylodin can be a good strategy for colitis therapy through inducing PPARα-dependent pathways.

N-Acetylserotonin (NAS) is an intermediate in the melatonin biosynthetic pathway. We investigated the anti-inflammatory activity of NAS by focusing on its chemical feature oxidizable to an electrophile. NAS was readily oxidized by reaction with HOCl, an oxidant produced in the inflammatory state. HOClreacted NAS (Oxi-NAS), but not NAS, activated the anti-inflammatory nuclear factor erythroid 2-related factor 2 (Nrf2)-heme oxygenase (HO)-1 pathway in cells. Chromatographic and mass analyses demonstrated that Oxi-NAS was the iminoquinone form of NAS and could react with N-acetylcysteine possessing a nucleophilic thiol to form a covalent adduct. Oxi-NAS bound to Kelch-like ECH-associated protein 1, resulting in Nrf2 dissociation. Moreover, rectally administered NAS increased the levels of nuclear Nrf2 and HO-1 proteins in the inflamed colon of rats. Simultaneously, NAS was converted to Oxi-NAS in the inflamed colon. Rectal NAS mitigated colonic damage and inflammation. The anticolitic effects were significantly compromised by the coadministration of an HO-1 inhibitor. K E Y W O R D Santi-inflammation, colitis, electrophile, melatonin, N-Acetylserotonin, nuclear factor erythroid 2-related factor 2 (Nrf2)-heme oxygenase (HO)-1 pathway, oxidation | INTRODUCTIONN-Acetylserotonin (NAS), also known as normelatonin, is produced from serotonin (ST) by arylalkylamine N-acetyltransferase (AANAT) and is converted to melatonin (MLT) by hydroxyindole-O-methyltransferase (HIOMT). 1-3 N-Acetylation of ST producing NAS occurs by alternative enzymes to AANAT in the cutaneous biosynthetic pathway for MLT. [4][5][6][7] MLT is predominantly synthesized in the mammalian pineal gland and the retina. However, its biosynthesis occurs in other tissues, such as the gastrointestinal tract (GIT), skin, and leukocytes. 1,8 Radioimmunoassay studies have revealed that 10-100 times higher concentrations of MLT are present in the GIT than in plasma. 9 About 15% of MLT is converted to NAS by O-demethylation in vivo, 10,11