Protective role of fermented mulberry leave extract in LPS­induced inflammation and autophagy of RAW264.7 macrophage cells.

Mulberry leaves have antioxidant activity and anti­inflammatory effects in several types of cells. However, the efficacy of mulberry leaves fermented with Cordyceps militaris remains unknown. Therefore, the present study aimed to investigate whether the ethanol extracts of mulberry leaves fermented with C. militaris (EMfC) can prevent lipopolysaccharide (LPS)­induced inflammation and autophagy in macrophages. To achieve this, RAW264.7 cells pretreated with three different dose of EMfCs were subsequently stimulated with LPS, and examined for alterations in the regulatory factors of inflammatory responses and key parameters of the autophagy signaling pathway. EMfC treatment inhibited the generation of reactive oxidative species; however, significant activity was observed for 2,2­diphenyl­1­picrylhydrazyl (DPPH) radical scavenging (IC50=579.6703 mg/ml). Most regulatory factors in inflammatory responses were significantly inhibited following treatment with EMfC, without any significant cellular toxicity. EMfC­treated groups exhibited marked suppression of nitrogen oxide (NO) levels, mRNA expression levels of iNOS/COX­2, levels of all inflammatory cytokines (TNF­α, IL­1ß and IL­6) and phosphorylation of MAPK members, as well as recovery of cell cycle progression. Furthermore, similar effects were observed in the LPS­induced autophagy signaling pathway of RAW264.7 cells. The expression levels of microtubule­associated protein 1A/1B­light chain 3 (LC3) and Beclin exhibited a dose­dependent decrease in the EMfC+LPS­treated groups compared with in the Vehicle+LPS­treated group, whereas the phosphorylation of PI3K and mTOR were enhanced in a dose­dependent manner in the same groups. Overall, the results of the present study provide evidence that exposure to EMfC protects against LPS­induced inflammation and autophagy in RAW264.7 cells. These results indicated that EMfC is a potential candidate for treatment of inflammatory diseases.

Synergic Laxative Effects of an Herbal Mixture of Liriope platyphylla, Glycyrrhiza uralensis, and Cinnamomum cassia in Loperamide-Induced Constipation of Sprague Dawley Rats.

Constipation is an acute or chronic illness attributed to various causes, ranging from lifestyle habits to side effects of a disease. To improve the laxative effects of some traditional medicines, herbal mixtures of Liriope platyphylla, Glycyrrhiza uralensis, and Cinnamomum cassia (LGC) were evaluated for their mechanism of action and therapeutic effects in loperamide (Lop)-induced constipated Sprague Dawley rats by examining alterations in excretion parameters, histological structure, mucin secretion, and related protein levels. Food intake and water consumption were constant for all animals. We observed that the Lop+LGC-treated group had significantly greater excretion of stool and urine than was observed in the Lop+Vehicle-treated group. Administration of LGC in the constipation model restored the intestinal transit ratio to normal levels, and increased the number of goblet cells, mucosal layer, and muscle thickness. Mucin secretion was greater in the Lop+LGC-treated group than in the Lop+Vehicle-treated group, and the expression of MUC2 and AQP8 genes were also increased. In addition, reverse transcription polymerase chain reaction and Western blot revealed an increase in the muscarinic acetylcholine receptors (mAChRs) in the Lop+LGC-treated group compared to the Lop+Vehicle-treated group. Furthermore, compared with the Lop+Vehicle-treated group, treatment with LGC reduced the phosphorylation of PKC and PI3K, and expression of Gα protein, but increased levels of IP3. Our results suggest that the traditional herbal mixture of LGC induces a potent laxative effect in Lop-induced constipation through mucosal tissue changes and mucin production. We also demonstrated that the laxative effect of LGC is closely related to the expression of mAChR and its downstream signals, suggesting the possibility of developing a constipation-laxative agent using LGC.