Huaying Fan1, Zhenfang Gao1, Kai Ji1, Xin Li1, Jingbao Wu2, Yue Liu1, Xuekai Wang1, Haiyue Liang3, Yanan Liu1, Xiaoting Li4, Pan Liu1, Daquan Chen1, Feng Zhao5. 1. School of Pharmacy, Key Laboratory of Molecular Pharmacology and Drug Evaluation (Yantai University), Ministry of Education, Collaborative Innovation Center of Advanced Drug Delivery System and Biotech Drugs in Universities of Shandong, Yantai University, 30 Qingquan Road of Laishan District, Yantai 264003, Shandong, PR China. 2. Department of Endocrinology, Yantaishan Hospital, Yantai 264000, PR China. 3. Yantai Center for Food and Drug Control, Yantai 264000, PR China. 4. School of Pharmacy, Key Laboratory of Molecular Pharmacology and Drug Evaluation (Yantai University), Ministry of Education, Collaborative Innovation Center of Advanced Drug Delivery System and Biotech Drugs in Universities of Shandong, Yantai University, 30 Qingquan Road of Laishan District, Yantai 264003, Shandong, PR China; Yantai Hospital of Traditional Chinese Medicine, Yantai 264000, PR China. 5. School of Pharmacy, Key Laboratory of Molecular Pharmacology and Drug Evaluation (Yantai University), Ministry of Education, Collaborative Innovation Center of Advanced Drug Delivery System and Biotech Drugs in Universities of Shandong, Yantai University, 30 Qingquan Road of Laishan District, Yantai 264003, Shandong, PR China. Electronic address: ytuzhaofeng@163.com.
Abstract
BACKGROUND: Ulcerative colitis (UC) is a chronic inflammatory condition of the intestines and is difficult to cure once diagnosed. The efficacy of the current clinical treatment for UC is limited. Common anti-inflammatory drugs are prone to adverse effects, while novel biological agents are expensive, although tolerated by patients. Therefore, an urgency exists to find more safe and effective drugs to treat UC. Osthole is an active constituent isolated from the fruit of Cnidium monnieri (L.) Cuss. Osthole has anti-inflammatory activities and offers certain intestinal protection. These characteristics indicate that osthole has the potential to inhibit UC. PURPOSE: The study was conducted to investigate the anti-inflammatory potential of osthole in LPS-induced RAW 264.7 cells and dextran sulphate sodium (DSS)-induced ulcerative colitis in mice. METHODS: In in vitro experiments, mouse monocyte-macrophage RAW 264.7 cells were stimulated by 1 μg/ml LPS to produce inflammatory mediators. Griess reagent was used to determine Nitric Oxide (NO) production, and ELISA kits were used to determine the levels of PGE2, TNF-α, and IL-6. The anti-inflammatory mechanisms of osthole were detected using western blot. In in vivo experiments, UC was induced via the intragastric administration of 3.5% DSS to BALB/C mice for 7 days. During the experiment, clinical signs and body weight were monitored and recorded daily to calculate the DAI score. At the end of the experiment, the colon lengths were measured. The colonic histopathological lesions were evaluated. MPO activity and TNF-α levels were determined using the corresponding kits. The protein expression of TNF-α and NF-κB pathways were analysed using western blot. RESULTS: In an in vitro study, osthole inhibited the production of NO, PGE2, TNF-α, and IL-6 in LPS-induced RAW 264.7 cells. The results of western blot showed that osthole inhibited the expression of iNOS, COX-2, p38 MAPK and IκB α in RAW 264.7 cells. On this basis, in DSS-induced UC mice, it was found that osthole relieved the symptoms of UC by inhibiting weight loss, colon shortening and the DAI score, and simultaneously alleviating colon tissue lesions. It was also found that osthole reduced the levels of TNF-α in serum and colon tissues and effectively inhibited the activity of MPO. The western blot results showed that osthole reduced the expression of NF-κB p65 and p-IκB α and increased the content of IκB α in colon tissues. CONCLUSION: Osthole exerted anti-inflammatory effects by blocking the activation of the NF-κB and MAPK/p38 pathways. Additionally, osthole possesses therapeutic potential in the treatment of UC.
BACKGROUND:Ulcerative colitis (UC) is a chronic inflammatory condition of the intestines and is difficult to cure once diagnosed. The efficacy of the current clinical treatment for UC is limited. Common anti-inflammatory drugs are prone to adverse effects, while novel biological agents are expensive, although tolerated by patients. Therefore, an urgency exists to find more safe and effective drugs to treat UC. Osthole is an active constituent isolated from the fruit of Cnidium monnieri (L.) Cuss. Osthole has anti-inflammatory activities and offers certain intestinal protection. These characteristics indicate that osthole has the potential to inhibit UC. PURPOSE: The study was conducted to investigate the anti-inflammatory potential of osthole in LPS-induced RAW 264.7 cells and dextran sulphate sodium (DSS)-induced ulcerative colitis in mice. METHODS: In in vitro experiments, mouse monocyte-macrophage RAW 264.7 cells were stimulated by 1 μg/ml LPS to produce inflammatory mediators. Griess reagent was used to determine Nitric Oxide (NO) production, and ELISA kits were used to determine the levels of PGE2, TNF-α, and IL-6. The anti-inflammatory mechanisms of osthole were detected using western blot. In in vivo experiments, UC was induced via the intragastric administration of 3.5% DSS to BALB/C mice for 7 days. During the experiment, clinical signs and body weight were monitored and recorded daily to calculate the DAI score. At the end of the experiment, the colon lengths were measured. The colonic histopathological lesions were evaluated. MPO activity and TNF-α levels were determined using the corresponding kits. The protein expression of TNF-α and NF-κB pathways were analysed using western blot. RESULTS: In an in vitro study, osthole inhibited the production of NO, PGE2, TNF-α, and IL-6 in LPS-induced RAW 264.7 cells. The results of western blot showed that osthole inhibited the expression of iNOS, COX-2, p38 MAPK and IκB α in RAW 264.7 cells. On this basis, in DSS-induced UC mice, it was found that osthole relieved the symptoms of UC by inhibiting weight loss, colon shortening and the DAI score, and simultaneously alleviating colon tissue lesions. It was also found that osthole reduced the levels of TNF-α in serum and colon tissues and effectively inhibited the activity of MPO. The western blot results showed that osthole reduced the expression of NF-κB p65 and p-IκB α and increased the content of IκB α in colon tissues. CONCLUSION:Osthole exerted anti-inflammatory effects by blocking the activation of the NF-κB and MAPK/p38 pathways. Additionally, osthole possesses therapeutic potential in the treatment of UC.