Yu Zhang1, Norio Takagi2, Bo Yuan2, Yanyan Zhou1, Nan Si1, Hongjie Wang1, Jian Yang1, Xiaolu Wei1, Haiyu Zhao3, Baolin Bian4. 1. Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing, 100700, China. 2. Department of Applied Biochemistry, Tokyo University of Pharmacy & Life Sciences, 1432-1 Horinouchi, Hachioji, Tokyo, 192-0392, Japan. 3. Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing, 100700, China. Electronic address: hyzhao@icmm.ac.cn. 4. Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing, 100700, China. Electronic address: blbian@icmm.ac.cn.
Abstract
ETHNOPHARMACOLOGICAL RELEVANCE: Toad skin came from Bufo bufo gargarizans Cantor and Bufo melanostictus Schneider. As the traditional Chinese medicine, it had the effect of clearing away heat and detoxification. In traditional applications, toad skin was often used for the treatment of cancer and inflammation. Total indolealkylamines (IAAs) from this medicine were proved the main compounds exert anti-inflammatory activity in our previous research. AIM OF THE STUDY: In the present study, we aimed to investigate the potential mechanism of anti-inflammatory activity of IAAs on LPS induced zebrafish. MATERIALS AND METHODS: LPS induced zebrafish was applicated as an in vivo inflammation model to clarify the structure-activity relationship of 4 major IAAs (N-methyl serotonin, bufotenine, dehydrobufotenine and bufothionine) from toad skin. Quantitative RT-PCR was applied to detect key cytokines and members of the MyD88-dependent signaling pathway. In addition, the targeted lipidomics was conducted to find out the potential biomarkers in the inflammatory zebrafish. Network pharmacology was used to unveil the main enzymes closely related to the target lipids. RESULTS: Our results showed that the anti-inflammatory activity of free IAAs (N-methyl serotonin, bufotenine and dehydrobufotenine) was more potent than that of combined IAAs (bufothionine). RT-PCR demonstrated that 4 IAAs exerted antiendotoxin inflammatory effect via suppressing the TLR4/MyD88/NF-κB and TLR4/MyD88/MAPKs signaling pathway. A total of 33 possible inflammatory biomarkers, including 14 SM, 6 Cer, 11 PC and 2 GlcCer, triggered by LPS were screened out. The levels of most of candidates could be regulated toward a normal level by IAAs, especially in N-methyl serotonin and dehydrobufotenine groups. Enzymes especially LBP, PLA2, CERK, SMPD and SGMS were found closely associated with the regulation of most lipid markers. CONCLUSIONS: Overall, the mechanism underlying the anti-inflammatory activity of IAAs probably attributed to their capability to suppress NF-κB and MAPKs inflammatory pathway. Meanwhile, IAAs could also interfere the metabolism of SM, Cer and PC probably by regulating LBP, PLA2, CERK, SMPD and SGMS.
ETHNOPHARMACOLOGICAL RELEVANCE: Toad skin came from Bufo bufo gargarizans Cantor and Bufo melanostictus Schneider. As the traditional Chinese medicine, it had the effect of clearing away heat and detoxification. In traditional applications, toad skin was often used for the treatment of cancer and inflammation. Total indolealkylamines (IAAs) from this medicine were proved the main compounds exert anti-inflammatory activity in our previous research. AIM OF THE STUDY: In the present study, we aimed to investigate the potential mechanism of anti-inflammatory activity of IAAs on LPS induced zebrafish. MATERIALS AND METHODS: LPS induced zebrafish was applicated as an in vivo inflammation model to clarify the structure-activity relationship of 4 major IAAs (N-methyl serotonin, bufotenine, dehydrobufotenine and bufothionine) from toad skin. Quantitative RT-PCR was applied to detect key cytokines and members of the MyD88-dependent signaling pathway. In addition, the targeted lipidomics was conducted to find out the potential biomarkers in the inflammatory zebrafish. Network pharmacology was used to unveil the main enzymes closely related to the target lipids. RESULTS: Our results showed that the anti-inflammatory activity of free IAAs (N-methyl serotonin, bufotenine and dehydrobufotenine) was more potent than that of combined IAAs (bufothionine). RT-PCR demonstrated that 4 IAAs exerted antiendotoxin inflammatory effect via suppressing the TLR4/MyD88/NF-κB and TLR4/MyD88/MAPKs signaling pathway. A total of 33 possible inflammatory biomarkers, including 14 SM, 6 Cer, 11 PC and 2 GlcCer, triggered by LPS were screened out. The levels of most of candidates could be regulated toward a normal level by IAAs, especially in N-methyl serotonin and dehydrobufotenine groups. Enzymes especially LBP, PLA2, CERK, SMPD and SGMS were found closely associated with the regulation of most lipid markers. CONCLUSIONS: Overall, the mechanism underlying the anti-inflammatory activity of IAAs probably attributed to their capability to suppress NF-κB and MAPKs inflammatory pathway. Meanwhile, IAAs could also interfere the metabolism of SM, Cer and PC probably by regulating LBP, PLA2, CERK, SMPD and SGMS.