The present study was designed to identify immunomodulatory components from the leech salivary gland of Haemadipsa sylvestris. The Sephadex G-50, ResourceTM S column chromatography and reverse-phase high performance l...The present study was designed to identify immunomodulatory components from the leech salivary gland of Haemadipsa sylvestris. The Sephadex G-50, ResourceTM S column chromatography and reverse-phase high performance liquid chromatography(RP-HPLC) were used to isolate and purify the salivary gland extracts(SGE). Structural analysis of isolated compounds was based on Edman degradation and matrix assisted laser desorption ionization time-of-flight mass spectrometer(MALDI-TOF-MS). The c DNA encoding the precursor of the compound was cloned from the c DNA library of the salivary gland of H. sylvestris. The levels of inflammatory mediators, including tumor necrosis factor-α(TNF-α), interferon ?(IFN-?), interleukin-6(IL-6), and monocyte chemotactic protein-1(MCP-1) were assayed using an enzyme-linked immunosorbent assay(ELISA). The effects on cell proliferation and cell viability were observed using MTT assay. A novel neuropeptide Y(Neuropeptide Y-HS) from the leech salivary gland of H. sylvestris was purified and characterized. It was composed of 36 amino acid residues and the amino acid sequence was determined to be FLEPPERPAVFTSVEQMKSYIKALNDYYLLLGRPRF-NH2, containing an amidated C-terminus. It showed significant inhibitory effects on the production of inflammatory cytokines including TNF-?, IFN-?, IL-6, and MCP-1. Neuropeptide Y was identified from leeches for the first time. The presence of neuropeptide Y-HS in leech salivary gland may help get blood meal from hosts and inhibit inflammation.展开更多
[Objectives]Peptides was extracted from Haemadipsa hainana and its activity was studied.[Methods]Electric stimulation,water extraction and ultrasonic extraction were used to extract the peptides from H.hainana.Then th...[Objectives]Peptides was extracted from Haemadipsa hainana and its activity was studied.[Methods]Electric stimulation,water extraction and ultrasonic extraction were used to extract the peptides from H.hainana.Then the protein content and molecular weight distribution of H.hainana peptides were detected by the BCA method and SDS-PAGE method,respectively.The antithrombin activity and analgesic activity of the three peptide extracts of H.hainana were detected by Markwardt thrombin titration method and mouse hot plate experiment,respectively.[Results]There extraction methods of electric stimulation,water extraction and ultrasonic extraction were used to extract the peptide extract of H.hainana,and the yields were as follows:water extraction>electrical stimulation>ultrasonic extraction.The three peptide extracts from H.hainana had antithrombin activity,and the antithrombin activity was as follows:water extraction>ultrasonic extraction>electrical stimulation.Through the hot plate experiment in mice,it was verified that the three peptide extracts of H.hainana had analgesic activity,and the analgesic activity was water extraction>electric stimulation>ultrasonic extraction.The analgesic activity of high-dose(100 mg/kg)group of H.hainana obtained by water extraction was slightly weaker than that of tramadol.[Conclusions]This study confirmed that the peptide extract of H.hainana had certain antithrombin and analgesic activity,laying a foundation for the subsequent development and application of H.hainana.展开更多
基金supported by Ministry of Science and Technology of China(Nos.2013CB911300,2010CB529800)the National Science Foundation of China(Nos.U1132601,31200590)the National Science Foundation of Yunnan Province(No.P0120150010)
文摘The present study was designed to identify immunomodulatory components from the leech salivary gland of Haemadipsa sylvestris. The Sephadex G-50, ResourceTM S column chromatography and reverse-phase high performance liquid chromatography(RP-HPLC) were used to isolate and purify the salivary gland extracts(SGE). Structural analysis of isolated compounds was based on Edman degradation and matrix assisted laser desorption ionization time-of-flight mass spectrometer(MALDI-TOF-MS). The c DNA encoding the precursor of the compound was cloned from the c DNA library of the salivary gland of H. sylvestris. The levels of inflammatory mediators, including tumor necrosis factor-α(TNF-α), interferon ?(IFN-?), interleukin-6(IL-6), and monocyte chemotactic protein-1(MCP-1) were assayed using an enzyme-linked immunosorbent assay(ELISA). The effects on cell proliferation and cell viability were observed using MTT assay. A novel neuropeptide Y(Neuropeptide Y-HS) from the leech salivary gland of H. sylvestris was purified and characterized. It was composed of 36 amino acid residues and the amino acid sequence was determined to be FLEPPERPAVFTSVEQMKSYIKALNDYYLLLGRPRF-NH2, containing an amidated C-terminus. It showed significant inhibitory effects on the production of inflammatory cytokines including TNF-?, IFN-?, IL-6, and MCP-1. Neuropeptide Y was identified from leeches for the first time. The presence of neuropeptide Y-HS in leech salivary gland may help get blood meal from hosts and inhibit inflammation.
基金Supported by"2020 Hainan Provincial Higher Education Teaching Reform Research and Scientific Research"Project of The Education Department of Hainan Province(Hnky2020-76)Research Fund of Hainan Health Management College(2020ZR01)。
文摘[Objectives]Peptides was extracted from Haemadipsa hainana and its activity was studied.[Methods]Electric stimulation,water extraction and ultrasonic extraction were used to extract the peptides from H.hainana.Then the protein content and molecular weight distribution of H.hainana peptides were detected by the BCA method and SDS-PAGE method,respectively.The antithrombin activity and analgesic activity of the three peptide extracts of H.hainana were detected by Markwardt thrombin titration method and mouse hot plate experiment,respectively.[Results]There extraction methods of electric stimulation,water extraction and ultrasonic extraction were used to extract the peptide extract of H.hainana,and the yields were as follows:water extraction>electrical stimulation>ultrasonic extraction.The three peptide extracts from H.hainana had antithrombin activity,and the antithrombin activity was as follows:water extraction>ultrasonic extraction>electrical stimulation.Through the hot plate experiment in mice,it was verified that the three peptide extracts of H.hainana had analgesic activity,and the analgesic activity was water extraction>electric stimulation>ultrasonic extraction.The analgesic activity of high-dose(100 mg/kg)group of H.hainana obtained by water extraction was slightly weaker than that of tramadol.[Conclusions]This study confirmed that the peptide extract of H.hainana had certain antithrombin and analgesic activity,laying a foundation for the subsequent development and application of H.hainana.
