Identification and characterization of a novel neuropeptide(neuropeptide Y-HS) from leech salivary gland of Haemadipsa sylvestris

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.