Regulatory Role Of Hemokinin-1(4-11) On Proliferation And Differentiation Of Human Leukemia Cells In Vitro And The Establishment And Application Of Mammalian Cell Expression For NK₁ Receptor

Human hemokinin-1 and its truncated form human hemokinin-1(4-11) in which three amino acid residues at the N-terminal are mammalian tachykinin peptides encoded by the recently identified TAC4 gene in human.The tachykinins have shown immuno-regulatory activities in humans,and human hemokinin-1 have an inhibitory effect on the proliferation of a human promyelocyte leukemia cell line, HL-60. So we investigated the effects of human hemokinin-1(4-11) on the proliferation and differentiation of this cell line. It is noteworthy that human hemokinin-1(4-11) (1-300μM) displayed inhibitory effects on the proliferation of HL-60 cells in a dose-and time-dependent manner. The effect of suppressing proliferation induced by this peptide was accompanied by an accumulation of cell cycle in the S phase. Moreover, this peptide induced differentiation of HL-60 cells by significantly increasing the NBT-reduction activity. However the effect induced by human hemokinin-1 (4-11)was not antagonized by the NK1 receptor antagonist SR140333 or the NK2 receptor antagonist SR48968. All the results indicated that human hemokinin-1(4-11) was able to significantly inhibit proliferation and induce differentiation and S phase arrest of a human promyelocyte leukemia cell line HL-60, which may not be mediated through the activation of classical tachykinin NK1 receptors and tachykinin NK2 receptors. Our observations also implied that human hemokinin-1(4-11) could act as an immunomodulatory factor in cancer chemotherapy.NK1 receptor mediates the effects of substance P (SP) that plays a key role in several physiological processes including pain transmission and inflammation. NK1 receptor is a member of G protein-coupled receptors (GPCRs). Recent studies indicated that SP binding with NK1 receptor could activate multiple cellular responses, including cAMP formation and the cascade reactions of mitogen-activated protein kinase (MAPK). Human hemokinin-1 and its truncated form human hemokinin-1(4-11) are also the members of mammalian tachykinin peptides.However, there were no research about the effects of human hemokinin-1 and human hemokinin-1 (4-11) on cell singals. So we transfected the plasmid pcDNA3.1-3×HA NK1 receptor into CHO cell, and selected the clone stable expression of NK1 receptor.Then we used the cell model to detect the effect of human hemokinin-1 and human hemokinin-1 (4-11) on the cAMP level and ERK1/2 phosphorylation. The results showed that both peptides could increase the intracellular cAMP, and induce the phosphorylation of ERK1/2.This section just investigated the effects of tachykinin peptide receptors binding with corresponding ligands on cell singals.It points out a new direction on the research about the interaction between newly discovered tachykinin peptide ligands and their receptors.