Induction of inflammatory cytokine release from human umbilical vein endothelial cells by agonists of proteinase-activated receptor-2.
1. Human endothelial cells express proteinase-activated receptor-2 (PAR-2), inflammatory cytokines and trypsin (EC 3.4.21.4). However, little is known about the mechanism through which trypsin induces cytokine release from endothelial cells. 2. In the present study, we investigated the effect of trypsin on cytokine release from primary cultures of human umbilical vein endothelial cells (HUVEC) using an antibody based protein microarray and ELISA. 3. The results showed that 1 microg/mL trypsin induced release of 32 different inflammatory factors, whereas 100 micromol/L Ser-Leu-Ile-Gly-Lys-Val-NH2 (SLIGKV-NH2) only stimulated secretion of 16 inflammatory factors from HUVEC, as assessed by an antibody based protein microarray. Because the release of interleukin (IL)-1a, IL-8, IL-10 and IL-12 was markedly increased following PAR-2 activation, their release was investigated further using ELISA. Increases in release of up to approximately 4.8-, 4.3-, 4.1- and 1.8-fold were observed for IL-1a, IL-10, IL-12 and IL-8, respectively, when HUVEC were challenged with trypsin for 16 h. Agonist peptides of PAR-2, namely SLIGKV-NH2 and trans-cinnamoyl-Leu-Ile-Gly-Arg-Leu-Orn-NH2 (tc-LIGRLO-NH2), also provoked significant release of IL-8. Trypsin-induced cytokine release was inhibited by its inhibitors soybean trypsin inhibitor, alpha1-antitrypsin and the inhibitor peptide of PAR-2 Phe-Ser-Leu-Leu-Arg-Tyr-NH2 (FSLLRY-NH2). 4. These data indicate the action of trypsin on HUVEC is most likely through activation of PAR-2, suggesting that PAR-2-related mechanisms are involved in the inflammatory process in humans.
Niu
Qing-Xia
QX
Allergy and Inflammation Research Institute, Key Immunopharmacology Laboratory of Guangdong Province, Shantou University Medical College, Shantou, Guangdong, China.
Chen
Han-Qiu
HQ
Chen
Zhuo-Yi
ZY
Fu
Yi-Ling
YL
Lin
Jie-Lian
JL
He
Shao-Heng
SH
D015703:Antigens, CD / Q000032:analysis; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D000071063:Endoglin; D042783:Endothelial Cells / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D018836:Inflammation Mediators / Q000378:metabolism*; D016753:Interleukin-10 / Q000378:metabolism; D018664:Interleukin-12 / Q000378:metabolism; D053582:Interleukin-1alpha / Q000378:metabolism; D016209:Interleukin-8 / Q000378:metabolism; D009842:Oligopeptides / Q000494:pharmacology*; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000032:analysis; D040081:Protein Array Analysis; D044464:Receptor, PAR-2 / Q000819:agonists* / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000032:analysis; D015398:Signal Transduction / Q000187:drug effects*; D014357:Trypsin / Q000378:metabolism* / Q000494:pharmacology; D014471:Umbilical Veins / Q000166:cytology / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D000515:alpha 1-Antitrypsin / Q000378:metabolism
D015703:Antigens, CD; D016207:Cytokines; C064206:ENG protein, human; D000071063:Endoglin; C508716:H-Phe-Ser-Leu-Leu-Arg-Tyr-NH2; C508609:IL10 protein, human; D018836:Inflammation Mediators; D053582:Interleukin-1alpha; D016209:Interleukin-8; D009842:Oligopeptides; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D044464:Receptor, PAR-2; D011956:Receptors, Cell Surface; D000515:alpha 1-Antitrypsin; C113425:cinnamoyl-LIGRLO-amide; C515807:seryl-leucyl-isoleucyl-glycyl-lysyl-valinamide; D016753:Interleukin-10; D018664:Interleukin-12; D014357:Trypsin