Wide proinflammatory effect of electronegative low-density lipoprotein on human endothelial cells assayed by a protein array.
Electronegative low-density lipoprotein (LDL(-)) is a modified subfraction of LDL present in plasma able to induce the release of interleukin 8 (IL-8) and monocyte chemotactic protein 1 (MCP-1) by human umbilical vein endothelial cells (HUVEC). To ascertain whether further inflammation mediator release could be induced by LDL(-), a protein array system was used to measure 42 cytokines and related compounds. Native LDL and LDL(-) isolated from normolipemic subjects were incubated for 24 h with HUVEC and culture supernatants were used to measure inflammation mediator release. The protein array revealed that IL-6, granulocyte/monocyte colony-stimulating factor (GM-CSF) and growth-related oncogene (GRO) release were increased by cultured HUVEC in response to LDL(-). LDL(-) enhanced production of IL-6 (4-fold vs. LDL(+)), GM-CSF (4-fold), GRObeta (2-fold) and GROgamma (7-fold) was confirmed by ELISA. Time-course experiments revealed that IL-6 was released earlier than the other inflammation mediators, suggesting a first-wave cytokine action. However, the addition of IL-6 alone did not stimulate the production of IL-8, MCP-1 or GM-CSF. Moreover, IL-8, MCP-1 or GM-CSF alone did not promote the release of the other inflammatory molecules. Modification of LDL(+) by phospholipase A(2)-mediated lipolysis or by loading with non-esterified fatty acids (NEFA) reproduced the action of LDL(-), thereby suggesting the involvement of NEFA and/or lysophosphatidylcholine in the release of these molecules. Our results indicate that LDL(-) promotes a proinflammatory phenotype in endothelial cells through the production of cytokines, chemokines and growth factors.
Benítez
Sònia
S
Department of Biochemistry and Institut de Recerca, Hospital de la Santa Creu i Sant Pau, C/Antoni Maria Claret 167, 08025 Barcelona, Spain.
Camacho
Mercedes
M
Bancells
Cristina
C
Vila
Luis
L
Sánchez-Quesada
José Luis
JL
Ordóñez-Llanos
Jordi
J
D002478:Cells, Cultured; D019743:Chemokines, CXC / Q000378:metabolism; D016207:Cytokines / Q000378:metabolism*; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism*; D005230:Fatty Acids, Nonesterified / Q000494:pharmacology*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000378:metabolism; D006801:Humans; D007249:Inflammation / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism; D008077:Lipoproteins, LDL / Q000494:pharmacology / Q000502:physiology*; D010741:Phospholipases A / Q000378:metabolism; D040081:Protein Array Analysis; D014471:Umbilical Veins / Q000166:cytology
D019743:Chemokines, CXC; D016207:Cytokines; D005230:Fatty Acids, Nonesterified; D015850:Interleukin-6; D008077:Lipoproteins, LDL; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D010741:Phospholipases A