Cyclophilin A differentially activates monocytes and endothelial cells: role of purity, activity, and endotoxin contamination in commercial preparations.
BACKGROUND
Cyclophilin A (CyPA) is a cytoplasmic protein secreted under inflammatory conditions. Extracellular CyPA is detected in atherosclerotic plaques and has been observed to activate endothelial cells as well as monocytes.


RESULTS
Commercially available recombinant CyPA-induced expression of tissue factor (TF) and vascular cell adhesion molecule-1 (VCAM-1) in human aortic endothelial cells (HAEC). However, CyPA from commercial sources contained lipopolysaccharide at concentrations up to 18.9 ng/ml; moreover, it exhibited low purity as determined by protein spectrum analysis and low activity as assessed by peptidyl prolyl cis-trans isomerase (PPIase) assay. An in-house preparation of pure, active, and uncontaminated CyPA failed to induce endothelial TF or VCAM-1 expression; moreover, it was not chemotactic for HAEC. In contrast, such CyPA exhibited potent chemotactic activity on monocytic THP-1 cells, with a maximal effect on migration occurring at a concentration of 5.5 x 10(-9)mol/l. Pretreatment of CyPA with cyclosporine A prevented its effect on THP-1 cell migration; similarly, PPIase-deficient mutant CyPA protein did not induce migration of these cells. In-house prepared CyPA induced the release of Il-6, but not TNF-alpha, from THP-1 cells.


CONCLUSIONS
Commercially available CyPA exhibits low purity and activity and may be contaminated by endotoxin. Pure, active, and uncontaminated CyPA does not induce endothelial TF or VCAM-1 expression; instead, it acts as a potent monocyte chemoattractant and induces monocyte Il-6 release, implying a role for extracellular CyPA in the pathogenesis of atherosclerosis via activation of monocytes rather than endothelial cells.
Payeli
Sravan K
SK
Cardiovascular Research, Institute of Physiology, University of Zurich, Switzerland.
Schiene-Fischer
Cordelia
C
Steffel
Jan
J
Camici
Giovanni G
GG
Rozenberg
Izabela
I
Lüscher
Thomas F
TF
Tanner
Felix C
FC
D001011:Aorta / Q000166:cytology; D002478:Cells, Cultured; D002630:Chemotactic Factors / Q000502:physiology; D021984:Cyclophilin A / Q000096:biosynthesis / Q000737:chemistry* / Q000502:physiology*; D004340:Drug Contamination; D042783:Endothelial Cells / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D008070:Lipopolysaccharides / Q000302:isolation & purification / Q000494:pharmacology; D009000:Monocytes / Q000187:drug effects / Q000502:physiology*; D013925:Thromboplastin / Q000378:metabolism; D019010:Vascular Cell Adhesion Molecule-1 / Q000378:metabolism
D002630:Chemotactic Factors; D008070:Lipopolysaccharides; D019010:Vascular Cell Adhesion Molecule-1; D013925:Thromboplastin; D021984:Cyclophilin A