Extracellular PBEF/NAMPT/visfatin activates pro-inflammatory signalling in human vascular smooth muscle cells through nicotinamide phosphoribosyltransferase activity.
OBJECTIVE
Extracellular pre-B cell colony-enhancing factor/nicotinamide phosphoribosyltransferase/visfatin (ePBEF/NAMPT/visfatin) is an adipocytokine, whose circulating levels are enhanced in metabolic disorders, such as diabetes mellitus and obesity. Here, we explored the ability of ePBEF/NAMPT/visfatin to promote vascular inflammation, as a condition closely related to atherothrombotic diseases. We specifically studied the ability of PBEF/NAMPT/visfatin to directly activate pathways leading to inducible nitric oxide synthase (iNOS) induction in cultured human aortic smooth muscle cells, as well as the mechanisms involved.


METHODS
iNOS levels and extracellular signal-regulated kinase (ERK) 1/2 activity were determined by western blotting. Nuclear factor (NF)-kappaB activity was assessed by electrophoretic mobility shift assay.


RESULTS
ePBEF/NAMPT/visfatin (10-250 ng/ml) induced iNOS in a concentration-dependent manner. At a submaximal concentration (100 ng/ml), ePBEF/NAMPT/visfatin time-dependently enhanced iNOS levels up to 18 h after stimulation. Over this time period, ePBEF/NAMPT/visfatin elicited a sustained activation of NF-kappaB and triggered a biphasic ERK 1/2 activation. By using the respective ERK 1/2 and NF-kappaB inhibitors, PD98059 and pyrrolidine dithiocarbamate, we established that iNOS induction by ePBEF/NAMPT/visfatin required the consecutive upstream activation of ERK 1/2 and NF-kappaB. The pro-inflammatory action of ePBEF/NAMPT/visfatin was not prevented by insulin receptor blockade. However, exogenous nicotinamide mononucleotide, the product of NAMPT activity, mimicked NF-kappaB activation and iNOS induction by ePBEF/NAMPT/visfatin, while the NAMPT inhibitor APO866 prevented the effects of ePBEF/NAMPT/visfatin on iNOS and NF-kappaB.


CONCLUSIONS
Through its intrinsic NAMPT activity, ePBEF/NAMPT/visfatin appears to be a direct contributor to vascular inflammation, a key feature of atherothrombotic diseases linked to metabolic disorders.
Romacho
T
T
Departamento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain.
Azcutia
V
V
Departamento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain.
Vázquez-Bella
M
M
Departamento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain.
Matesanz
N
N
Departamento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain.
Cercas
E
E
Departamento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain.
Nevado
J
J
Instituto de Genética Médica y Molecular (INGEMM), Hospital Universitario La Paz, Madrid, Spain.
Carraro
R
R
Servicio de Endocrinología, Hospital Universitario de La Princesa y Universidad Autónoma de Madrid, Madrid, Spain.
Rodríguez-Mañas
L
L
Unidad de Investigación y Servicio de Geriatría, Hospital Universitario de Getafe, Getafe, Spain.
Sánchez-Ferrer
C F
CF
Departamento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain.
Peiró
C
C
Departamento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain. concha.peiro@uam.es.
D001011:Aorta / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D018929:Cell Culture Techniques; D016207:Cytokines / Q000494:pharmacology* / Q000502:physiology*; D003920:Diabetes Mellitus / Q000097:blood / Q000503:physiopathology; D005419:Flavonoids / Q000494:pharmacology; D006801:Humans; D050417:Insulin-Secreting Cells / Q000187:drug effects / Q000502:physiology; D007700:Kinetics; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000502:physiology*; D016328:NF-kappa B / Q000187:drug effects / Q000378:metabolism; D054409:Nicotinamide Phosphoribosyltransferase / Q000378:metabolism* / Q000494:pharmacology / Q000502:physiology; D052247:Nitric Oxide Synthase Type II / Q000187:drug effects / Q000378:metabolism; D009765:Obesity / Q000097:blood / Q000503:physiopathology; D015398:Signal Transduction
D016207:Cytokines; D005419:Flavonoids; D016328:NF-kappa B; D052247:Nitric Oxide Synthase Type II; D054409:Nicotinamide Phosphoribosyltransferase; C085455:nicotinamide phosphoribosyltransferase, human; D048052:Mitogen-Activated Protein Kinase 3; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one