Objective-Obesity and hypertension are comorbid in epidemic proportion, yet their biological connection is largely a mystery. The peptide chemerin is a candidate for connecting fat deposits around the blood vessel (perivascular adipose tissue) to arterial contraction. We presently tested the hypothesis that chemerin is expressed in perivascular adipose tissue and is vasoactive, supporting the existence of a chemerin axis in the vasculature. Approach and Results-Real-time polymerase chain reaction, immunohistochemistry, and Western analyses supported the synthesis and expression of chemerin in perivascular adipose tissue, whereas the primary chemerin receptor ChemR23 was expressed both in the tunica media and endothelial layer. The ChemR23 agonist chemerin-9 caused receptor, concentration-dependent contraction in the isolated rat thoracic aorta, superior mesenteric artery, and mesenteric resistance artery, and contraction was significantly amplified (more than 100%) when nitric oxide synthase was inhibited and the endothelial cell mechanically removed or tone was placed on the arteries. The novel ChemR23 antagonist CCX832 inhibited phenylephrine-induced and prostaglandin F2α-induced contraction (+perivascular adipose tissue), suggesting that endogenous chemerin contributes to contraction. Arteries from animals with dysfunctional endothelium (obese or hypertensive) demonstrated a pronounced contraction to chemerin-9. Finally, mesenteric arteries from obese humans demonstrate amplified contraction to chemerin-9. Conclusions-These Watts et al Chemerin as a Vasoconstrictor 1321also play a role in obesity. Additionally, chemerin regulates adipocyte differentiation [18][19][20] and production of several proinflammatory cytokines. We hypothesized that a chemerin axis exists in blood vessels. We propose that chemerin and the primary receptor for chemerin, ChemR23, are present and mediate contraction in the vasculature. Materials and MethodsMaterials and Methods are available in the online-only Supplement. Results Arterial Chemerin AxisIsolated rat arteries express chemerin protein in the PVAT ( Figure 1A). Real-time polymerase chain reaction supports the expression of chemerin (RARRES2) mRNA in the rat thoracic aortic PVAT (whole PVAT; threshold cycle [C T ] =22.78±0.35; β2-microglobulin as control = 19.32±0.27; n=6). Chemerin signal does not wholly derive from resident mast cells because there was negligible CD68 staining in PVAT ( Figure 1B, + control below), and staining for chemerin was, in many places, not punctate. Positive staining was observed within the cytoplasm of the fat cell, outside the rounded lipid droplet. The predominant receptor for chemerin, ChemR23, is expressed in the tunica media and endothelial cell layer ( Figure 1C) and is observed as 3 dominant bands in homogenates (−PVAT) of the thoracic aorta and superior mesenteric artery cleaned of PVAT ( Figure 1D and 1E). Two bands (at arrows) are consistent with that observed in a JAR (choriocarcinoma) positive control and were 42 kDa (expected size for...

BackgroundSerotonin (5-hydroxytryptamine, 5-HT) was named for its source (sero-) and ability to modify smooth muscle tone (tonin). The biological effects of 5-HT are believed to be carried out by stimulation of serotonin receptors at the plasma membrane. Serotonin has recently been shown to be synthesized in vascular smooth muscle and taken up from external sources, placing 5-HT inside the cell. The enzyme transglutaminase uses primary amines such as 5-HT to covalently modify proteins on glutamine residues. We tested the hypothesis that 5-HT is a substrate for transglutaminase in arterial vascular smooth muscle, with protein serotonylation having physiological function.Methodology/Principal FindingsThe model was the rat aorta and cultured aortic smooth muscle cells. Western analysis demonstrated that transglutaminase II was present in vascular tissue, and transglutaminase activity was observed as a cystamine-inhibitable incorporation of the free amine pentylamine-biotin into arterial proteins. Serotonin-biotin was incorporated into α -actin, β-actin, γ-actin, myosin heavy chain and filamin A as shown through tandem mass spectrometry. Using antibodies directed against biotin or 5-HT, immunoprecipitation and immunocytochemistry confirmed serotonylation of smooth muscle α–actin. Importantly, the α-actin-dependent process of arterial isometric contraction to 5-HT was reduced by cystamine.Conclusions5-HT covalently modifies proteins integral to contractility and the cytoskeleton. These findings suggest new mechanisms of action for 5-HT in vascular smooth muscle and consideration for intracellular effects of primary amines.