Chemerin connects fat to arterial contraction

Abstract

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 data support a new role for chemerin as an endogenous vasoconstrictor that operates through a receptor typically attributed to function only in immune cells.

Keywords: ChemR23; PVAT; adipose tissue; chemerin; vasoconstriction.

Figure 1. Chemerin Axis in Rat Arteries

Immunohistochemical staining of isolated rat thoracic aorta for chemerin (a), CD68 (b) and ChemR23 (c). Representative of six different rats. + = bone marrow positive control (left image with antibody, right image without). d, Western analyses of homogenates of seven separate samples isolated from the thoracic aorta (left) and superior mesenteric artery (right) −PVAT from normal Sprague Dawley rats. + = human choriocarcinoma (JAR) whole cell lysate. Quantification of ChemR23 expression normalized to alpha actin is in e, presented as means±SEM for number of animals in parentheses. Bars = 100 microns. * p<0.05, unpaired Students t test aorta vs mesentery. L = arterial lumen.

Figure 2. Chemerin-9-induced Arterial Contraction

a, Concentration-dependent response to chemerin from baseline and above PE-induced contraction in the endothelium intact (+E) isolated thoracic aorta. Dotted line is point from which contraction was measured. b, Quantification of data in tissues in which PVAT was left intact (left) or removed (right). * significant difference of maximum response in baseline vs PE contracted tissue (p <0.05). c, Concentration-dependent response to chemerin-9 in a PE-induced contraction of the endothelium-intact isolated superior mesenteric artery, quantified on the far right. * significant difference in maximum response in baseline vs PE contracted tissue (p<0.05). d, Chemerin-9-induced contraction in thoracic aorta (no PVAT, endothelium intact) contracted with different agonists (left) and effect of endothelium removal and LNNA on baseline induced contraction (right). * significantly different from baseline values, † different from LNNA values. Points represent means±SEM for number of animals in parentheses.

Figure 3. Amplification and ChemR23-dependence of Chemerin-9 Contraction

a, left: Sample tracing of the ability of chemerin-9 to cause concentration-, ChemR23-dependent contraction in the aorta with endothelium removed (−E). Right, Quantification of the ability of CCX832 to inhibit chemerin-9 induced contraction. b, left: representative tracing of CCX832 reversal of chemerin-9-induced contraction. Right demonstrates quantification of the effects of equivalent and increasing concentrations of CCX826 (inactive analog, DMSO) and CCX832 (ChemR23 antagonist, DMSO) in reversing chemerin-9-induced contraction. Bars/points represent means±SEM for number of animals in parentheses. * significantly different from CCX826 (p<0.05). c, Ability of ChemR23 receptor antagonist CCX832 to inhibit PE (left), PGF2α (middle) but not KCl (right) -induced contraction in tissues + PVAT, * = significantly different from appropriate (+ or −PVAT) vehicle-incubated responses.

Figure 4. The Chemerin Axis in DOCA-salt Hypertension

Immunohistochemical staining of chemerin (a), ChemR23 (b) and CD68 (c) in thoracic aorta and superior mesenteric artery from the sham normotensive (left) and DOCA-salt hypertensive rat (right). Representative of 4-5 different animals for each group. Arrows are regions of interest with boxed insets magnified. Bars = 100 microns.

Figure 5. Amplification of Chemerin-9 induced Contraction in DOCA-salt hypertension

Baseline contraction of the thoracic aorta (a) and superior mesenteric artery (b) from sham normotensive and DOCA-salt hypertensive (+E, −PVAT). Points represent means±SEM for number of animals in parentheses, * Significant differences from sham response (p<0.05). c, relationship of state of endothelial cell integrity (relaxation of acetylcholine, x-axis) to magnitude of contraction to chemerin-9 (1 μM). Thirty-two different samples (aorta, superior mesenteric artery) from rodents (sham, DOCA-salt, SHRSP, WKY, diet-induced obese and control).

Figure 6. The Chemerin Axis in Human Arteries

Immunohistochemical localization of ChemR23 (a, higher magnification right) and chemerin (b) in small mesenteric arteries isolated from intestinal bariatric specimens (obese humans). Representative of seven (7) different humans. c, Chemerin-9 induced contraction from baseline and in norepinephrine contracted arteries. Bars are means±SEM for number of patients in parentheses. * indicates statistically significant increase from baseline (p<0.05, unpaired Students t test). d, Chemerin-9 induced contraction from baseline and in PE contracted resistance arteries from the mesenteric arcade of the normal male Sprague-Dawley rat. e, Depiction of hypothesis. Chemerin, released from fat (PVAT, visceral fat) or tissue (liver), stimulates the ChemR23 receptor on vascular smooth muscle to elicit contraction, a process that is revealed in the presence of damaged endothelial function (gray endothelial cell).