Adiponectin improves endothelial function in mesenteric arteries of rats fed a high-fat diet: role of perivascular adipose tissue

Abstract

Background and purpose: Adiponectin, the most abundant peptide secreted by adipocytes, is involved in the regulation of energy metabolism and vascular physiology. Here, we have investigated the effects of exogenous administration of adiponectin on metabolism, vascular reactivity and perivascular adipose tissue (PVAT) of mesenteric arteries in Wistar rats fed a high-fat diet.

Experimental approach: The effects of adiponectin on NO-dependent and independent vasorelaxation were investigated in isolated mesenteric arteries from 12-month-old male Wistar rats (W12m) fed a high-fat diet (HFD) for 4 months and compared with those from age-matched rats given a control diet. Adiponectin ((96 μg·day^-1 ) was administered by continuous infusion with a minipump, implanted subcutaneously, for 28 days.

Key results: Chronic adiponectin treatment reduced body weight, total cholesterol, free fatty acids, fasting glucose and area under the curve of intraperitoneal glucose tolerance test, compared with HFD rats. It also normalized NO-dependent vasorelaxation increasing endothelial NO synthase (eNOS) phosphorylation in mesenteric arteries of HFD rats. In PVAT from aged (W12m) and HFD rats there was increased expression of chemokines and pro-inflammatory adipokines, the latter being important contributors to endothelial dysfunction. Infusion of adiponectin reduced these changes.

Conclusions and implications: Adiponectin normalized endothelial cell function by a mechanism that involved increased eNOS phoshorylation and decreased PVAT inflammation. Detailed characterization of the adiponectin signalling pathway in the vasculature and perivascular fat is likely to provide novel approaches to the management of atherosclerosis and metabolic disease.

Linked articles: This article is part of a themed section on Molecular Mechanisms Regulating Perivascular Adipose Tissue - Potential Pharmacological Targets? To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v174.20/issuetoc.

Figure 1

Effects of HFD and adiponectin on blood glucose levels during an IPGTT (A), AUC of IPGTT (B), fasting glycaemia (C) and plasma FFA (D) in W12m rats, compared with W6m rats. Blood glucose and FFA levels were determined at the end of treatment in all experimental groups [W6, W12m, HFD, HFDV or adiponectin (HFDA)]. The blood glucose was determined using a blood glucose monitoring system at 0, 30, 60 and 120 min after the glucose injection. Results are mean ± SE (n = 11 animals in each group). *P < 0.05, significantly different from W6m group; §P < 0.05, significantly different from W12m group; φP < 0.05, significantly different from HFD group; ΔP < 0.05, significantly different from HFDV group.

Figure 2

Effects of HFD and adiponectin treatment on relaxant responses to ACh (A) and sodium nitroprusside (B) in segments of mesenteric arteries from W12m rats, compared with W6m rats, after phenylephrine precontraction. Relaxation was measured using an isometric force displacement transducer in all groups including HFD treated with HFDV or adiponectin (HFDA). Data are expressed as mean ± SE (n = 11, 22 vascular ring preparations in 11 animals per group). *P < 0.05 significantly different from W6m group; §P < 0.05, significantly different from W12m group; φP < 0.05, significantly different from HFD group; ΔP < 0.05, significantly different from HFDV group.

Figure 3

In situ detection of superoxide (A–E) and nitrotyrosine levels (F–J) in rat mesenteric arteries. Representative DHE‐stained mesenteric artery sections reflect production of superoxide (A–E) with the different treatments. The endothelium is facing up in all layers. At identical settings, fluorescence in sections from W12m rats (B) was increased compared those from W6m rats (A). Note the markedly increased fluorescence, reflecting superoxide levels in the endothelium, intima and media of HFD rats (HFD and HFDV; C and D respectively). Fluorescence decreased to basal levels (as in W6m rats) in the adiponectin (HFDA)‐treated group (E). Panel (K) shows quantification of the fluorescence ethidium signal in the different groups of arteries. Representative mesenteric sections showing nitrotyrosine staining (F–J), indicative of increased peroxynitrite formation, in W6m (F), W12m (G), HFD (H), HFDV (I) and HFD treated with adiponectin (HFDA; J) rats. Panel (L) contains quantification of the green fluorescence in the different groups of arteries. Data are mean ± SE (n = 11 animals per group). *P < 0.05, significantly different from W6m group; §P < 0.05, significantly different from W12m group; φ P < 0.05, significantly different from HFD group; ΔP < 0.05, HFDV group.

Figure 4

Effects of HFD and adiponectin treatment on mesenteric eNOS levels in W12m rats, compared with W6m rats. Representative mesenteric sections demonstrating decreased p‐eNOS staining in W12m, HFD and HFDV arteries (B–D). Panel presents mesenteric arteries from W6m (A), W12m (B), HFD (C), HFDV (D) and HFDA (E) groups of rats. Panel (F) contains quantification of the red (p eNOS) to green fluorescence (total eNOS;t eNOS) ratio in the different groups of arteries. Data are mean ± SE (n = 11 animals per group). *P < 0.05, significantly different from W6m group; § P < 0.05, significantly different from W12m group; φ P < 0.05, significantly different from HFD group; Δ P < 0.05, significantly different from HFDV group.

Figure 5

Effects of HFD and adiponectin treatment on systemic leptin/adiponectin ratio (A) and PVAT leptin levels (B, C) in W12m and W6m rats. PVAT lysates were analysed by SDS‐PAGE. Representative Western blot analyses of leptin expression in PVAT of the different groups of rats (B). Averaged densitometric data for the different groups expressed as a percentage of elevation over the W6m value, set to 100% after normalisation to actin by densitometry. Data are mean ± SE (n = 11 animals per group). * P < 0.05, significantly different from W6m group; § P < 0.05, significantly different from W12m group; φ P < 0.05, significantly different from HFD group; ΔP < 0.05, significantly different from HFDV group.

Figure 6

Effects of HFD and adiponectin treatment on chemokine (CCL2 and CCL5) levels in PVAT from W12m and W6m rats. (A, C) Representative Western blot analyses of CCL2 and CCL5 expressions in PVAT of the different groups of rats. (B, D) Averaged densitometric data for the different groups expressed as a percentage of elevation over the W6m value, set to 100% after normalisation to actin by densitometry. Data are mean ± SE (n = 11 animals per group). *P < 0.05, significantly different from W6m group; §P < 0.05, significantly different from W12m group; φ P < 0.05, significantly different from HFD group; ΔP < 0.05, significantly different from HFDV group.

Figure 7

Effects of HFD and adiponectin treatment on lipocalin‐2 levels in PVAT from W12m and W6m rats. (A) Representative Western blot analyses of lipocalin‐2 expression in PVAT of mesenteric arteries of the different groups of rats. (B) Averaged densitometric data for groups expressed as a percentage of elevation over the W6m value, set to 100% after normalisation to actin by densitometry. Data are expressed as mean ± SE (n = 11 animals per group). *P < 0.05, significantly different from W6m group; §P < 0.05, significantly different from W12m group; φP < 0.05, significantly different from HFD group; ΔP < 0.05, significantly different from HFDV group.