Perivascular adipose tissue from female rats fed a high‐fat diet impaired mesenteric artery vasodilation

Figures

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  FIGURE 1. Summary of experimental methods. Thoracic PVAT from rats fed a HFD or NCD for 16 weeks was excised and used for PVAT phenotyping or incubated in media for 24 h. Pressure myography was used to quantify the effects of PVAT‐conditioned media on mesenteric artery vasodilation. Finally, we determined how PVAT‐conditioned media affects endothelial ROS production and eNOS activation as markers for NO bioavailability.

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  FIGURE 2. HFD did not significantly increase male and female Sprague Dawley rat body or PVAT weight over NCD. Body weight over 16 weeks of NCD or HFD feeding in female (a) and male (b) rats. Body weight at study endpoint in NCD or HFD female and male rats (c). Thoracic PVAT weight at study endpoint in NCD or HFD female and male rats (d). Thoracic PVAT weight per body weight at study endpoint in NCD or HFD female and male rats (e). n = 6 rats per group. Data represented as mean ± standard deviation. Statistical significance was determined using a repeated measures ANOVA (a, b) or a two‐way ANOVA followed by Fisher's LSD test (c–e).

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  FIGURE 3. HFD increased some inflammatory biomarkers in PVAT from female rats compared to male rats. Relative gene expression of PVAT leptin (a), adiponectin (b), and CD68 (c). Representative Western blot and quantification of SIRT1 (d) and acetyl‐NF‐κB:NF‐κB ratio (e). n = 6 rats per group. Data are represented as mean ± standard deviation. Statistical significance was determined using a two‐way ANOVA followed by Fisher's LSD test. The full Western blots are provided in the Data S1.

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  FIGURE 4. PVAT‐conditioned media from HFD female rats impaired mesenteric artery vasodilation. Pressure myography and pEC50 from mesenteric arteries undergoing acetylcholine dose–response after 30‐min treatment with PSS (untreated; n = 4–9) or 10% PVAT conditioned media from NCD male rats (n = 7; a, b), HFD male rats (n = 4; c, d), NCD female rats (n = 7; e, f), and HFD female rats (n = 5; g, h). PVAT conditioned media effects on vasodilation were tested on mesenteric arteries from sex and diet‐matched rats. Data are represented as mean ± standard deviation. Statistical significance for panels b, d, f, and h was determined using a Mann–Whitney test.

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  FIGURE 5. PVAT‐conditioned media from male and female rats did not impair smooth muscle cell relaxation. Pressure myography of mesenteric arteries undergoing DEA‐NONOate (NO donor) dose–response after 30‐min treatment with PSS (n = 4–7; untreated), or 10% conditioned media from NCD male rats (n = 7; a), HFD male rats (n = 3; b), NCD female rats (n = 6; c), and HFD female rats (n = 4; d). PVAT conditioned media effects on smooth muscle cell relaxation were tested on mesenteric arteries from sex and diet‐matched rats. Data are represented as mean ± standard deviation. Statistical significance was tested using a repeated measures ANOVA.

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  FIGURE 6. PVAT‐conditioned media from HFD rats did not increase endothelial oxidative stress. Representative fluorescent microscopy images and quantification of RAECs treated with PVAT‐conditioned media from male and female rats fed an NCD or HFD. Scale bar = 200 μm. All data were normalized to cell count. Data are represented as mean ± standard deviation using conditioned media from n = 5 rats. Statistical significance was determined using a one‐way ANOVA.

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  FIGURE 7. PVAT‐conditioned media from HFD rats did not impair eNOS activation or deactivation. Representative Western blots and quantification of eNOS (Ser1177; a), and eNOS (Thr495) phosphorylation (b) in RAEC pretreated with PVAT‐conditioned media from male and female rats fed an NCD or HFD and then treated with 2 μM Yoda1 (Y) for 30 min. All data normalized to GAPDH and one cell sample that was included on each Western blot. Data are represented as mean ± standard deviation from n = 6 rats per group. Statistical significance was determined using a two‐way ANOVA followed by Tukey multiple comparison test. The full Western blots are provided in the Data S1.