Enhanced beta-1 adrenergic receptor responsiveness in coronary arterioles following intravenous stromal vascular fraction therapy in aged rats

Figures

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  Figure 1. Coronary flow reserve using Doppler echocardiography in rats. Stress test was performed on experimental groups using dobutamine (A) or adenosine (B) and CFR was calculated. O+SVF group exhibited increased CFR vs. OC in both adenosine and dobutamine conditions. P≤0.05 vs Young Control (*), vs Old Control (#), and vs Old+ SVF($); Data are presented as means±SD, analyzed with one-way ANOVA followed by post-hoc Dunn’s (A) or Holm-Sidak (B) test.

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  Figure 2. Diastolic function assessment using echocardiography and pressure- volume loop (PV-loop). Compared to YC rats, there was an age-related deterioration in diastolic function as measured echocardiographically by IVRT (A), E/A ratio (B), E/e’ ratio (C), and hemodynamically by Tau (D). Old rats treated with SVF significantly reversed this dysfunction in measures of E/A ratio (B) and E/e’ ratio (C) compared to OC, and normalized diastolic function to YC levels in IVRT (A) and Tau (D). P≤0.05 vs Young Control (*), vs Old Control (#), and vs Old+SVF ($); Data are presented as means±SD, analyzed with one-way ANOVA followed by post-hoc Holm-Sidak (B, D) or Dunn’s (A, C) test.

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  Figure 3. Isolated coronary arteriolar vasoreactivity to β1- and β2-AR agonists. Dobutamine, primarily a β1-AR agonist, induced vasorelaxation in all groups (A). Coronary arterioles from YC animals exhibited a significantly greater dilation compared to OC and O+BM at concentrations 1e-6 and 1e-5 [M] (*) (A). Salbutamol, a β2-AR agonist, induced mild vasorelaxation that was similar between all the groups (B). P≤0.05 vs Young Control (*), vs Old Control (#), and vs Old+SVF ($); data are presented as means±SEM and analyzed with two-way repeated measures ANOVA followed by post-hoc Bonferroni test.

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  Figure 4. Contribution of β1- and β2-AR to isoproterenol-induced vasodilation from isolated coronary arterioles. Vasorelaxation to isoproterenol, primarily a non-selective β1-, β2-, and β3-AR agonist, was significantly impaired in the O+BM group compared to O+SVF ($) (A). Isoproterenol with ICI118551, a β2-AR antagonist, eliminated differences between the groups. Compared to pre-incubation, all groups except OC (YC, O+SVF, and O+BM) had significant attenuation in the inhibited dose response (^) at several concentrations (B). Isoproterenol with CPG20712A, a β1-AR antagonist, also eliminated differences between the groups, and all groups exhibited significant attenuation in the response compared to pre-inhibition (^) (C). No group differences to isoproterenol were noted following inhibition with both ICI118551 and CPG20712A, and all groups exhibited significant attenuation in the response compared to pre-inhibition (^) at every concentration (D). P ≤ 0.05 vs Young Control (*), vs Old Control (#), vs Old+SVF, and pre- vs post-inhibition (^); data are presented as means±SEM and analyzed with two-way repeated measures ANOVA, paired for inhibitor analysis, followed by post-hoc Bonferroni test.

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  Figure 5. Contribution of β2-AR to norepinephrine-induced vasoreactivity in isolated coronary arterioles. Relaxation to NE was significantly impaired in OC animals compared to YC and O+SVF groups at all concentrations (A). NE with CPG20712A, a β1-AR antagonist, attenuated (^) the majority of the vasodilation response (3e-7 – 1e-4 [M]) in YC and O+BM treated groups (B). P≤0.05 vs Young Control (*), vs Old Control (#), vs Old+SVF ($), and pre- vs post-inhibition (^); data are presented as means±SEM and analyzed with two-way repeated measures ANOVA, paired for inhibitor analysis, followed by post-hoc Bonferroni test.

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  Figure 6. β1- and β2-AR immunofluorescence in isolated coronary arterioles. Representative images with ROI boxes used for fluorescent intensity analysis are shown on isolated coronary arterioles stained for β1- or β2-AR (A). There is no significant difference between the groups in the expression of β1- or β2-AR as measured by quantification of fluorescent intensity (B). Data are represented as means+SEM and analyzed with one-way ANOVA (n≥4). Scale bar is 50 micrometers.