Vaccine Targeting Alpha 1D-Adrenergic Receptor Improved Metabolic Syndrome in Mice

Figures

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  Fig. 1. ADRQβ-004 vaccine improved obesity and decreased SBP. a Experimental design. C57BL/6N mice were maintained on HFD + L-NAME diet and analyzed at 18 weeks. b The sodium dodecyl sulfate-polyacrylamide gel electrophoresis image of the ADRQβ-004 vaccine. c ADR-004 specific antibody titers were screened on days 0, 18, 24, 37, 52, 80, 95, 111, and 126. d Body weight of mice from different experimental groups during 18 weeks of diet. e Representative photographs of body size in different experimental groups. f The SBP of each group during the study measured by tail-cuff method. All data are expressed as the mean ± SD. *P < 0.05, **P < 0.01 and ***P < 0.001 vs the control group; #P < 0.05, ##P < 0.01 and ###P < 0.001 vs the HFD + L-NAME group. ADRQβ-004 indicates the ADRQβ-004 vaccine group; Con, the control group; HFD + L-NAME, the HFD + L-NAME group; HFD, high-fat diet; L-NAME, Nω-nitro-L-arginine methyl ester; SBP, systolic blood pressure; VLP, the Qβ virus-like particle

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  Fig. 2. The ADRQβ-004 vaccine significantly improved fasting blood glucose and insulin resistance, and partly ameliorated dyslipidemia. a–c Fasting blood glucose, fasting serum insulin, HOMA-IR values, and non-fasting serum insulin measured after 18 weeks of diet. d The level of serum leptin at the end of the experiment. e, f IPGTT and ITT as well as their AUC values of each after 18 weeks of diet. g–j The serum TG, TC, LDL-C, and HDL-C concentrations after 18 weeks of diet. k The level of serum FFA at the end of the experiment. All data are expressed as the mean ± SD. *P < 0.05, **P < 0.01 and ***P < 0.001 vs the control group; #P < 0.05, ##P < 0.01 and ###P < 0.001 vs the HFD + L-NAME group. ADRQβ-004 indicates the ADRQβ-004 vaccine group; AUC, area under the curve; Con, the control group; FFA, free fatty acid; HDL-C, high-density lipoprotein cholesterol; HFD + L-NAME, the HFD + L-NAME group; HOMA-IR, homeostasis model assessment of insulin resistance; IPGTT, intraperitoneal glucose-tolerance test; ITT, insulin-tolerance test; LDL-C, low-density lipoprotein cholesterol; TC, total cholesterol; TG, triglyceride

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  Fig. 3. ADRQβ-004 vaccine attenuated LV remodeling and diastolic dysfunction. a Representative images of H&E (upper bar = 500 μm, lower bar = 50 μm) and WGA (bar = 50 μm) staining in transversal sections of left ventricle of different experimental groups. b Ratio between heart weight and tibia length. c Quantification of LV cardiomyocyte CSA. d CD31 (bar = 100 μm) staining in transversal sections of LV. e Quantification of LV myocardial capillary density. f LV mass calculated by echocardiography. g Ratio between wet and dry lung weight. h The serum level of NT-proBNP. i Representative LV M-mode echocardiographic tracings. j, k Percentage of LVEF, and LVFS calculated by echocardiography. l Representative LV pulsed-wave Doppler tracings. m, n Ratio between mitral E wave and A wave, and IVRT calculated by Doppler imaging. All data are expressed as the mean ± SD. ADRQβ-004 indicates the ADRQβ-004 vaccine group; CD31, cluster of differentiation 31; Con, the control group; CSA, cross-sectional area; HFD + L-NAME, the HFD + L-NAME group; H&E, hematoxylin and eosin; IVRT, isovolumic relaxation time; LV, left ventricular; LVEF, left ventricular ejection fraction; LVFS, left ventricular fractional…

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  Fig. 4. ADRQβ-004 vaccine decreased fat mass and adipocyte size, and relieved tissue inflammation in eWAT. a Representative H&E (bar = 200 μm) staining of cross-sections of eWAT isolated from each experimental group. b Ratio between eWAT weight and tibia length. c Quantification of adipocyte’s CSA. d Representative F4/80 (bar = 200 μm) staining of cross-sections of eWAT. e Quantification of macrophageocyte density in eWAT. f Relative mRNA level of F4/80. g Relative mRNA levels of pro- and anti-inflammatory genes. h Relative mRNA levels of lipid metabolism. All data are expressed as the mean ± SD. ADRQβ-004 indicates the ADRQβ-004 vaccine group; Arcp30, adiponectin; Arg1, Arginase-1; Con, the control group; CD36, fatty acid translocase; CSA, cross-sectional area; eWAT, epididymal white adipose tissue; FABP4, fatty acid-binding protein 4; FATP1, fatty acid transport protein 1; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HFD + L-NAME, the HFD + L-NAME group; H&E, hematoxylin and eosin; MCP1, monocyte chemoattractant protein 1; PPARγ, peroxisome proliferator activated receptor γ; TNFα, tumor necrosis factor alpha; VLDLR, very-low-density lipoprotein receptor

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  Fig. 5. ADRQβ-004 vaccine alleviated hepatic steatosis elicited by HFD + L-NAME-feeding. a, b Representative images of H&E (upper bar = 500 μm, lower bar = 20 μm) staining and Oil Red O (upper bar = 500 μm, lower bar = 20 μm) staining of liver section from each group. c Ratio between liver weight and tibia length. d Percentage of OR area. e, f The hepatic TG and TC contents. g, h Gene expressions of lipogenic transcription factors, lipid uptake, and transport, respectively. i The protein expression level of PPARβ in the liver. All data are expressed as the mean ± SD. ADRQβ-004 indicates the ADRQβ-004 vaccine group; CD36, fatty acid translocase; Con, the control group; Fas, fatty acid synthase; FATP4, fatty acid transport protein 4; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; H&E, hematoxylin and eosin; HFD + L-NAME, the HFD + L-NAME group; LXRα, liver X receptor α; OR, Oil Red O; PPAR, peroxisome proliferator activated receptor; TC, total cholesterol; TG, triglyceride; SREBP1, sterol regulatory element-binding protein; VLDLR, very-low-density lipoprotein receptor

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  Fig. 6. ADRQβ-004 vaccine regulated systematic sympathetic activity and that of end-organs. a, b The blood CA and NE concentrations. c–e The protein expression level of TH in eWAT, liver, and heart respectively. f–h mRNA levels of α1-AR subtypes in eWAT, liver, and heart. Data are expressed as mean ± SD. ADRQβ-004 indicates the ADRQβ-004 vaccine group; α1-AR, α1-adrenergic receptor; CA, catecholamine; Con, the control group; eWAT, epididymal white adipose tissue; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HFD + L-NAME, the HFD + L-NAME group; NE, norepinephrine; TH, tyrosine hydroxylase