Angiogenic growth factors have recently been linked to tissue metabolism. We have used genetic gain‐ and loss‐of function models to elucidate the effects and mechanisms of action of vascular ...endothelial growth factor‐B (VEGF‐B) in the heart. A cardiomyocyte‐specific VEGF‐B transgene induced an expanded coronary arterial tree and reprogramming of cardiomyocyte metabolism. This was associated with protection against myocardial infarction and preservation of mitochondrial complex I function upon ischemia‐reperfusion. VEGF‐B increased VEGF signals via VEGF receptor‐2 to activate Erk1/2, which resulted in vascular growth. Akt and mTORC1 pathways were upregulated and AMPK downregulated, readjusting cardiomyocyte metabolic pathways to favor glucose oxidation and macromolecular biosynthesis. However, contrasting with a previous theory, there was no difference in fatty acid uptake by the heart between the VEGF‐B transgenic, gene‐targeted or wildtype rats. Importantly, we also show that VEGF‐B expression is reduced in human heart disease. Our data indicate that VEGF‐B could be used to increase the coronary vasculature and to reprogram myocardial metabolism to improve cardiac function in ischemic heart disease.
Synopsis
Transgenic expression of VEGF‐B in the rat heart leads to expansion of the coronary arterial tree and an increase in functional coronary reserve, accompanied by a shift in myocardial metabolism from fatty acid to glucose utilization.
VEGF‐B‐induced coronary artery growth protects the heart from ischemic damage
Cardiac hypertrophy in VEGF‐B transgenic rats is physiological even in old age
AAV‐VEGF‐B gene transfer to adult rats phenocopies the VEGF‐B transgenic rat heart
Glucose uptake is increased and expression of fatty acid oxidation genes decreased in the VEGF‐B transgenic hearts
VEGF‐B increases VEGF signaling via VEGFR‐2
Transgenic expression of VEGF‐B in the rat heart leads to expansion of the coronary arterial tree and an increase in functional coronary reserve, accompanied by a shift in myocardial metabolism from fatty acid to glucose utilization.
: The expression of matrix metalloproteinases 2/9 (MMP-2/9) has been implicated in arterial remodeling and inflammation in atherosclerosis. We evaluated a gallium-68 labeled peptide for the detection ...of MMP-2/9 in atherosclerotic mouse aorta.
: We studied sixteen low-density lipoprotein receptor deficient mice (LDLR
ApoB
) kept on a Western-type diet. Distribution of intravenously-injected MMP-2/9-targeting peptide,
GaGa-DOTA-TCTP-1, was studied by combined positron emission tomography (PET) and contrast-enhanced computed tomography (CT). At 60 min post-injection, aortas were cut into cryosections for autoradiography analysis of tracer uptake, histology, and immunohistochemistry. Zymography was used to assess MMP-2/9 activation and pre-treatment with MMP-2/9 inhibitor to assess the specificity of tracer uptake.
: Tracer uptake was not visible by in vivo PET/CT in the atherosclerotic aorta, but ex vivo autoradiography revealed 1.8 ± 0.34 times higher tracer uptake in atherosclerotic plaques than in normal vessel wall (
= 0.0029). Tracer uptake in plaques correlated strongly with the quantity of Mac-3-positive macrophages (R = 0.91,
< 0.001), but weakly with MMP-9 staining (R = 0.40,
= 0.099). Zymography showed MMP-2 activation in the aorta, and pre-treatment with MMP-2/9 inhibitor decreased tracer uptake by 55% (
= 0.0020).
: The MMP-2/9-targeting
GaGa-DOTA-TCTP-1 shows specific uptake in inflamed atherosclerotic lesions; however, a low target-to-background ratio precluded in vivo vascular imaging. Our results suggest, that the affinity of gelatinase imaging probes should be steered towards activated MMP-2, to reduce the interference of circulating enzymes on the target visualization in vivo.
Purpose
Rupture-prone atherosclerotic plaques are characterized by accumulation of macrophages, which have shown to express somatostatin type 2 receptors. We aimed to investigate whether somatostatin ...receptor-targeting positron emission tomography (PET) tracers,
68
GaDOTANOC,
18
FFDR-NOC, and
68
GaDOTATATE, can detect inflamed atherosclerotic plaques.
Procedures
Atherosclerotic IGF-II/LDLR
−/−
ApoB
100/100
mice were studied
in vivo
and
ex vivo
for tracer uptake into atherosclerotic plaques. Furthermore,
68
GaDOTANOC and
68
GaDOTATATE were compared in a head-to-head setting for
in vivo
PET/X-ray computed tomography (CT) imaging characteristics.
Results
Ex vivo
uptake of
68
GaDOTANOC and
68
GaDOTATATE in the aorta was higher in atherosclerotic mice compared to control C57Bl/6N mice, while the aortic uptake of
18
FFDR-NOC showed no genotype difference. Unlike
18
FFDR-NOC,
68
GaDOTANOC and
68
GaDOTATATE showed preferential binding to atherosclerotic plaques with plaque-to-wall ratio of 1.7 ± 0.3 and 2.1 ± 0.5, respectively. However, the aortic uptake and aorta-to-blood ratio of
68
GaDOTANOC were higher compared to
68
GaDOTATATE in
in vivo
PET/CT imaging.
Conclusion
Our results demonstrate superior applicability for
68
GaDOTANOC and
68
GaDOTATATE in the detection of atherosclerotic plaques compared to
18
FFDR-NOC.
Atherosclerosis is characterized by the accumulation of oxidized lipids in the artery wall, which triggers an inflammatory response. Oxidized low-density lipoprotein (ox-LDL) presents amyloid-like ...structural properties, and different amyloid species have recently been recognized in atherosclerotic plaques. Therefore, we studied the uptake of the amyloid imaging agent
FFlutemetamol in atherosclerotic plaques. The binding of
FFlutemetamol to human carotid artery plaque was studied in vitro. In vivo uptake of the tracer was studied in hypercholesterolemic IGF-II/LDLR
ApoB
mice and C57BL/6N controls. Tracer biodistribution was studied in vivo with PET/CT, and ex vivo by gamma counter and digital ex vivo autoradiography. The presence of amyloid, ox-LDL, and macrophages in the plaques was examined by immunohistochemistry.
FFlutemetamol showed specific accumulation in human carotid plaque, especially in areas positive for amyloid beta. The aortas of IGF-II/LDLR
ApoB
mice showed large thioflavin-S-positive atherosclerotic plaques containing ox-LDL and macrophages. Autoradiography revealed 1.7-fold higher uptake in the plaques than in a lesion-free vessel wall, but no difference in aortic tissue uptake between mouse strains were observed in the in vivo PET/CT. In conclusion,
FFlutemetamol binds to amyloid-positive areas in human atherosclerotic plaques. Further studies are warranted to clarify the uptake mechanisms, and the potential of the tracer for in vivo imaging of atherosclerosis in patients.
Given the important role of inflammation and the potential association of the leukocyte trafficking-associated adhesion molecule vascular adhesion protein 1 (VAP-1) with atherosclerosis, this study ...examined whether functional VAP-1 is expressed in atherosclerotic lesions and, if so, whether it could be targeted by positron emission tomography (PET). First, immunohistochemistry revealed that VAP-1 localized to endothelial cells of intra-plaque neovessels in human carotid endarterectomy samples from patients with recent ischemic symptoms. In low-density lipoprotein receptor-deficient mice expressing only apolipoprotein B100 (LDLR
ApoB
), VAP-1 was expressed on endothelial cells lining inflamed atherosclerotic lesions; normal vessel walls in aortas of C57BL/6N control mice were VAP-1-negative. Second, we discovered that the focal uptake of VAP-1 targeting sialic acid-binding immunoglobulin-like lectin 9 based PET tracer
GaDOTA-Siglec-9 in atherosclerotic plaques was associated with the density of activated macrophages (r = 0.58, P = 0.022). As a final point, we found that the inhibition of VAP-1 activity with small molecule LJP1586 decreased the density of macrophages in inflamed atherosclerotic plaques in mice. Our results suggest for the first time VAP-1 as a potential imaging target for inflamed atherosclerotic plaques, and corroborate VAP-1 inhibition as a therapeutic approach in the treatment of atherosclerosis.
The purpose of this study was to explore the feasibility of (11)C-choline in the assessment of the degree of inflammation in atherosclerotic plaques.
Uptake of (11)C-choline was studied ex vivo in ...tissue samples and aortic sections excised from 6 atherosclerotic mice deficient for both low-density lipoprotein receptor and apolipoprotein B48 (LDLR(-/-)ApoB(100/100)) and 5 control mice. The autoradiographs were compared with the immunohistology of the arterial sites.
The uptake of (11)C-choline (percentage of the injected activity per gram of tissue) in the atherosclerotic aortas of the LDLR(-/-)ApoB(100/100) mice was significantly higher (1.9-fold, P = 0.0016) than that in the aortas of the control mice. The autoradiography analysis showed significantly higher uptake of (11)C-choline in the plaques than in healthy vessel wall (mean ratio, 2.3 +/- 0.6; P = 0.014), prominently in inflamed plaques, compared with noninflamed plaque areas.
We observed a high (11)C-choline uptake in the aortic plaques of atherosclerotic mice. Our data suggest that macrophages may be responsible for the uptake of (11)C-choline in the plaques.
The balance between pro- and anti-inflammatory cytokines released by immune and non-immune cells plays a decisive role in the progression of atherosclerosis. Interleukin (IL)-17A has been shown to ...accelerate atherosclerosis. In this study, we investigated the effect on pro-inflammatory mediators and atherosclerosis development of an Affibody molecule that targets IL17A. Affibody molecule neutralizing IL17A, or sham were administered
to human aortic smooth muscle cells (HAoSMCs) and murine NIH/3T3 fibroblasts and
to atherosclerosis-prone, hyperlipidaemic ApoE
mice. Levels of mediators of inflammation and development of atherosclerosis were compared between treatments. Exposure of human smooth muscle cells and murine NIH/3T3 fibroblasts
to αIL-17A Affibody molecule markedly reduced IL6 and CXCL1 release in supernatants compared with sham exposure. Treatment of ApoE
mice with αIL-17A Affibody molecule significantly reduced plasma protein levels of CXCL1, CCL2, CCL3, HGF, PDGFB, MAP2K6, QDPR, and splenocyte mRNA levels of
, and
compared with sham exposure. There was no significant difference in atherosclerosis burden between the groups. In conclusion, administration of αIL17A Affibody molecule reduced levels of pro-inflammatory mediators and attenuated inflammation in ApoE
mice.
Atherosclerosis progression is modulated by interactions with the adaptive immune system. Humoral immunity can help protect against atherosclerosis formation; however, the existence, origin, and ...function of putative atherogenic antibodies are controversial. How such atherosclerosis-promoting antibodies could affect the specific composition and stability of plaques, as well as the vasculature generally, remains unknown.
We addressed the overall contribution of antibodies to atherosclerosis plaque formation, composition, and stability in vivo (1) with mice that displayed a general loss of antibodies, (2) with mice that had selectively ablated germinal center-derived IgG production, or (3) through interruption of T-B-cell interactions and further studied the effects of antibody deficiency on the aorta by transcriptomics.
Here, we demonstrate that atherosclerosis-prone mice with attenuated plasma cell function manifest reduced plaque burden, indicating that antibodies promote atherosclerotic lesion size. However, the composition of the plaque was altered in antibody-deficient mice, with an increase in lipid content and decreases in smooth muscle cells and macrophages, resulting in an experimentally validated vulnerable plaque phenotype. Furthermore, IgG antibodies enhanced smooth muscle cell proliferation in vitro in an Fc receptor-dependent manner, and antibody-deficient mice had decreased neointimal hyperplasia formation in vivo. These IgG antibodies were shown to be derived from germinal centers, and mice genetically deficient for germinal center formation had strongly reduced atherosclerosis plaque formation. mRNA sequencing of aortas revealed that antibodies are required for the sufficient expression of multiple signal-induced and growth-promoting transcription factors and that aortas undergo large-scale metabolic reprograming in their absence. Using an elastase model, we demonstrated that absence of IgG results in an increased severity of aneurysm formation.
We propose that germinal center-derived IgG antibodies promote the size and stability of atherosclerosis plaques, through promoting arterial smooth muscle cell proliferation and maintaining the molecular identity of the aorta. These results could have implications for therapies that target B cells or B-T-cell interactions because the loss of humoral immunity leads to a smaller but less stable plaque phenotype.
Autoantibodies to cardiac troponins (cTnAAbs) can interfere with the measurement of cardiac troponin I (cTnI) by immunoassays for the diagnosis of myocardial infarction. Therefore, we determined the ...cTnI binding sites and IgG subclasses of circulating cTnAAbs.
We studied epitope specificity with sandwich-type immunoassays by measuring the recovery of troponin complex added to 10 cTnAAb-negative and 10 cTnAAb-positive sera from healthy volunteers. To study the IgG subclasses, we analyzed admission and 3-month follow-up sera from chest pain patients with a reference assay measuring total IgG (14 cTnAAb negative and 14 cTnAAb positive at 3 months) and with 4 subclass-specific assays measuring exclusively IgG subclasses 1-4.
Mean recoveries of troponin complex in cTnAAb-positive samples for single cTnI epitopes ranged from 37% to 211%, being lowest for the cTnI midfragment (aa 30-110). However, the lowest sample-specific recoveries, 4%-92%, showed that none of the studied epitopes completely escaped the cTnAAb-related interference. Eight chest pain patients of the cTnAAb-positive group became positive between sampling points, and according to all 5 cTnAAb assays, specific signals were generally higher at follow-up. IgG4, with the highest prevalence, was detected in 68% of samples in the cTnAAb-positive group.
IgG subclass studies confirm that cTnAAb formation may be triggered/boosted in acute cardiac events. This new information about the epitope specificity of cTnAAbs should be used to reevaluate existing recommendations regarding use of midfragment epitopes in cTnI assays. To circumvent the negative interference of the highly heterogeneous cTnAAbs, use of 3 or more unconventionally selected epitopes should be considered.
Dipeptidyl peptidase 4 (DPP-4) inhibitors have anti-inflammatory and atheroprotective effects. We evaluated the effects of the DPP-4 inhibitor linagliptin on atherosclerotic plaque and hepatic ...inflammation using histology and 2-deoxy-2-18F-fluoro-d-glucose (18F-FDG), a positron emission tomography tracer of inflammation, in a mouse model of hypercholesterolemia and type 2 diabetes.
Igf2/Ldlr−/−Apob100/100 mice were fed a high-fat diet (HFD) for 8 weeks and then randomly allocated to receive HFD (n = 14), or HFD with added linagliptin (n = 15) for additional 12 weeks. Five mice fed a chow diet were studied as an additional control. At the end of the study, glucose tolerance, aortic and liver uptake of 18F-FDG, and histology were studied.
Mice in linagliptin and HFD groups had similar fasting glucose concentrations, but linagliptin improved glucose tolerance. Aortas of linagliptin and HFD groups showed advanced atherosclerotic plaques with no difference in the mean intima-to-media ratio or number of macrophages in the plaques. Autoradiography showed similar 18F-FDG uptake by atherosclerotic plaques in linagliptin and HFD groups (plaque-to-wall ratio: 1.7 ± 0.25 vs. 1.6 ± 0.21; p = 0.24). In the liver, linagliptin reduced the histologic inflammation score but had no effect on 18F-FDG uptake. Compared with chow diet, uptake of 18F-FDG was similar in the aorta, but higher in the liver after HFD.
Linagliptin therapy improved glucose tolerance and reduced hepatic inflammation but had no effect on plaque burden or atherosclerotic inflammation, as determined by histology and 18F-FDG uptake, in atherosclerotic mice with type 2 diabetes.
Display omitted
•Potential anti-inflammatory effects of linagliptin were studied by a PET tracer 18F-FDG in a mouse model of atherosclerosis and diabetes.•Linagliptin treatment did not affect atherosclerotic plaque size or inflammation.•Linagliptin treatment, however, did reduce hepatic inflammation.
PDF
![Evaluation of [ 68 Ga]Ga-DO...](http://api.summon.serialssolutions.com/2.0.0/image/issn/XR4PS5YY4K/1420-3049_1433-1373/small "Evaluation of [ 68 Ga]Ga-DOTA-TCTP-1 for the Detection of Metalloproteinase 2/9 Expression in Mouse Atherosclerotic Plaques")
