The anti-inflammatory effects of exercise training promote atherosclerotic plaque stabilization in apolipoprotein E knockout mice with diabetic atherosclerosis

Physical exercise is the cornerstone of cardiovascular disease treatment. The present study investigated whether exercise training affects atherosclerotic plaque composition through the modification of inflammatory-related pathways in apolipoprotein E knockout (apoE-/-) mice with diabetic atherosclerosis. Forty-five male apoE-/- mice were randomized into three equivalent (n=15) groups: control (CO), sedentary (SED), and exercise (EX). Diabetes was induced by streptozotocin administration. High-fat diet was administered to all groups for 12 weeks. Afterwards, CO mice were euthanatized, while the sedentary and exercise groups continued high-fat diet for 6 additional weeks. Exercising mice followed an exercise program on motorized-treadmill (5 times/week, 60 min/session). Then, blood samples and atherosclerotic plaques in the aortic root were examined. A considerable (P<0.001) regression of the atherosclerotic lesions was observed in the exercise group (180.339±75.613 x103μm2) compared to the control (325.485±72.302 x103μm2) and sedentary (340.188±159.108 x103μm2) groups. We found decreased macrophages, matrix metalloproteinase-2 (MMP-2), MMP-3, MMP-8 and interleukin-6 (IL-6) concentrations (P<0.05) in the atherosclerotic plaques of the exercise group. Compared to both control and sedentary groups, exercise training significantly increased collagen (P<0.05), elastin (P<0.001), and tissue inhibitor of matrix metalloproteinase-2 (TIMP-2) (P<0.001) content in the atherosclerotic plaques. Those effects paralleled with increased fibrous cap thickness and less internal elastic lamina ruptures after exercise training (P<0.05), while body-weight and lipid parameters did not significantly change. Plasma MMP-2 and MMP-3 concentrations in atherosclerotic tissues followed a similar trend. From our study we can conclude that exercise training reduces and stabilizes atherosclerotic lesions in apoE-/- mice with diabetic atherosclerosis. A favorable modification of the inflammatory regulators seems to explain those beneficial effects

Ablation of the chaperone protein ERDJ5 results in a Sjögren's syndrome-like phenotype in mice, consistent with an upregulated unfolded protein response in human patients

Objective: Sjögren's syndrome (SS) is a chronic autoimmune disorder that affects mainly the exocrine glands. Endoplasmic reticulum (ER) stress proteins have been suggested to participate in autoimmune and inflammatory responses, either acting as autoantigens, or by modulating factors of inflammation. The chaperone protein ERdj5 is an ER-resident disulfide reductase, required for the translocation of misfolded proteins during ER-associated protein degradation. In this study we investigated the role of ERdj5 in the salivary glands (SGs), in association with inflammation and autoimmunity. Methods: In situ expression of ERdj5 and XBP1 activation were studied immunohistochemically in minor SG tissues from primary SS patients and non-SS sicca-complaining controls. We used the mouse model of ERdj5 ablation and characterized its features: Histopathological, serological (antinuclear antibodies and cytokine levels), and functional (saliva flow rate). Results: ERdj5 was highly expressed in the minor SGs of SS patients, with stain intensity correlated to inflammatory lesion severity and antiSSA/Ro positivity. Moreover, SS patients demonstrated higher XBP1 activation within the SGs. Remarkably, ablation of ERdj5 in mice conveyed many of the cardinal features of SS, like spontaneous inflammation in SGs with infiltrating T and B lymphocytes, distinct cytokine signature, excessive cell death, reduced saliva flow, and production of anti-SSA/Ro and anti-SSB/La autoantibodies. Notably, these features were more pronounced in female mice. Conclusions: Our findings suggest a critical connection between the function of the ER chaperone protein ERdj5 and autoimmune inflammatory responses in the SGs and provide evidence for a new, potent animal model of SS. Copyright © 2019 Apostolou, Moustardas, Iwawaki, Tzioufas and Spyrou. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms

Deregulation of the Kallikrein Protease Family in the Salivary Glands of the Sjögren’s Syndrome ERdj5 Knockout Mouse Model

Introduction: The purpose of this study was to identify differentially expressed proteins in salivary glands of the ERdj5 knockout mouse model for Sjögren’s syndrome and to elucidate possible mechanisms for the morbid phenotype development. At the same time, we describe for the first time the sexual dimorphism of the murine submandibular salivary gland at the proteome level. Methods: We performed Liquid Chromatography/Mass Spectrometry in salivary gland tissues from both sexes of ERdj5 knockout and 129SV wildtype mice. The resulting list of proteins was evaluated with bioinformatic analysis and selected proteins were validated by western blot and immunohistochemistry and further analyzed at the transcription level by qRT-PCR. Results: We identified 88 deregulated proteins in females, and 55 in males in wildtype vs knockout comparisons. In both sexes, Kallikrein 1b22 was highly upregulated (fold change>25, ANOVA p<0.0001), while all other proteases of this family were either downregulated or not significantly affected by the genotype. Bioinformatic analysis revealed a possible connection with the downregulated NGF that was further validated by independent methods. Concurrently, we identified 416 proteins that were significantly different in the salivary gland proteome of wildtype female vs male mice and highlighted pathways that could be driving the strong female bias of the pathology. Conclusion: Our research provides a list of novel targets and supports the involvement of an NGF-mediating proteolytic deregulation pathway as a focus point towards the better understanding of the underlying mechanism of Sjögren’s syndrome. © Copyright © 2021 Moustardas, Yamada-Fowler, Apostolou, Tzioufas, Turkina and Spyrou

The beneficial effects of a direct thrombin inhibitor, dabigatran etexilate, on the development and stability of atherosclerotic lesions in apolipoprotein e-deficient mice

Purpose Dabigatran etexilate (DE) constitutes a novel, direct thrombin inhibitor. Regarding the association of thrombin with atherogenesis, we assessed the effects of DE on the development and stability of atherosclerotic lesions in apolipoprotein-E deficient (ApoE-/-) mice. Materials-methods Fifty male ApoE-/- mice were randomized to receive western-type diet either supplemented with DE 7.5 mg DE/g chow) (DE-group, n025) or matching placebo as control (CO-group, n025) for 12 weeks. After this period, all mice underwent carotid artery injury with ferric chloride and the time to thrombotic total occlusion (TTO) was measured. Then, mice were euthanatized and each aortic arch was analyzed for the mean plaque area, the content of macrophages, elastin, collagen, nuclear factor kappaB (NF?B), vascular cell adhesion molecule-1 (VCAM-1), matrix metalloproteinase-9 (MMP-9) and its inhibitor (TIMP-1). Results DE-group showed significantly longer TTO compared to CO-group (8.9±2.3 min vs 3.5±1.1 min, p<0.001) and the mean plaque area was smaller in DE-group than CO-group (441.00±160.01×103μm2 vs 132.12±32.17× 103μm2, p<0.001). Atherosclerotic lesions derived from DE-treated mice showed increased collagen (p00.043) and elastin (p00.031) content, thicker fibrous caps (p<0.001) and reduced number of internal elastic lamina ruptures per mm of arterial girth (p<0.001) when compared to COgroup. Notably, DE treatment seemed to promote plaque stability possibly by reducing concentrations of NF?B, VCAM-1, macrophages and MMP-9 and increasing TIMP-1 within atherosclerotic lesions (p<0.05). Conclusions DE attenuates arterial thrombosis, reduces lesion size and may promote plaque stability in ApoE-/- mice. The plaque-stabilizing effects of chronic thrombin inhibition might be the result of the favorable modification of inflammatory mechanisms. © Springer Science+Business Media, LLC 2012

The role of exercise training and the endocannabinoid system in atherosclerotic plaque burden and composition in apo-E-deficient mice

Introduction: We investigated the effect of combining exercise training and treatment with an endocannabinoid receptor 1 inhibitor (Rimonabant) on atherosclerosis burden and composition. Methods: Forty-eight apolipoprotein E-deficient (ApoE-/-) mice were kept on a 16-week highfat diet. Mice were then placed on a normal diet and were randomized to the following groups with n=12 mice for 6 more weeks: 1) Control (Co)-no intervention; 2) Exercise (Ex)-exercise training on treadmill; 3) Rimonabant (Ri)-oral administration of rimonabant (10 mg/kg/day); or 4) RimonabantþExercise (RiEx)-combination of Ri and Ex groups treatment. At the end, all animals were sacrificed, and blood samples, as well as aortic root specimens, were obtained for histomorphometric analysis and quantification of the serum and plaque content of matrix metalloproteinases (MMPs). Results: The mean plaque area was significantly smaller (RiEx: 43.18 ±1.72%, Ri: 44.66 ± 3.1%, Ex: 49 ± 4.10%, Co: 70.43 ± 2.83%) in all active treatment groups relative to the Co group (p < 0.01). Conversely, the relative concentrations of collagen and elastin were increased significantly across all treatment groups compared to Co (p < 0.05). Immunohistochemical analysis revealed significantly reduced macrophage content within plaques after all interventions, with the most pronounced effect observed after combined treatment (RiEx: 9.4 ± 3.92%, Ri: 15 ± 2.45%, Ex: 19.78 ± 2.79%, Co: 34.25 ± 4.99%; p < 0.05). Within plaques, the TIMP-1 concentration was significantly upregulated in exercise-treated groups. MMP-3 and MMP-9 concentrations were equivalently decreased in all three active treatment groups compared to controls (p < 0.001). Discussion: Both exercise and rimonabant treatments induced plaque regression and promoted plaque stability. The combined treatment failed to show additive or synergistic benefits relative to either intervention alone. © 2016 Hellenic Society of Cardiology

The anti-inflammatory effects of exercise training promote atherosclerotic plaque stabilization in apolipoprotein E knockout mice with diabetic atherosclerosis

Physical exercise is the cornerstone of cardiovascular disease treatment. The present study investigated whether exercise training affects atherosclerotic plaque composition through the modification of inflammatoryrelated pathways in apolipoprotein E knockout (apoE-/-) mice with diabetic atherosclerosis. Forty-five male apoE-/mice were randomized into three equivalent (n=15) groups: control (CO), sedentary (SED), and exercise (EX). Diabetes was induced by streptozotocin administration. High-fat diet was administered to all groups for 12 weeks. Afterwards, CO mice were euthanatized, while the sedentary and exercise groups continued high-fat diet for 6 additional weeks. Exercising mice followed an exercise program on motorizedtreadmill (5 times/week, 60 min/session). Then, blood samples and atherosclerotic plaques in the aortic root were examined. A considerable (P<0.001) regression of the atherosclerotic lesions was observed in the exercise group (180.339±75.613×103μm2) compared to the control (325.485±72.302 ×103 μm2) and sedentary (340.188±159.108×103μm2) groups. We found decreased macrophages, matrix metalloproteinase-2 (MMP-2), MMP-3, MMP-8 and interleukin-6 (IL-6) concentrations (P<0.05) in the atherosclerotic plaques of the exercise group. Compared to both control and sedentary groups, exercise training significantly increased collagen (P<0.05), elastin (P<0.001), and tissue inhibitor of matrix metalloproteinase-2 (TIMP-2) (P<0.001) content in the atherosclerotic plaques. Those effects paralleled with increased fibrous cap thickness and less internal elastic lamina ruptures after exercise training (P<0.05), while body-weight and lipid parameters did not significantly change. Plasma MMP-2 and MMP-3 concentrations in atherosclerotic tissues followed a similar trend. From our study we can conclude that exercise training reduces and stabilizes atherosclerotic lesions in apoE-/-mice with diabetic atherosclerosis. A favorable modification of the inflammatory regulators seems to explain those beneficial effects. © N.P.E. Kadoglou et al., 2013 Licensee PAGEPress, Italy

Effects of exercise training on the severity and composition of atherosclerotic plaque in apoE-deficient mice

Aim: To investigate the effects of exercise on atherosclerotic plaque composition, the concentration of matrix metalloproteinases (MMPs) in the atherosclerotic plaque and the systemic circulation. Methods: Ninety apolipoprotein E-deficient (apoE-/-) mice (45 male) were randomized to the following groups (n = 15 each): control male/female; sedentary male/female; exercise male/female. Mice were kept on a 16-week high-fat diet. Subsequently, the control groups were sacrificed, while the rest of the animals were placed on a normal diet for 6 more weeks. During the latter period, the exercise groups were trained daily on treadmill. At the end of the study, mice were euthanized, and blood samples as well as aortic root specimens were obtained. Results: Compared to control and sedentary animals, exercise training reduced atherosclerotic plaques (-30%; p < 0.01) and increased elastin and collagen content in both genders (p < 0.05). Body weight or lipid profile did not change significantly. Decreased macrophages and MMP-9 as well as increased tissue inhibitor of metalloproteinases 1 (TIMP-1) levels were observed in the atherosclerotic plaques of the exercise-treated groups (p < 0.05). Plasma concentrations of MMP-9 decreased, while plasma TIMP-1 levels increased in the exercise compared to control and sedentary groups (p < 0.05). Conclusions: Exercise training had a favorable effect on the size and composition of the atherosclerotic plaque in apoE-/- mice, associated with suppressed MMP activity. Copyright © 2011 S. Karger AG