Myeloid cell-derived interleukin-6 induces vascular dysfunction and vascular and systemic inflammation.

Aims: The cytokine interleukin-6 (IL-6) plays a central role in the inflammation cascade as well as cardiovascular disease progression. Since myeloid cells are a primary source of IL-6 formation, we aimed to generate a mouse model to study the role of myeloid cell-derived IL-6 in vascular disease. Methods and results: Interleukin-6-overexpressing (IL-6OE) mice were generated and crossed with LysM-Cre mice, to generate mice (LysM-IL-6OE mice) overexpressing the cytokine in myeloid cells. Eight- to 12-week-old LysM-IL-6OE mice spontaneously developed inflammatory colitis and significantly impaired endothelium-dependent aortic relaxation, increased aortic reactive oxygen species (ROS) formation, and vascular dysfunction in resistance vessels. The latter phenotype was associated with decreased survival. Vascular dysfunction was accompanied by a significant accumulation of neutrophils, monocytes, and macrophages in the aorta, increased myeloid cell reactivity (elevated ROS production), and vascular fibrosis associated with phenotypic changes in vascular smooth muscle cells. In addition to elevated Mcp1 and Cxcl1 mRNA levels, aortae from LysM-IL-6OE mice expressed higher levels of inducible NO synthase and endothelin-1, thus partially accounting for vascular dysfunction, whereas systemic blood pressure alterations were not observed. Bone marrow (BM) transplantation experiments revealed that vascular dysfunction and ROS formation were driven by BM cell-derived IL-6 in a dose-dependent manner. Conclusion: Mice with conditional overexpression of IL-6 in myeloid cells show systemic and vascular inflammation as well as endothelial dysfunction. A decrease in circulating IL-6 levels by replacing IL-6-producing myeloid cells in the BM improved vascular dysfunction in this model, underpinning the relevant role of IL-6 in vascular disease.

Role of inflammatory signaling pathways involving the CD40-CD40L-TRAF cascade in diabetes and hypertension-insights from animal and human studies.

CD40L-CD40-TRAF signaling plays a role in atherosclerosis progression and affects the pathogenesis of coronary heart disease (CHD). We tested the hypothesis that CD40L-CD40-TRAF signaling is a potential therapeutic target in hyperlipidemia, diabetes, and hypertension. In mouse models of hyperlipidemia plus diabetes (db/db mice) or hypertension (1 mg/kg/d angiotensin-II for 7 days), TRAF6 inhibitor treatment (2.5 mg/kg/d for 7 or 14 days) normalized markers of oxidative stress and inflammation. As diabetes and hypertension are important comorbidities aggravating CHD, we explored whether the CD40L-CD40-TRAF signaling cascade and their associated inflammatory pathways are expressed in CHD patients suffering from comorbidities. Therefore, we analyzed vascular bypass material (aorta or internal mammary artery) and plasma from patients with CHD with diabetes and/or hypertension. Our Olink targeted plasma proteomic analysis using the IMMUNO-ONCOLOGY panel revealed a pattern of step-wise increase for 13/92 markers of low-grade inflammation with significant changes. CD40L or CD40 significantly correlated with 38 or 56 other inflammatory targets. In addition, specific gene clusters that correlate with the comorbidities were identified in isolated aortic mRNA of CHD patients through RNA-sequencing. These signaling clusters comprised CD40L-CD40-TRAF, immune system, hemostasis, muscle contraction, metabolism of lipids, developmental biology, and apoptosis. Finally, immunological analysis revealed key markers correlated with comorbidities in CHD patients, such as CD40L, NOX2, CD68, and 3-nitrotyrosine. These data indicate that comorbidities increase inflammatory pathways in CHD, and targeting these pathways will be beneficial in reducing cardiovascular events in CHD patients with comorbidities.

Vascular dysfunction and arterial hypertension in experimental celiac disease are mediated by gut-derived inflammation and oxidative stress.

AIMS: We examined the cardiovascular effects of celiac disease (CeD) in a humanized mouse model, with a focus on vascular inflammation, endothelial dysfunction, and oxidative stress. METHODS AND RESULTS: NOD.DQ8 mice genetically predisposed to CeD were subjected to a diet regime and oral gavage to induce the disease (gluten group vs. control). We tested vascular function, confirmed disease indicators, and evaluated inflammation and oxidative stress in various tissues. Plasma proteome profiling was also performed. CeD markers were confirmed in the gluten group, indicating increased blood pressure and impaired vascular relaxation. Pro-inflammatory genes were upregulated in this group, with increased CD11b+ myeloid cell infiltration and oxidative stress parameters observed in aortic and heart tissue. However, heart function remained unaffected. Plasma proteomics suggested the cytokine interleukin-17A (IL-17A) as a link between gut and vascular inflammation. Cardiovascular complications were reversed by adopting a gluten-free diet. CONCLUSION: Our study sheds light in the heightened cardiovascular risk associated with active CeD, revealing a gut-to-cardiovascular inflammatory axis potentially mediated by immune cell infiltration and IL-17A. These findings augment our understanding of the link between CeD and cardiovascular disease providing clinically relevant insight into the underlying mechanism. Furthermore, our discovery that cardiovascular complications can be reversed by a gluten-free diet underscores a critical role for dietary interventions in mitigating cardiovascular risks associated with CeD.

Vascular protein disulfide isomerase A1 mediates endothelial dysfunction induced by angiotensin II in mice.

AIM: Protein disulfide isomerases (PDIs) are involved in platelet aggregation and intravascular thrombosis, but their role in regulating endothelial function is unclear. Here, we characterized the involvement of vascular PDIA1 in angiotensin II (Ang II)-induced endothelial dysfunction in mice. METHODS: Endothelial dysfunction was induced in C57BL/6JCmd male mice via Ang II subcutaneous infusion, and PDIA1 was inhibited with bepristat. Endothelial function was assessed in vivo with magnetic resonance imaging and ex vivo with a myography, while arterial stiffness was measured as pulse wave velocity. Nitric oxide (NO) bioavailability was measured in the aorta (spin-trapping electron paramagnetic resonance) and plasma (NO2 - and NO3 - levels). Oxidative stress, eNOS uncoupling (DHE-based aorta staining), and thrombin activity (thrombin-antithrombin complex; calibrated automated thrombography) were evaluated. RESULTS: The inhibition of PDIA1 by bepristat in Ang II-treated mice prevented the impairment of NO-dependent vasodilation in the aorta as evidenced by the response to acetylcholine in vivo, increased systemic NO bioavailability and the aortic NO production, and decreased vascular stiffness. Bepristat's effect on NO-dependent function was recapitulated ex vivo in Ang II-induced endothelial dysfunction in isolated aorta. Furthermore, bepristat diminished the Ang II-induced eNOS uncoupling and overproduction of ROS without affecting thrombin activity. CONCLUSION: In Ang II-treated mice, the inhibition of PDIA1 normalized the NO-ROS balance, prevented endothelial eNOS uncoupling, and, thereby, improved vascular function. These results indicate the importance of vascular PDIA1 in regulating endothelial function, but further studies are needed to elucidate the details of the mechanisms involved.