Search

The human cationic anti-microbial peptide LL-37 is a T cell self-antigen in patients with psoriasis, who have increased risk of cardiovascular events. However, the role of LL-37 as a T cell self-antigen in the context of atherosclerosis remains unclear. The objective of this study was to test for the presence of T cells reactive to LL-37 in patients with acute coronary syndrome (ACS). Furthermore, the role of T cells reactive to LL-37 in atherosclerosis was assessed using apoE-/- mice immunized with the LL-37 mouse ortholog, mCRAMP. Peripheral blood mononuclear cells (PBMCs) from patients with ACS were stimulated with LL-37. PBMCs from stable coronary artery disease (CAD) patients or self-reported subjects served as controls. T cell memory responses were analyzed with flow cytometry. Stimulation of PBMCs with LL-37 reduced CD8+ effector T cell responses in controls and patients with stable CAD but not in ACS and was associated with reduced programmed cell death protein 1 (PDCD1) mRNA expression. For the mouse studies, donor apoE-/- mice were immunized with mCRAMP or adjuvant as controls, then T cells were isolated and adoptively transferred into recipient apoE-/- mice fed a Western diet. Recipient mice were euthanized after 5 weeks. Whole aortas and hearts were collected for analysis of atherosclerotic plaques. Spleens were collected for flow cytometric and mRNA expression analysis. Adoptive transfer experiments in apoE-/- mice showed a 28% reduction in aortic plaque area in mCRAMP T cell recipient mice (P < 0.05). Fifty six percent of adjuvant T cell recipient mice showed calcification in atherosclerotic plaques, compared to none in the mCRAMP T cell recipient mice (Fisher's exact test P = 0.003). Recipients of T cells from mice immunized with mCRAMP had increased IL-10 and IFN- γ expression in CD8+ T cells compared to controls. In conclusion, the persistence of CD8+ effector T cell response in PBMCs from patients with ACS stimulated with LL-37 suggests that LL-37-reactive T cells may be involved in the acute event. Furthermore, studies in apoE-/- mice suggest that T cells reactive to mCRAMP are functionally active in atherosclerosis and may be involved in modulating plaque calcification., (Copyright © 2020 Chernomordik, Cercek, Lio, Mihailovic, Yano, Herscovici, Zhao, Zhou, Chyu, Shah and Dimayuga.)

Generalized arterial calcification of infancy (GACI) and pseudoxanthoma elasticum (PXE) are characterized by pathologic calcifications in the media of large- and medium sized arteries. GACI is associated with biallelic mutations in ENPP1 in the majority of cases, whereas mutations in ABCC6 are known to cause PXE. Different treatment approaches including bisphosphonates and orally administered pyrophosphate (PP i ) were investigated in recent years, but reversion of calcification could not be achieved. With this study, we pursued the idea of a combination of controlled drug delivery through nanoparticles and active targeting via antibody conjugation to develop a treatment for GACI and PXE. To establish a suitable drug delivery system, the chelating drug diethylenetriamine pentaacetic acid (DTPA) was conjugated to nanoparticles composed of human serum albumin (HSA) as biodegradable and non-toxic particle matrix. To accomplish an active targeting of the elastic fibers exposed through calcification of the affected areas, the nanoparticle surface was functionalized with an anti-elastin antibody. Cytotoxicity and cell interaction studies revealed favorable preconditions for the intended i.v. application. The chelating ability was evaluated in vitro and ex vivo on aortic ring culture isolated from two mouse models of GACI and PXE. The positive results led to the conclusion that the produced nanoparticles might be a promising therapy in the treatment of GACI and PXE., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Aims: Smooth muscle cells (SMCs) play a role in medial vascular calcification, which can be stimulated by high levels of serum phosphate and inflammatory mediators. The aim of this study was to investigate whether mitogen-activated protein kinases (MAPKs) (p38 MAPK, ERK1/2, and JNK) and protein kinase A (PKA) can participate in inorganic phosphate (Pi)- and inflammation response-stimulated SMC calcification., Main Methods: We examined the change of Pi- and/or inflammation-stimulated human aortic smooth muscle cell (HASMC) calcification in the presence and absence of inhibitors or small interfering RNAs., Key Findings: Ca levels were increased in HASMCs incubated with 1.5-3.9 mM Pi, but not with 0.9 mM Pi or compared with non-Pi-treated HASMCs. Furthermore, the addition of interferon-γ (IFN-γ) increased pro-inflammatory cytokines [interleukin (IL)-1α, IL-6, and tumor necrosis factor-α (TNF-α)] in media containing Raw 264.7 cells. Ca levels were significantly increased in HASMCs cultured in IFN-γ-treated medium, compared with non-IFN-γ-treated medium in the presence of Pi (0.9-2.4 mM). The inhibition of p38 MAPK and PKA decreased HASMC calcification stimulated by Pi and IFN-γ-treated medium, though PKA inhibition produced a more significant reduction in calcification than p38 MAPK inhibition., Significance: These results indicate that PKA inhibition can efficiently reduce Pi- and inflammation-stimulated SMC calcification., (Copyright © 2018. Published by Elsevier Inc.)

Background: Given the link between cholesterol and activation of inflammation via interleukin 1β (IL-1β), we tested the effects of IL-1β inhibition on atherosclerotic calcification in mice. Patients with familial hypercholesterolemia develop extensive aortic calcification and calcific aortic stenosis. Although statins delay this process, low-density lipoprotein (LDL) cholesterol lowering alone is not enough to avert it. Data suggest that vascular inflammation initiated by hypercholesterolemia is followed by unchecked mineralization at sites of atherosclerotic plaques. The LDL-receptor (LDLR)-deficient (Ldlr(-/-)) and LDLR-attenuated Pcsk9(Tg) mice are available animal models for pharmacological testing., Methods: A mouse monoclonal antibody (mAb) against IL-1β or placebo was administered subcutaneously in Ldlr(-/-) and Pcsk9(Tg) models fed a Western diet. Drug level, anthropometric, lipid, and glucose profiles were determined. Expressions of proprotein convertase subtilisin/kexin type 9 (PCSK9), serum amyloid A1, and cytokine were measured by enzyme-linked immunosorbent assay. Aortic calcification was determined by microcomputerized tomography (micro-CT) and X-ray densitometry, and aortic flow velocity was assessed by ultrasound., Results: Circulating levels of IL-1β in Ldlr(-/-) mice were significantly greater (2-fold) than observed in Pcsk9(Tg) mice. Placebo- and mAb-treated mice did not differ in their growth, lipid, glucose profiles, and other cytokines. Calcifications were significantly diminished in mAb-treatment Ldlr(-/-) mice (a reduction of ∼ 75% by X-ray and ∼ 90% by micro-CT) and reduced insignificantly in mAb-treatment Pcsk9(Tg) mice, whereas aortic flow velocity was unchanged in both models., Conclusions: Herein, we demonstrate that aortic calcifications can be inhibited by an IL-1β mAb in LDLR-deficient mice. These results have a translational component to prevent vascular calcification in human and represent new evidence to rationalize targeting inflammation in cardiovascular disease., (© The Author(s) 2015.)