Single-cell RNAseq identifies clonally expanded antigen-specific T-cells following intradermal injection of gold nanoparticles loaded with diabetes autoantigen in humans.

Gold nanoparticles (GNPs) have been used in the development of novel therapies as a way of delivery of both stimulatory and tolerogenic peptide cargoes. Here we report that intradermal injection of GNPs loaded with the proinsulin peptide C19-A3, in patients with type 1 diabetes, results in recruitment and retention of immune cells in the skin. These include large numbers of clonally expanded T-cells sharing the same paired T-cell receptors (TCRs) with activated phenotypes, half of which, when the TCRs were re-expressed in a cell-based system, were confirmed to be specific for either GNP or proinsulin.

Using gold nanoparticles for enhanced intradermal delivery of poorly soluble auto-antigenic peptides.

Ultra-small 1-2 nm gold nanoparticles (NP) were conjugated with a poorly-soluble peptide auto-antigen, associated with type 1 diabetes, to modify the peptide pharmacokinetics, following its intradermal delivery. Peptide distribution was characterized, in vivo, after delivery using either conventional intradermal injection or a hollow microneedle device. The poorly-soluble peptide was effectively presented in distant lymph nodes (LN), spleen and draining LN when conjugated to the nanoparticles, whereas peptide alone was only presented in the draining LN.

Noninvasive Molecular Imaging of Mouse Atherosclerosis.

Molecular imaging offers great potential for noninvasive visualization and quantitation of the cellular and molecular components involved in atherosclerotic plaque stability. In this chapter, we review emerging molecular imaging modalities and approaches for quantitative, noninvasive detection of early biological processes in atherogenesis, including vascular endothelial permeability, endothelial adhesion molecule up-regulation, and macrophage accumulation, with special emphasis on mouse models. We also highlight a number of targeted imaging nanomaterials for assessment of advanced atherosclerotic plaques, including extracellular matrix degradation, proteolytic enzyme activity, and activated platelets using mouse models of atherosclerosis.

Endothelial cell-specific reactive oxygen species production increases susceptibility to aortic dissection.

Background: Increased production of reactive oxygen species (ROS) throughout the vascular wall is a feature of cardiovascular disease states, but therapeutic strategies remain limited by our incomplete understanding of the role and contribution of specific vascular cell ROS to disease pathogenesis. To investigate the specific role of endothelial cell (EC) ROS in the development of structural vascular disease, we generated a mouse model of endothelium-specific Nox2 overexpression and tested the susceptibility to aortic dissection after angiotensin II (Ang II) infusion.

Methods And Results: A specific increase in endothelial ROS production in Nox2 transgenic mice was sufficient to cause Ang II-mediated aortic dissection, which was never observed in wild-type mice.

Targeted molecular imaging of vascular inflammation in cardiovascular disease using nano- and micro-sized agents.

Molecular imaging is emerging as a key experimental tool for the identification of inflammatory cellular and molecular processes involved in the development of cardiovascular disease. This review summarises current molecular imaging approaches for the detection of vascular inflammation using a range of nano- and micro-sized contrast agents. We highlight strategies for detection of cell adhesion molecules, which are key regulators of endothelial activation and leukocyte recruitment in atherogenesis and ischaemia-reperfusion in jury.

A leukocyte-mimetic magnetic resonance imaging contrast agent homes rapidly to activated endothelium and tracks with atherosclerotic lesion macrophage content.

Objective: Endothelial cell activation is an important mediator of monocyte recruitment to sites of vascular inflammation. We hypothesized that high-affinity dual-ligand microparticles of iron oxide (MPIO), targeted to P-selectin and vascular cell adhesion molecule-1 (PV-MPIO), would identify activated endothelial cells during atherosclerosis progression.

Methods And Results: In vivo magnetic resonance imaging in apolipoprotein E-deficient mice showed rapid binding of PV-MPIO to the aortic root, which was maximal 30 minutes post-MPIO injection and maintained at 60 minutes.

Molecular MRI enables early and sensitive detection of brain metastases.

Metastasis to the brain is a leading cause of cancer mortality. The current diagnostic method of gadolinium-enhanced MRI is sensitive only to larger tumors, when therapeutic options are limited. Earlier detection of brain metastases is critical for improved treatment.

Development and application of endothelium-targeted microparticles for molecular magnetic resonance imaging.

Molecular imaging of disease states can enhance diagnosis allowing for accurate and more effective treatment. By specifically targeting molecules differentially expressed in disease states, researchers and clinicians have a means of disease characterization at a cellular or tissue level. Targeted micron-sized particles of iron oxide (MPIO) have been used as molecule-specific contrast agents for use with magnetic resonance imaging (MRI), and early evidence suggests they may be suitable for use with other imaging modalities.

VCAM-1-targeted magnetic resonance imaging reveals subclinical disease in a mouse model of multiple sclerosis.

Diagnosis of multiple sclerosis (MS) currently requires lesion identification by gadolinium (Gd)-enhanced or T(2)-weighted magnetic resonance imaging (MRI). However, these methods only identify late-stage pathology associated with blood-brain barrier breakdown. There is a growing belief that more widespread, but currently undetectable, pathology is present in the MS brain.

Molecular imaging with optical coherence tomography using ligand-conjugated microparticles that detect activated endothelial cells: rational design through target quantification.

Objectives: Optical coherence tomography (OCT) is a high resolution imaging technique used to assess superficial atherosclerotic plaque morphology. Utility of OCT may be enhanced by contrast agents targeting molecular mediators of inflammation.

Methods And Results: Microparticles of iron oxide (MPIO; 1 and 4.

Molecular MRI approaches to the detection of CNS inflammation.

Inflammation is a key component of many neurological diseases, yet our understanding of the contribution of these processes to tissue damage remains poor. For many such diseases, magnetic resonance imaging (MRI) has become the method of choice for clinical diagnosis. However, many of the MRI parameters that enable disease detection, such as passive contrast enhancement across a compromised blood-brain barrier, are weighted towards late-stage disease.

Magnetic resonance imaging of brain inflammation using microparticles of iron oxide.

For molecular magnetic resonance imaging (mMRI), microparticles of iron oxide (MPIO) create potent hypointense contrast effects that extend a distance far exceeding their physical size. The potency of the contrast effects derive from their high iron content and are significantly greater than that of ultra-small particles of iron oxide (USPIO), commonly used for MRI. Due to their size and incompressible nature, MPIO are less susceptible to nonspecific vascular egress or uptake by endothelial cells.

Detection of brain pathology by magnetic resonance imaging of iron oxide micro-particles.

Contrast agents are widely used with magnetic resonance imaging (MRI) to increase the contrast between regions of interest and the background signal, thus providing better quality information. Such agents can work in one of two ways, either to specifically enhance the signal that is produced or to localize in a specific cell type of tissue. Commonly used image contrast agents are typically based on gadolinium complexes or super-paramagnetic iron oxide, the latter of which is used for imaging lymph nodes.

In vivo quantification of VCAM-1 expression in renal ischemia reperfusion injury using non-invasive magnetic resonance molecular imaging.

Rationale And Objective: Vascular cell adhesion molecule-1 (VCAM-1) is upregulated in ischemia reperfusion injury (IRI), persisting after restoration of blood flow. We hypothesized that microparticles of iron oxide targeting VCAM-1 (VCAM-MPIO) would depict "ischemic memory" and enable in vivo assessment of VCAM-1 expression.

Methodology And Findings: Mice subject to unilateral, transient (30 minutes) renal ischemia and subsequent reperfusion received intravenous VCAM-MPIO (4.

Molecular magnetic resonance imaging of acute vascular cell adhesion molecule-1 expression in a mouse model of cerebral ischemia.

The pathogenesis of stroke is multifactorial, and inflammation is thought to have a critical function in lesion progression at early time points. Detection of inflammatory processes associated with cerebral ischemia would be greatly beneficial in both designing individual therapeutic strategies and monitoring outcome. We have recently developed a new approach to imaging components of the inflammatory response, namely endovascular adhesion molecule expression on the brain endothelium.

An approach to molecular imaging of atherosclerosis, thrombosis, and vascular inflammation using microparticles of iron oxide.

The rapidly evolving field of molecular imaging promises important advances in the diagnosis, characterization and pharmacological treatment of vascular disease. Magnetic resonance imaging (MRI) provides a modality that is well suited to vascular imaging as it can provide anatomical, structural and functional data on the arterial wall. Its capabilities are further enhanced by the use of a range of increasingly sophisticated contrast agents that target specific molecules, cells and biological processes.

Chapter 4 - Applications of nanotechnology in molecular imaging of the brain.

Rapid advances in the field of nanotechnology promise revolutionary improvements in the diagnosis and therapy of neuroinflammatory disorders. An array of iron oxide nano- and microparticle agents have been developed for in vivo molecular magnetic resonance imaging (mMRI) of cerebrovascular endothelial targets, such as vascular cell adhesion molecule-1 (VCAM-1), E-selectin, and the glycoprotein receptor GP IIb/IIIa expressed on activated platelets. Molecular markers of glioma cells, such as matrix metalloproteinase-2 (MMP-2), and markers for brain tumor angiogenesis, such as alpha (v) beta (3) integrin (alpha(v)beta(3)), have also been successfully targeted using nanoparticle imaging probes.

Visualization of activated platelets by targeted magnetic resonance imaging utilizing conformation-specific antibodies against glycoprotein IIb/IIIa.

Ruptured atherosclerotic plaques, lined with activated platelets, constitute an attractive target for magnetic resonance imaging (MRI). This study evaluated whether microparticles of iron oxide (MPIO) targeting ligand-induced binding sites (LIBS) on the activated conformation of glycoprotein IIb/IIIa could be used to image platelets. MPIO (size: 1 microm) were conjugated to anti-LIBS or control single-chain antibody.

Magnetic resonance imaging of endothelial adhesion molecules in mouse atherosclerosis using dual-targeted microparticles of iron oxide.

Objective: Microparticles of iron oxide (MPIO) distort magnetic field creating marked contrast effects far exceeding their physical size. We hypothesized that antibody-conjugated MPIO would enable magnetic resonance imaging (MRI) of endothelial cell adhesion molecules in mouse atherosclerosis.

Methods And Results: MPIO (4.

In vivo magnetic resonance imaging of acute brain inflammation using microparticles of iron oxide.

Multiple sclerosis is a disease of the central nervous system that is associated with leukocyte recruitment and subsequent inflammation, demyelination and axonal loss. Endothelial vascular cell adhesion molecule-1 (VCAM-1) and its ligand, alpha4beta1 integrin, are key mediators of leukocyte recruitment, and selective inhibitors that bind to the alpha4 subunit of alpha4beta1 substantially reduce clinical relapse in multiple sclerosis. Urgently needed is a molecular imaging technique to accelerate diagnosis, to quantify disease activity and to guide specific therapy.

High-resolution, multicontrast three-dimensional-MRI characterizes atherosclerotic plaque composition in ApoE-/- mice ex vivo.

Purpose: To systematically investigate intrinsic MR contrast mechanisms that would facilitate plaque characterization and quantification in the aortic root and brachiocephalic artery of ApoE-/- mice ex vivo.

Materials And Methods: To establish unambiguous MR parameters for routinely analyzing atherosclerotic plaque ex vivo at 11.7 T, relaxation times of plaque components were quantitatively assessed.

Quantification and 3D reconstruction of atherosclerotic plaque components in apolipoprotein E knockout mice using ex vivo high-resolution MRI.

Objective: To investigate the ability of high-resolution MRI to determine composition and microanatomy of atherosclerosis in mouse aortic root and brachiocephalic artery.

Methods And Results: Aortic root and brachiocephalic arteries of apolipoprotein E knockout (apoE-/-) mice fed Western diet for 10, 20, or 30 weeks were imaged ex vivo (11.7 T; 3D multiecho sequence; resolution 47x47x62.

Increased endothelial tetrahydrobiopterin synthesis by targeted transgenic GTP-cyclohydrolase I overexpression reduces endothelial dysfunction and atherosclerosis in ApoE-knockout mice.

Objective: Increased production of reactive oxygen species and loss of endothelial nitric oxide (NO) bioactivity are key features of vascular disease states such as atherosclerosis. Tetrahydrobiopterin (BH4) is a required cofactor for NO synthesis by endothelial nitric oxide synthase (eNOS); pharmacologic studies suggest that reduced BH4 availability may be an important mediator of endothelial dysfunction in atherosclerosis. We aimed to investigate the importance of endothelial BH4 availability in atherosclerosis using a transgenic mouse model with endothelial-targeted overexpression of the rate-limiting enzyme in BH4 synthesis, GTP-cyclohydrolase I (GTPCH).

Dietary cholesterol reduces lipoprotein lipase activity in the atherosclerosis-susceptible Bio F(1)B hamster.

We have compared lipoprotein metabolism in, and susceptibility to atherosclerosis of, two strains of male Golden Syrian hamster, the Bio F(1)B hybrid and the dominant spot normal inbred (DSNI) strain. When fed a normal low-fat diet containing approximately 40 g fat and 0.3 g cholesterol/kg, triacylglycerol-rich lipoprotein (chylomicron+VLDL) and HDL-cholesterol were significantly higher (P<0.