Inhibition of Inflammatory Arthritis Using Fullerene Nanomaterials

Inflammatory arthritis (e.g. rheumatoid arthritis; RA) is a complex disease driven by the interplay of multiple cellular lineages. Fullerene derivatives have previously been shown to have anti-inflammatory capabilities mediated, in part, by their ability to prevent inflammatory mediator release by mast cells (MC). Recognizing that MC can serve as a cellular link between autoantibodies, soluble mediators, and other effector populations in inflammatory arthritis, it was hypothesized that fullerene derivatives might be used to target this inflammatory disease. A panel of fullerene derivatives was tested for their ability to affect the function of human skin-derived MC as well as other lineages implicated in arthritis, synovial fibroblasts and osteoclasts. It is shown that certain fullerene derivatives blocked FcγR- and TNF-α-induced mediator release from MC; TNF-α-induced mediator release from RA synovial fibroblasts; and maturation of human osteoclasts. MC inhibition by fullerene derivatives was mediated through the reduction of mitochondrial membrane potential and FcγR-mediated increases in cellular reactive oxygen species and NF-κB activation. Based on these in vitro data, two fullerene derivatives (ALM and TGA) were selected for in vivo studies using K/BxN serum transfer arthritis in C57BL/6 mice and collagen-induced arthritis (CIA) in DBA/1 mice. Dye-conjugated fullerenes confirmed localization to affected joints in arthritic animals but not in healthy controls. In the K/BxN moldel, fullerenes attenuated arthritis, an effect accompanied by reduced histologic inflammation, cartilage/bone erosion, and serum levels of TNF-α. Fullerenes remained capable of attenuating K/BxN arthritis in mast cell-deficient mice Cre-Master mice, suggesting that lineages beyond the MC represent relevant targets in this system. These studies suggest that fullerene derivatives may hold promise both as an assessment tool and as anti-inflammatory therapy of arthritis

Human Tumor Targeted Cytotoxic Mast Cells for Cancer Immunotherapy

The diversity of autologous cells being used and investigated for cancer therapy continues to increase. Mast cells (MCs) are tissue cells that contain a unique set of anti-cancer mediators and are found in and around tumors. We sought to exploit the anti-tumor mediators in MC granules to selectively target them to tumor cells using tumor specific immunoglobin E (IgE) and controllably trigger release of anti-tumor mediators upon tumor cell engagement. We used a human HER2/neu-specific IgE to arm human MCs through the high affinity IgE receptor (FcεRI). The ability of MCs to bind to and induce apoptosis of HER2/neu-positive cancer cells in vitro and in vivo was assessed. The interactions between MCs and cancer cells were investigated in real time using confocal microscopy. The mechanism of action using cytotoxic MCs was examined using gene array profiling. Genetically manipulating autologous MC to assess the effects of MC-specific mediators have on apoptosis of tumor cells was developed using siRNA. We found that HER2/neu tumor-specific IgE-sensitized MCs bound, penetrated, and killed HER2/neu-positive tumor masses in vitro. Tunneling nanotubes formed between MCs and tumor cells are described that parallel tumor cell apoptosis. In solid tumor, human breast cancer (BC) xenograft mouse models, infusion of HER2/neu IgE-sensitized human MCs co-localized to BC cells, decreased tumor burden, and prolonged overall survival without indications of toxicity. Gene microarray of tumor cells suggests a dependence on TNF and TGFβ signaling pathways leading to apoptosis. Knocking down MC-released tryptase did not affect apoptosis of cancer cells. These studies suggest MCs can be polarized from Type I hypersensitivity-mediating cells to cytotoxic cells that selectively target tumor cells and specifically triggered to release anti-tumor mediators. A strategy to investigate which MC mediators are responsible for the observed tumor killing is described so that rational decisions can be made in the future when selecting which mediators to target for deletion or those that could further polarize them to cytotoxic MC by adding other known anti-tumor agents. Using autologous human MC may provide further options for cancer therapeutics that offers a unique anti-cancer mechanism of action using tumor targeted IgE’s

Functionalization of gadolinium metallofullerenes for detecting atherosclerotic plaque lesions by cardiovascular magnetic resonance

BACKGROUND: The hallmark of atherosclerosis is the accumulation of plaque in vessel walls. This process is initiated when monocytic cells differentiate into macrophage foam cells under conditions with high levels of atherogenic lipoproteins. Vulnerable plaque can dislodge, enter the blood stream, and result in acute myocardial infarction and stroke. Imaging techniques such as cardiovascular magnetic resonance (CMR) provides one strategy to identify patients with plaque accumulation. METHODS: We synthesized an atherosclerotic-targeting contrast agent (ATCA) in which gadolinium (Gd)-containing endohedrals were functionalized and formulated into liposomes with CD36 ligands intercalated into the lipid bilayer. In vitro assays were used to assess the specificity of the ATCA for foam cells. The ability of ATCA to detect atherosclerotic plaque lesions in vivo was assessed using CMR. RESULTS: The ATCA was able to detect scavenger receptor (CD36)-expressing foam cells in vitro and were specifically internalized via the CD36 receptor as determined by focused ion beam/scanning electron microscopy (FIB-SEM) and Western blotting analysis of CD36 receptor-specific signaling pathways. The ATCA exhibited time-dependent accumulation in atherosclerotic plaque lesions of ApoE −/− mice as determined using CMR. No ATCA accumulation was observed in vessels of wild type (C57/b6) controls. Non-targeted control compounds, without the plaque-targeting moieties, were not taken up by foam cells in vitro and did not bind plaque in vivo. Importantly, the ATCA injection was well tolerated, did not demonstrate toxicity in vitro or in vivo, and no accumulation was observed in the major organs. CONCLUSIONS: The ATCA is specifically internalized by CD36 receptors on atherosclerotic plaque providing enhanced visualization of lesions under physiological conditions. These ATCA may provide new tools for physicians to non-invasively detect atherosclerotic disease

Human Mast Cells From Adipose Tissue Target and Induce Apoptosis of Breast Cancer Cells

Mast cells (MC) are important immune sentinels found in most tissue and widely recognized for their role as mediators of Type I hypersensitivity. However, they also secrete anti-cancer mediators such as tumor necrosis factor alpha (TNF-α) and granulocyte-macrophage colony-stimulating factor (GM-CSF). The purpose of this study was to investigate adipose tissue as a new source of MC in quantities that could be used to study MC biology focusing on their ability to bind to and kill breast cancer cells. We tested several cell culture media previously demonstrated to induce MC differentiation. We report here the generation of functional human MC from adipose tissue. The adipose-derived mast cells (ADMC) are phenotypically and functionally similar to connective tissue expressing tryptase, chymase, c-kit, and FcεRI and capable of degranulating after cross-linking of FcεRI. The ADMC, sensitized with anti-HER2/neu IgE antibodies with human constant regions (trastuzumab IgE and/or C6MH3-B1 IgE), bound to and released MC mediators when incubated with HER2/neu-positive human breast cancer cells (SK-BR-3 and BT-474). Importantly, the HER2/neu IgE-sensitized ADMC induced breast cancer cell (SK-BR-3) death through apoptosis. Breast cancer cell apoptosis was observed after the addition of cell-free supernatants containing mediators released from FcεRI-challenged ADMC. Apoptosis was significantly reduced when TNF-α blocking antibodies were added to the media. Adipose tissue represents a source MC that could be used for multiple research purposes and potentially as a cell-mediated cancer immunotherapy through the expansion of autologous (or allogeneic) MC that can be targeted to tumors through IgE antibodies recognizing tumor specific antigens

A prospective registry of emergency department patients admitted with infection

<p>Abstract</p> <p>Background</p> <p>Patients with infections account for a significant proportion of Emergency Department (ED) workload, with many hospital patients admitted with severe sepsis initially investigated and resuscitated in the ED. The aim of this registry is to systematically collect quality observational clinical and microbiological data regarding emergency patients admitted with infection, in order to explore in detail the microbiological profile of these patients, and to provide the foundation for a significant programme of prospective observational studies and further clinical research.</p> <p>Methods/design</p> <p>ED patients admitted with infection will be identified through daily review of the computerised database of ED admissions, and clinical information such as site of infection, physiological status in the ED, and components of management abstracted from patients' charts. This information will be supplemented by further data regarding results of investigations, microbiological isolates, and length of stay (LOS) from hospital electronic databases. Outcome measures will be hospital and intensive care unit (ICU) LOS, and mortality endpoints derived from a national death registry.</p> <p>Discussion</p> <p>This database will provide substantial insights into the characteristics, microbiological profile, and outcomes of emergency patients admitted with infections. It will become the nidus for a programme of research into compliance with evidence-based guidelines, optimisation of empiric antimicrobial regimens, validation of clinical decision rules and identification of outcome determinants. The detailed observational data obtained will provide a solid baseline to inform the design of further controlled trials planned to optimise treatment and outcomes for emergency patients admitted with infections.</p

The Role of Horseshoe Crabs in the Biomedical Industry and Recent Trends Impacting Species Sustainability

Every year the Atlantic horseshoe crab (Limulus polyphemus) arrives on shore to spawn, a sight once taken for granted. However, in addition to the gradual climate changes impacting all ecosystems, commercial demand from the widespread application of Atlantic horseshoe crab blood in industrial endotoxin testing and steady use as eel and whelk bait has brought the future of this enduring species into question. In response, regulations have been adopted to enhance the traceability and record keeping of horseshoe crab harvest, which has historically been difficult to track. However, these regulations do not restrict or limit LAL harvest in any significant manner. Still, sometimes-lethal biomedical bleeding process and associated behavioral changes pose a risk to horseshoe crab viability after bleeding and once returned to the waters. As a result, regulators and environmentalists are concerned that current trends and overfishing of this marine arthropod will significantly impact the surrounding ecosystem. This review examines their role and recent trends in the biomedical industry that are impacting these ancient creatures and the derivative effect on shorebirds, while considering emerging alternatives where feasible, as well as ways to ensure sustainable and pragmatic harvesting strategies. Ultimately, healthy populations of horseshoe crabs are vital to restoring and maintaining ecosystems while balancing the need for medical and research applications entirely dependent on these unique creatures