Molecular imaging of inflammation - Current and emerging technologies for diagnosis and treatment

Inflammation is a key factor in multiple diseases including primary immune-mediated inflammatory diseases e.g. rheumatoid arthritis but also, less obviously, in many other common conditions, e.g. cardiovascular disease and diabetes. Together, chronic inflammatory diseases contribute to the majority of global morbidity and mortality. However, our understanding of the underlying processes by which the immune response is activated and sustained is limited by a lack of cellular and molecular information obtained in situ. Molecular imaging is the visualization, detection and quantification of molecules in the body. The ability to reveal information on inflammatory biomarkers, pathways and cells can improve disease diagnosis, guide and monitor therapeutic intervention and identify new targets for research. The optimum molecular imaging modality will possess high sensitivity and high resolution and be capable of non-invasive quantitative imaging of multiple disease biomarkers while maintaining an acceptable safety profile. The mainstays of current clinical imaging are computed tomography (CT), magnetic resonance imaging (MRI), ultrasound (US) and nuclear imaging such as positron emission tomography (PET). However, none of these have yet progressed to routine clinical use in the molecular imaging of inflammation, therefore new approaches are required to meet this goal. This review sets out the respective merits and limitations of both established and emerging imaging modalities as clinically useful molecular imaging tools in addition to potential theranostic applications

Plasmacytoid Dendritic Cells Capture and Cross-Present Viral Antigens from Influenza-Virus Exposed Cells

Among the different subsets of dendritic cells (DC), plasmacytoid dendritic cells (PDC) play a unique role in secreting large amounts of type I interferons upon viral stimulation, but their efficiency as antigen-presenting cells has not been completely characterized. We show here, by flow cytometry, with human primary blood PDC and with a PDC cell line, that PDC display poor endocytic capacity for soluble or cellular antigens when compared to monocyte-derived myeloid DC. However, immature PDC efficiently take up cellular material from live influenza-exposed cells, subsequently mature and cross-present viral antigens very efficiently to specific CD8+ T cells. Therefore, during viral infection PDC not only secrete immunomodulatory cytokines, but also recognize infected cells and function as antigen cross-presenting cells to trigger the anti-viral immune response

Plasmacytoid Dendritic Cells Capture and Cross-Present Viral Antigens from Influenza-Virus Exposed Cells

Among the different subsets of dendritic cells (DC), plasmacytoid dendritic cells (PDC) play a unique role in secreting large amounts of type I interferons upon viral stimulation, but their efficiency as antigen-presenting cells has not been completely characterized. We show here, by flow cytometry, with human primary blood PDC and with a PDC cell line, that PDC display poor endocytic capacity for soluble or cellular antigens when compared to monocyte-derived myeloid DC. However, immature PDC efficiently take up cellular material from live influenza-exposed cells, subsequently mature and cross-present viral antigens very efficiently to specific CD8+ T cells. Therefore, during viral infection PDC not only secrete immunomodulatory cytokines, but also recognize infected cells and function as antigen cross-presenting cells to trigger the anti-viral immune response

Poly(I:C) Enhances the Susceptibility of Leukemic Cells to NK Cell Cytotoxicity and Phagocytosis by DC

α Active specific immunotherapy aims at stimulating the host's immune system to recognize and eradicate malignant cells. The concomitant activation of dendritic cells (DC) and natural killer (NK) cells is an attractive modality for immune-based therapies. Inducing immunogenic cell death to facilitate tumor cell recognition and phagocytosis by neighbouring immune cells is of utmost importance for guiding the outcome of the immune response. We previously reported that acute myeloid leukemic (AML) cells in response to electroporation with the synthetic dsRNA analogue poly(I:C) exert improved immunogenicity, demonstrated by enhanced DC-activating and NK cell interferon-γ-inducing capacities. To further invigorate the potential of these immunogenic tumor cells, we explored their effect on the phagocytic and cytotoxic capacity of DC and NK cells, respectively. Using single-cell analysis, we assessed these functionalities in two- and three-party cocultures. Following poly(I:C) electroporation AML cells become highly susceptible to NK cell-mediated killing and phagocytosis by DC. Moreover, the enhanced killing and the improved uptake are strongly correlated. Interestingly, tumor cell killing, but not phagocytosis, is further enhanced in three-party cocultures provided that these tumor cells were upfront electroporated with poly(I:C). Altogether, poly(I:C)-electroporated AML cells potently activate DC and NK cell functions and stimulate NK-DC cross-talk in terms of tumor cell killing. These data strongly support the use of poly(I:C) as a cancer vaccine component, providing a way to overcome immune evasion by leukemic cells

Mir-27b as Biomarker and Regulator of IL-6R Pathway in Resistant Rheumatoid Arthritis Monocyte

Background/Purpose: Whereas molecular mechanisms mediating treatment responses to biologic DMARDS in Rheumatoid Arthritis (RA) are emerging, those pathways that subserve treatment resistance are poorly explored. MicroRNAs (miRs) post-transcriptionally regulate different elements of given signal pathways, and as such capture a ‘snapshot’ of the cell state. We previously assembled a cross-sectional cohort of RA patients with distinct therapy response characteristics including rapid sustained response to conventional (c)DMARD, cDMARD failure and recurrent DMARD (conventional and biologic) resistance. We sought herein to identify dysregulated miR species in therapy resistant patients. <br> Methods: Peripheral blood (PB) CD14+ monocytes were isolated from three groups of RA patients meeting ACR 2010 criteria: good responders to (n30). Comparative samples from age-matched healthy volunteers (n23) were obtained. Candidate miR profiling (based on cytokine receptor targeting miRs) was performed using real-time quantitative PCR. Potential miR-27a/b targets were identified using TargetScan with transcriptional pathway analysis data from PB RA CD14+ cells and functionally validated by Luciferase reporter assay. THP-1 cells were transfected with control or miRNA-27a/b antagomiR and membrane IL-6R expression evaluated by flow cytometry.</br> <br>Results: We identified that miR-27b was significantly down-regulated in sorted CD14+ monocytes obtained from cDMARD failure and biologic treatment resistant, compared to cells from patients responding well to cDMARD and HV (p<0.03). Elevated levels of serum IL-6 were evident in both therapy resistant groups (p+lt;0.001). TargetScan identified several potential miR-27 targets in the IL-6R signal pathway, including soluble and membrane bound variants of IL-6 receptor (IL-6R). Both IL-6R isoforms were confirmed as direct targets of miR-27a/b by luciferase reporter assay. Inhibition of endogenous miR-27a/b via antagomiR in THP-1 cells (myeloid line) significantly increased surface levels of Il-6R (p=0.05, n=4), demonstrating that miR-27a/b actively regulates membrane IL-6R expression.</br> Conclusion: Together our data uncover a previously unrecognized miR-27:IL-6R pathway interaction that operates particularly in monocytes of RA patients with active or treatment refractory disease. Our study thereby identifies a potential new biomarker and functional mechanism associated with disease chronicity

Expression of IL-21 receptor in synovial tissue and blood of patients with rheumatoid arthritis

Background and objectives: Cytokines regulate a broad range of inflammatory pathways in the pathogenesis of Rheumatoid Arthritis (RA), and cytokine blockade against tumour necrosis factor α and IL-6 has offered substantial advances in the treatment of articular inflammation. However, a large proportion of patients will not respond or exhibit only a partial response to treatment and new therapies are thus required. IL-21 is a member of the four-α-helix bundle family of cytokines that mediates pleiotropic effects through the IL-21 receptor (IL-21R). Since potency of IL-21 is mainly dependent on presence and abundance of its receptor on different cells types, the objective of this study was to characterise expression of IL-21R in the synovium and blood of patients with RA. <p/>Materials and methods: Immunohistochemistry for IL-21R was carried out on synovial tissue samples derived by arthroplasty from patients with RA (n=5) obtained from the Institute of Infection, Immunity and Inflammation Research Tissue Bank. Mononuclear cells were separated out from peripheral blood (PBMC) of 10 RA patients or 3 healthy controls on a density gradient using Histopaque (Sigma) and analysed by flow cytometry for IL-21R expression on T cells (CD3/CD4/CD8), B cells (CD19/CD27) and NK cells (CD16/CD56). <p/>Results: Expression of IL-21R was detected in 5/5 synovial RA tissues. The IL-21R+ cells were located in the synovial intimal and sublining layers and in lymphoid aggregates. Flow cytometric analysis on blood PBMC revealed that IL-21R is highly expressed on both CD4+ (73.04%, 12.58 MFI) and CD8+ (50.88%, 13.69 MFI) T cells, as well as on a proportion of NK cells (73.83%, 19.15 MFI) in RA patients. On B cells, IL-21R expression was higher on the CD27- fraction of naïve B cells (95.19%, 37.02 MFI), with lower expression on the CD27+ memory B cells (15.2%, 32.22 MFI). <p/>Conclusions: Our results show increased expression of IL-21R in established RA synovial tissue and peripheral blood, and indicate that targeting of the IL-21/IL-21R pathway may be a valid therapeutic strategy for the treatment of RA