Georgia Schäfer

The Immunomodulation and Anti-Inflammatory Effects of Garlic Organosulfur Compounds in Cancer Chemoprevention

Garlic (Allium sativum) has been used for centuries as a prophylactic and therapeutic medicinal agent. Importantly, garlic has been suggested to have both cancer-preventive potential as well as significant enhancing effects on the immune system. While these observations are supported experimentally both in vitro and in vivo, the impact of garlic in assisting the immune system in the prevention of cancer still lacks experimental confirmation. Studies addressing the immunomodulatory effects of garlic reveal conflicting data as to pro- or anti-inflammatory responses depending on the particular experimental set-ups and the garlic preparation used (i.e. garlic extract versus chemically pure garlic compounds). Here we provide an overview of the chemistry of the major garlic organosulfur compounds, summarize the current understanding and propose a link between the immunomodulating activity of garlic and the prevention of cancer. We hypothesize that garlic rather elicits anti-inflammatory and anti-oxidative responses that aid in priming the organism towards eradication of an emerging tumor.

The C-type Lectin Receptor CLECSF8 (CLEC4D) Is Expressed by Myeloid Cells and Triggers Cellular Activation through Syk Kinase

Background: C-type lectins play important roles in immunity and homeostasis. Results: CLECSF8 is expressed on neutrophils and monocytes and can mediate phagocytosis, the respiratory burst and inflammatory cytokine production, in part through association with a novel adaptor. Conclusion: CLECSF8 can trigger cellular activation. Significance: This study identifies a novel C-type lectin that can control immune cell function. CLECSF8 is a poorly characterized member of the “Dectin-2 cluster” of C-type lectin receptors and was originally thought to be expressed exclusively by macrophages. We show here that CLECSF8 is primarily expressed by peripheral blood neutrophils and monocytes and weakly by several subsets of peripheral blood dendritic cells. However, expression of this receptor is lost upon in vitro differentiation of monocytes into dendritic cells or macrophages. Like the other members of the Dectin-2 family, which require association of their transmembrane domains with signaling adaptors for surface expression, CLECSF8 is retained intracellularly when expressed in non-myeloid cells. However, we demonstrate that CLECSF8 does not associate with any known signaling adaptor molecule, including DAP10, DAP12, or the FcRγ chain, and we found that the C-type lectin domain of CLECSF8 was responsible for its intracellular retention. Although CLECSF8 does not contain a signaling motif in its cytoplasmic domain, we show that this receptor is capable of inducing signaling via Syk kinase in myeloid cells and that it can induce phagocytosis, proinflammatory cytokine production, and the respiratory burst. These data therefore indicate that CLECSF8 functions as an activation receptor on myeloid cells and associates with a novel adaptor molecule. Characterization of the CLECSF8-deficient mice and screening for ligands using oligosaccharide microarrays did not provide further insights into the physiological function of this receptor.

Helicobacter pylori stimulates host vascular endothelial growth factor-A (vegf-A) gene expression via MEK/ERK-dependent activation of Sp1 and Sp3

VEGF‐A is a key regulator of inflammatory and tumor‐associated angiogenesis. H. pylori plays a critical role in the pathogenesis of benign and malignant gastric diseases. It has been suggested that H. pylori infection is associated with activation of host angiogenesis, however, underlying mechanisms as well as angiogenic growth factors activated by the bacterium have not yet been identified. Therefore, we investigated the influence of the bacterium on VEGF‐A as a candidate host target gene in vivo and in vitro. We show that H. pylori potently up‐regulates production and release of VEGF‐A protein as well as vegf‐A mRNA levels, and we provide strong evidence that enhanced recruitment of Sp1 and Sp3 transcription factors to two proximal GC‐rich vegf‐A promoter elements mediates H. pylori‐triggered vegf‐A gene expression. In addition, H. pylori infection increased the transactivating capacity of both Sp1 and Sp3, which suggests additional mechanism(s) of vegf‐A gene regulation by the bacterium. Signaling studies identified the MEK>ERK1/‐2 kinase cascade as principal host signaling pathway mediating H. pylori‐stimulated vegf‐A transcription. By identifying H. pylori as potent activator of vegf‐A gene expression and characterization of underlying molecular mechanisms, our results provide novel insights into pathways linking the bacterium to host angiogenesis and may help to develop strategies to influence vegf‐A gene expression in the setting of H. pylori infection.

The role of scavenger receptor B1 in infection with Mycobacterium tuberculosis in a murine model

Background The interaction between Mycobacterium tuberculosis (Mtb) and host cells is complex and far from being understood. The role of the different receptor(s) implicated in the recognition of Mtb in particular remains poorly defined, and those that have been found to have activity in vitro were subsequently shown to be redundant in vivo. Methods and Findings To identify novel receptors involved in the recognition of Mtb, we screened a macrophage cDNA library and identified scavenger receptor B class 1 (SR-B1) as a receptor for mycobacteria. SR-B1 has been well-described as a lipoprotein receptor which mediates both the selective uptake of cholesteryl esters and the efflux of cholesterol, and has also recently been implicated in the recognition of other pathogens. We show here that mycobacteria can bind directly to SR-B1 on transfected cells, and that this interaction could be inhibited in the presence of a specific antibody to SR-B1, serum or LDL. We define a variety of macrophage populations, including alveolar macrophages, that express this receptor, however, no differences in the recognition and response to mycobacteria were observed in macrophages isolated from SR-B1−/− or wild type mice in vitro. Moreover, when wild type and SR-B1−/− animals were infected with a low dose of Mtb (100 CFU/mouse) there were no alterations in survival, bacterial burdens, granuloma formation or cytokine production in the lung. However, significant reduction in the production of TNF, IFNγ, and IL10 were observed in SR-B1−/− mice following infection with a high dose of Mtb (1000 CFU/mouse), which marginally affected the size of inflammatory foci but did not influence bacterial burdens. Deficiency of SR-B1 also had no effect on resistance to disease under conditions of varying dietary cholesterol. We did observe, however, that the presence of high levels of cholesterol in the diet significantly enhanced the bacterial burdens in the lung, but this was independent of SR-B1. Conclusion SR-B1 is involved in mycobacterial recognition, but this receptor plays only a minor role in anti-mycobacterial immunity in vivo. Like many other receptors for these pathogens, the loss of SR-B1 can be functionally compensated for under normal conditions.

Non-Opsonic Recognition of Mycobacterium tuberculosis by Phagocytes

The interactions between Mycobacterium tuberculosis and host phagocytes such as macrophages and dendritic cells are central to both immunity and pathogenesis. Many receptors have been implicated in recognition and binding of M. tuberculosis such as the mannose receptor, dendritic-cell-specific intercellular adhesion molecule-3 grabbing nonintegrin, dectin-1 and complement receptor 3 as well as Toll-like receptors, scavenger receptors and CD14. While in vitro studies have demonstrated clear roles for particular recep- tor(s), in vivo work in receptor-deficient animals often revealed only a minor, or no role, in infection with M. tuberculosis. The initial encounter of phagocytic cells with myco- bacteria appears to be complex and depends on various parameters. It seems likely that infection with M. tuberculosis does not occur via a single receptor-mediated pathway. Rather, multiple receptors play different roles in M. tuberculosis infection, and the overall effect depends on the expression and availability of a particular receptor on a particular cell type and its triggered downstream responses. Moreover, the role of membrane cholesterol for M. tuberculosis interactions with phagocytes adds to the complexity of mycobacterial recognition and response. This review summarizes current knowledge on non-opsonic receptors involved in binding of mycobacteria and discusses the contribution of individual receptors to the recognition process.

The Saccharomyces cerevisiae linker histone Hho1p is essential for chromatin compaction in stationary phase and is displaced by transcription

The importance of core histones in the regulation of DNA function by chromatin is clear. However, little is known about the role of the linker histone. We investigated the role of H1 in Saccharomyces cerevisiae during extensive transcriptional reprogramming in stationary phase. Although the levels of linker histone Hho1p remained constant during growth to semiquiescence, there was a genome-wide increase in binding to chromatin. Hho1p was essential for compaction of chromatin in stationary phase, but not for general transcriptional repression. A clear, genome-wide anticorrelation was seen between the level of bound Hho1p and gene expression. Surprisingly, the rank order of gene activity was maintained even in the absence of Hho1p. Based on these findings, we suggest that linker histone Hho1p has a limited role in transcriptional regulation and that the dynamically exchanging linker histone may be evicted from chromatin by transcriptional activity.

Dynamic reciprocity: the role of annexin A2 in tissue integrity

Interactions between cells and the extracellular matrix are integral to tissue development, remodelling and pathogenesis. This is underlined by bi-directional flow of information signalling, referred to as dynamic reciprocity. Annexin A2 is a complex and multifunctional protein that belongs to a large family of Ca2+-dependent anionic phospholipid and membrane-binding proteins. It has been implicated in diverse cellular processes at the nuclear, cytoplasmic and extracellular compartments including Ca2+-dependent regulation of endocytosis and exocytosis, focal adhesion dynamics, transcription and translation, cell proliferation, oxidative stress and apoptosis. Most of these functions are mediated by the annexin A2-S100A10 heterotetramer (AIIt) via its ability to simultaneously interact with cytoskeletal, membrane and extracellular matrix components, thereby mediating regulatory effects of extracellular matrix adhesion on cell behaviour and vice versa. While Src kinase-mediated phosphorylation of filamentous actin-bound AIIt results in membrane-cytoskeletal remodelling events which control cell polarity, cell morphology and cell migration, AIIt at the cell surface can bind to a number of extracellular matrix proteins and catalyse the activation of serine and cysteine proteases which are important in facilitating tissue remodelling during tissue repair, neoangiogenesis and pathological situations. This review will focus on the role of annexin A2 in regulating tissue integrity through intercellular and cell-extracellular matrix interaction. Annexin A2 is differentially expressed in various tissue types as well as in many pathologies, particularly in several types of cancer. These together suggest that annexin A2 acts as a central player during dynamic reciprocity in tissue homeostasis.

The role of inflammation in HPV infection of the Oesophagus

BackgroundSeveral human cancers are known to be associated with inflammation and/or viral infections. However, the influence of tumour-related inflammation on viral uptake is largely unknown. In this study we used oesophageal squamous cell carcinoma (OSCC) as a model system since this type of cancer is associated with chronic irritation, inflammation and viral infections. Although still debated, the most important viral infection seems to be with Human Papillomavirus (HPV). The present study focused on a possible correlation between inflammation, OSCC development and the influence of HPV infection.MethodsA total of 114 OSCC biopsies and corresponding normal tissue were collected at Groote Schuur Hospital and Tygerberg Hospital, Cape Town (South Africa), that were subjected to RNA and DNA isolation. RNA samples were analysed by quantitative Light Cycler RT-PCR for the expression of selected genes involved in inflammation and infection, while conventional PCR was performed on the DNA samples to assess the presence of integrated viral DNA. Further, an in vitro infection assay using HPV pseudovirions was established to study the influence of inflammation on viral infectivity using selected cell lines.ResultsHPV DNA was found in about 9% of OSCC patients, comprising predominantly the oncogenic type HPV18. The inflammatory markers IL6 and IL8 as well as the potential HPV receptor ITGA6 were significantly elevated while IL12A was downregulated in the tumour tissues. However, none of these genes were expressed in a virus-dependent manner. When inflammation was mimicked with various inflammatory stimulants such as benzo-α-pyrene, lipopolysaccharide and peptidoglycan in oesophageal epithelial cell lines in vitro, HPV18 pseudovirion uptake was enhanced only in the benzo-α-pyrene treated cells. Interestingly, HPV pseudovirion infectivity was independent of the presence of the ITGA6 receptor on the surface of the tested cells.ConclusionThis study showed that although the carcinogen benzo-α-pyrene facilitated HPV pseudovirion uptake into cells in culture, HPV infectivity was independent of inflammation and seems to play only a minor role in oesophageal cancer.

Interaction of Human Tumor Viruses with Host Cell Surface Receptors and Cell Entry

Currently, seven viruses, namely Epstein-Barr virus (EBV), Kaposi’s sarcoma-associated herpes virus (KSHV), high-risk human papillomaviruses (HPVs), Merkel cell polyomavirus (MCPyV), hepatitis B virus (HBV), hepatitis C virus (HCV) and human T cell lymphotropic virus type 1 (HTLV-1), have been described to be consistently associated with different types of human cancer. These oncogenic viruses belong to distinct viral families, display diverse cell tropism and cause different malignancies. A key to their pathogenicity is attachment to the host cell and entry in order to replicate and complete their life cycle. Interaction with the host cell during viral entry is characterized by a sequence of events, involving viral envelope and/or capsid molecules as well as cellular entry factors that are critical in target cell recognition, thereby determining cell tropism. Most oncogenic viruses initially attach to cell surface heparan sulfate proteoglycans, followed by conformational change and transfer of the viral particle to secondary high-affinity cell- and virus-specific receptors. This review summarizes the current knowledge of the host cell surface factors and molecular mechanisms underlying oncogenic virus binding and uptake by their cognate host cell(s) with the aim to provide a concise overview of potential target molecules for prevention and/or treatment of oncogenic virus infection.

Targeting neddylation in cancer therapy

The neddylation conjugation pathway has a pivotal role in mediating ubiquitination of proteins and regulation of numerous biological processes. Dysregulation in the ubiquitination and neddylation pathways is associated with many cancers. Ubiquitination involves covalent attachment of ubiquitin to target proteins, leading to protein degradation by the proteasome system. The activity of the E3-ubiquitin ligase family, cullin-RING ligases, is essential for promoting ubiquitin transfer to the appropriate substrates. Neddylation, a process mediated by the protein NEDD8, is required for conformational changes of cullins, a scaffolding protein situated in the core of cullin-RING ligases, and regulation of E3 ligase activity. In this review, we present a comprehensive discussion of the recent findings on the neddylation pathway and its importance during tumorigenesis. The ramifications regarding the potential therapeutic use of ubiquination and neddylation inhibition are also discussed.