Molecularly Imprinted Nanobeacons Redirect Innate Immune Killing towards Triple Negative Breast Cancer

Harnessing innate immunity is an appealing strategy for cancer treatment. Herein, we report a new strategy called molecularly imprinted nanobeacons (MINBs) for redirecting innate immune killing towards triple-negative breast cancer (TNBC). The MINBs were molecularly imprinted nanoparticles with the N-epitope of glycoprotein nonmetastatic B (GPNMB) as the template and grafted with plentiful fluorescein moieties as the hapten. The MINBs could tag the TNBC cells via binding with GPNMB and thereby provide navigation for recruiting hapten-specific antibodies. The gathered antibodies could further trigger effective Fc-domain-mediated immune killing towards the tagged cancer cells. In vivo experiments showed that the TNBC growth was significantly inhibited after MINBs treatment by intravenous injection as compared with control groups. This study not only opens a new access for redirecting innate immunity towards TNBC but also paves the way for innate immunity-based therapy of other diseases.     

Structures of the dimeric and monomeric chromanones, gonytolides A-C, isolated from the fungus Gonytrichum sp. and their promoting activities of innate immune responses

Innate immunity is the front line of self-defense against microbial infection. After searching for natural substances that regulate innate immunity using an ex vivo Drosophila culture system, we identified a novel dimeric chromanone, gonytolide A, as an innate immune promoter from the fungus Gonytrichum sp. along with gonytolides B and C. Gonytolide A also increased TNF-α-stimulated production of IL-8 in human umbilical vein endothelial cells.     

Human and Limulus Blood: Opportunity for Development of A Medical Biotechnology Project in Taiwan

The function of blood is to deliver nourishments to and remove wastes from all parts of the body. It is made up of different kinds of cells bathed in a fluid called plasma. The major cellular components of blood include (1) red blood cells for carrying oxygen to the various tissues, (2) white blood cells for providing defense against infectious agents and cancer, and (3) platelets for inducing a cascade of events leading to blood clot formation that stops bleeding. The plasma also contains numerous proteins for maintaining normal balance in our body, and include (1) clotting factors such as factor VIII, factor IX, fibrinogen, and thrombin, (2) protease inhibitors and anticoagulants that regulate the coagulation pathway, the complement system, or the fibrinolytic system, (3) immunoglobulin which are antibodies directed against different infectious agents, and (4) albumin which functions as a volume expander to maintain our blood volume and is essential when threatened with severe blood loss. The plasma of the invertebrate horseshoe crab, contains three major proteins: hemocyanin, C-reactive protein (CRP), and α₂-macroglobulin. Hemocyanin functions as an oxygen-carrying protein. CRP is a lectin that binds to phosphocholine of the pneumococcus C-polysaccharide, to the chromatins of damaged cells, and to the galactose moiety of desialylated glycoprotein as a membrane-associated protein on liver marcrophages. CRP exists, however, as a normal component of the invertebrate hemolymph. α₂-macroglobulin exhibits proteinase inhibitory activity with a broad specificity that can block the activities of protease secreted from invading microorganisms. The Limulus CRP, along with the C3 homologue α₂-macroglobulin, participates in a complement - like hemolytic activity in horseshoe crab hemolymph. Whereas the vertebrate evolved to use both the innate and the adaptive immunity, the invertebrate only uses the innate immunity. The innate immunity uses germ-line encoded receptors to recognize conserved molecular constituents of infectious microorganisms, is phylogenetically older, with some of its form presumably presents in all multicellular organisms. The adaptive immunity is mediated by highly specific antigen receptors that are distributed clonally on the two types of lymphocytes, the T-cells and the B-cells. Evidence has accumulated in recent years to suggest that the innate immune system provides signals that are essential for the adaptive immune response to generate information on the origin of the antigen and the type of response to be induced. This linkage invites renewed interest in the study of the innate immune system of the horseshoe crab.     

New era for mucosal mast cells: their roles in inflammation,allergic immune responses and adjuvant development

To achieve immune homeostasis in such a harsh environment as the intestinal mucosa, both active and quiescent immunity operate simultaneously. Disruption of gut immune homeostasis leads to the development of intestinal immune diseases such as colitis and food allergies. Among various intestinal innate immune cells, mast cells (MCs) play critical roles in protective immunity against pathogenic microorganisms, especially at mucosal sites. This suggests the potential for a novel MC-targeting type of vaccine adjuvant. Dysregulated activation of MCs also results in inflammatory responses in mucosal compartments. The regulation of this yin and yang function of MCs remains to be elucidated. In this review, we focus on the roles of mucosal MCs in the regulation of intestinal allergic reaction, inflammation and their potential as a new target for the development of mucosal adjuvants.     

Immunomodulatory functional foods and their molecular mechanisms

The immune system comprises a complex group of processes that provide defense against diverse pathogens. These defenses can be divided into innate and adaptive immunity, in which specific immune components converge to limit infections. In addition to genetic factors, aging, lifestyle, and environmental factors can influence immune function, potentially affecting the susceptibility of the host to disease-causing agents. Chemical compounds in certain foods have been shown to regulate signal transduction and cell phenotypes, ultimately impacting pathophysiology. Research has shown that the consumption of specific functional foods can stimulate the activity of immune cells, providing protection against cancer, viruses, and bacteria. Here, we review a number of functional foods reported to strengthen immunity, including ginseng, mushrooms, chlorella, and probiotics (Lactobacillus plantarum). We also discuss the molecular mechanisms involved in regulating the activity of various types of immune cells. Identifying immune-enhancing functional foods and understanding their mechanisms of action will support new approaches to maintain proper health and combat immunological diseases.Subject terms: Health care, Pathogenesis     

Innate Immunity Modulating Impurities and the Immunotoxicity of Nanobiotechnology-Based Drug Products

Innate immunity can be triggered by the presence of microbial antigens and other contaminants inadvertently introduced during the manufacture and purification of bionanopharmaceutical products. Activation of these innate immune responses, including cytokine secretion, complement, and immune cell activation, can result in unexpected and undesirable host immune responses. These innate modulators can also potentially stimulate the activation of adaptive immune responses, including the formation of anti-drug antibodies which can impact drug effectiveness. To prevent induction of these adverse responses, it is important to detect and quantify levels of these innate immunity modulating impurities (IIMIs) that may be present in drug products. However, while it is universally agreed that removal of IIMIs from drug products is crucial for patient safety and to prevent long-term immunogenicity, there is no single assay capable of directly detecting all potential IIMIs or indirectly quantifying downstream biomarkers. Additionally, there is a lack of agreement as to which of the many analytical assays currently employed should be standardized for general IIMI screening. Herein, we review the available literature to highlight cellular and molecular mechanisms underlying IIMI-mediated inflammation and its relevance to the safety and efficacy of pharmaceutical products. We further discuss methodologies used for direct and indirect IIMI identification and quantification.     

A re-evaluation of the role of host defence peptides in mammalian immunity

Host defence peptides are found in all classes of life and are a fundamental component of the innate immune response. Initially it was believed that their sole role in innate immunity was to kill invading microorganisms, thus providing direct defence against infection. Evidence now suggests that these peptides play diverse and complex roles in the immune response and that, in higher animals, their functions are not restricted to the innate immune response. In in vitro experiments certain host defence peptides have been demonstrated to be potent antimicrobial agents at modest concentrations, although their antimicrobial activity is often strongly reduced or ablated in the presence of physiological concentrations of ions such as Na(+) and Mg(2+). In contrast, in experiments done in standard tissue culture media, the composition of which more accurately represents physiological levels of ions, mammalian host defence peptides have been demonstrated to have a number of immunomodulatory functions including altering host gene expression, acting as chemokines and/or inducing chemokine production, inhibiting lipopolysaccharide induced pro-inflammatory cytokine production, promoting wound healing, and modulating the responses of dendritic cells and cells of the adaptive immune response. Animal models indicate that host defence peptides are crucial for both prevention and clearance of infection. As interest in the in vivo functions of host defence peptides is increasing, it is important to consider whether in mammals the direct antimicrobial and immunomodulatory properties observed in vitro are physiologically relevant, especially since many of these activities are concentration dependent. In this review we summarize the concentrations of host defence peptides and ions reported throughout the body and compare that information with the concentrations of peptides that are known have antimicrobial or immunomodulatory functions in vitro.     

Pellino1 promotes chronic inflammatory skin disease via keratinocyte hyperproliferation and induction of the T helper 17 response

Psoriasis is one of the most common immune-mediated chronic inflammatory skin diseases. However, little is known about the molecular mechanism underlying the immunological circuits that maintain innate and adaptive immune responses in established psoriasis. In this study, we found that the Pellino1 (Peli1) ubiquitin E3 ligase is activated by innate pattern-recognition receptors (PRRs), such as Toll-like receptors (TLRs), and is highly upregulated in human psoriatic skin lesions and murine psoriasis-like models. Increased Peli1 expression is strongly correlated with the immunopathogenesis of psoriasis by activating hyperproliferation of keratinocytes in the S and G2/M phases of the cell cycle and promoting chronic skin inflammation. Furthermore, Peli1-induced psoriasis-like lesions showed significant changes in the expression levels of several T helper 17 (Th17)-related cytokines, such as IL-17a, IL-21, IL-22, IL-23, and IL-24, indicating that overexpression of Peli1 resulted in the sequential engagement of the Th17 cell response. However, the overexpression of Peli1 in T cells was insufficient to trigger psoriasis, while T cells were indispensable for disease manifestation. In summary, our findings demonstrate that Peli1 is a critical cell cycle activator of innate immunity, which subsequently links Th17 cell immune responses to the psoriatic microenvironment.Subject terms: Chronic inflammation, Immunoproliferative disorders     

Combination of 5-Fluorouracil with Photodynamic Therapy: Enhancement of Innate and Adaptive Immune Responses in a Murine Model of Actinic Keratosis†

We previously showed that a combination of differentiation-inducing agents (5-fluorouracil [5FU], vitamin D3 or methotrexate) and aminolevulinate-based photodynamic therapy (PDT) improves clinical responses by enhancing protoporphyrin IX (PpIX) photosensitizer levels and cell death. Here, we show that in addition to its previously known effects, 5FU enhances PDT-induced tumor-regressing immunity. Murine actinic keratoses were treated with topical 5FU or vehicle for 3 days prior to aminolevulinic acid application, followed by blue light illumination (~417 nm). Lesions were harvested for time-course analyses of innate immune cell recruitment into lesions, i.e. neutrophils (Ly6G+) and macrophages (F4/80+), which peaked at 72 h and 1 week post-PDT, respectively, and were greater in 5FU-treated lesions. Enhanced infiltration of activated T cells (CD3+) throughout the time course, and of cytotoxic T cells (CD8+) at 1–2 weeks post-PDT, also occurred in 5FU-treated lesions. 5FU pretreatment reduced the presence of cells expressing the immune checkpoint marker PD-1 at ~72 h post-PDT, favoring cytotoxic T cell activity. A combination of 5FU and PDT, each individually known to induce long-term tumor-targeting immune responses in addition to their more immediate effects on cancer cells, may synergize to provide better management of squamous precancers.     

Quantitative Mapping of Endosomal DNA Processing by Single Molecule Counting

Extracellular DNA is engulfed by innate immune cells and digested by endosomal DNase II to generate an immune response. Quantitative information on endosomal stage‐specific cargo processing is a critical parameter to predict and model the innate immune response. Biochemical assays quantify endosomal processing but lack organelle‐specific information, while fluorescence microscopy has provided the latter without the former. Herein, we report a single molecule counting method based on fluorescence imaging that quantitatively maps endosomal processing of cargo DNA in innate immune cells with organelle‐specific resolution. Our studies reveal that endosomal DNA degradation occurs mainly in lysosomes and is negligible in late endosomes. This method can be used to study cargo processing in diverse endocytic pathways and measure stage‐specific activity of processing factors in endosomes.     

Combination of anti-angiogenic therapy and immune checkpoint blockade normalizes vascular-immune crosstalk to potentiate cancer immunity

Cancer immunotherapy with immune checkpoint inhibitors (ICIs) has revolutionized the treatment of advanced cancers. However, the tumor microenvironment (TME) functions as a formidable barrier that severely impairs the efficacy of ICIs. While the crosstalk between tumor vessels and immune cells determines the nature of anti-tumor immunity, it is skewed toward a destructive cycle in growing tumors. First, the disorganized tumor vessels hinder CD8⁺ T cell trafficking into the TME, disable effector functions, and even kill T cells. Moreover, VEGF, the key driver of angiogenesis, interferes with the maturation of dendritic cells, thereby suppressing T cell priming, and VEGF also induces TOX-mediated exhaustion of CD8⁺ T cells. Meanwhile, a variety of innate and adaptive immune cells contribute to the malformation of tumor vessels. Protumoral M2-like macrophages as well as T_H2 and Treg cells secrete pro-angiogenic factors that accelerate uncontrolled angiogenesis and promote vascular immaturity. While CD8⁺ T and CD4⁺ T_H1 cells suppress angiogenesis and induce vascular maturation by secreting IFN-γ, they are unable to infiltrate the TME due to malformed tumor vessels. These findings led to preclinical studies that demonstrated that simultaneous targeting of tumor vessels and immunity is a viable strategy to normalize aberrant vascular-immune crosstalk and potentiate cancer immunotherapy. Furthermore, this combination strategy has been evidently demonstrated through recent pivotal clinical trials, granted approval from FDA, and is now being used in patients with kidney, liver, lung, or uterine cancer. Overall, combining anti-angiogenic therapy and ICI is a valid therapeutic strategy that can enhance cancer immunity and will further expand the landscape of cancer treatment.Subject terms: Cancer immunotherapy, Cancer microenvironment, Tumour angiogenesis, Tumour immunology, Targeted therapies     

Regulation of antiviral innate immune signaling and viral evasion following viral genome sensing

A harmonized balance between positive and negative regulation of pattern recognition receptor (PRR)-initiated immune responses is required to achieve the most favorable outcome for the host. This balance is crucial because it must not only ensure activation of the first line of defense against viral infection but also prevent inappropriate immune activation, which results in autoimmune diseases. Recent studies have shown how signal transduction pathways initiated by PRRs are positively and negatively regulated by diverse modulators to maintain host immune homeostasis. However, viruses have developed strategies to subvert the host antiviral response and establish infection. Viruses have evolved numerous genes encoding immunomodulatory proteins that antagonize the host immune system. This review focuses on the current state of knowledge regarding key host factors that regulate innate immune signaling molecules upon viral infection and discusses evidence showing how specific viral proteins counteract antiviral responses via immunomodulatory strategies.Subject terms: Innate immunity, Post-translational modifications     

Innate immunity probed by lipopolysaccharides affinity strategy and proteomics

Lipopolysaccharides (LPSs) are ubiquitous and vital components of the cell surface of Gram-negative bacteria that have been shown to play a relevant role in the induction of the immune-system response. In animal and plant cells, innate immune defenses toward microorganisms are triggered by the perception of pathogen associated molecular patterns. These are conserved and generally indispensable microbial structures such as LPSs that are fundamental in the Gram-negative immunity recognition. This paper reports the development of an integrated strategy based on lipopolysaccharide affinity methodology that represents a new starting point to elucidate the molecular mechanisms elicited by bacterial LPS and involved in the different steps of innate immunity response. Biotin-tagged LPS was immobilized on streptavidin column and used as a bait in an affinity capture procedure to identify protein partners from human serum specifically interacting with this effector. The complex proteins/lipopolysaccharide was isolated and the protein partners were fractionated by gel electrophoresis and identified by mass spectrometry. This procedure proved to be very effective in specifically binding proteins functionally correlated with the biological role of LPS. Proteins specifically bound to LPS essentially gathered within two functional groups, regulation of the complement system (factor H, C4b, C4BP, and alpha 2 macroglobulin) and inhibition of LPS-induced inflammation (HRG and Apolipoproteins). The reported strategy might have important applications in the elucidation of biological mechanisms involved in the LPSs-mediated molecular recognition and anti-infection responses.     

锌对动物免疫功能的影响

锌是机体必需的微量元素,它对动物免疫系统的正常发育和功能方面有重要作用。综述了微量元素锌对动物免疫器官、免疫细胞和免疫分子等的可能作用机制和对免疫功能的影响。     

Iron chelator induces MIP-alpha/CCL20 in human intestinal epithelial cells: implication for triggering mucosal adaptive immunity

A previous report by this laboratory demonstrated that bacterial iron chelator (siderophore) triggers inflammatory signals, including the production of CXC chemokine IL-8, in human intestinal epithelial cells (IECs). Microarray-based gene expression profiling revealed that iron chelator also induces macrophage inflammatory protein 3 alpha (MIP-3alpha)/CC chemokine-ligand 20 (CCL20). As CCL20 is chemotactic for the cells involved in host adaptive immunity, this suggests that iron chelator may stimulate IECs to have the capacity to link mucosal innate and adaptive immunity. The basal medium from iron chelator deferoxamine (DFO)-treated HT-29 monolayers was as chemotactic as recombinant human CCL20 at equivalent concentrations to attract CCR6(+) cells. The increase of CCL20 protein secretion appeared to correspond to that of CCL20 mRNA levels, as determined by real-time quantitative RT-PCR. The efficacy of DFO at inducing CCL20 mRNA was also observed in human PBMCs and in THP-1 cells, but not in human umbilical vein endothelial cells. Interestingly, unlike other proinflammatory cytokines, such as TNF-alpha and IL-1beta, a time-dependent experiment revealed that DFO slowly induces CCL20, suggesting a novel mechanism of action. A pharmacologic study also revealed that multiple signaling pathways are differentially involved in CCL20 production by DFO, while some of those pathways are not involved in TNF-alpha-induced CCL20 production. Collectively, these results demonstrate that, in addition to some bacterial products known to induce host adaptive immune responses, direct chelation of host iron by infected bacteria may also contribute to the initiation of host adaptive immunity in the intestinal mucosa.     

靶向Toll样受体4的小分子调节剂

Toll样受体(Toll-like receptors,TLRs)是进化保守的天然免疫模式识别受体,能够识别外源的病原菌相关分子模式(Pathogen-associated molecular patterns,PAMPs)、内源的损害相关分子模式(Damage-associated molecular patterns,DAMPs)和异源物相关分子模式(Xenobiotic-associated molecular patterns),诱导炎症免疫反应。 其中,TLR4(Toll-like receptor 4)是目前研究最为广泛的Toll样受体之一,TLR4是脂多糖(lipopoiysaccharide,LPS)的主要受体,LPS激活的TLR4信号通路在炎症信号的传递中发挥着重要作用,而此信号转导需要通过LPS与TLR4及其附属蛋白髓样分化因子2(myeloid differentiation factor 2,MD-2)的相互作用来实现。 因此,TLR4/MD-2成为炎症反应和免疫调控最重要的研究热点。 本文综述靶向TLR4/MD-2的小分子激动剂和抑制剂的研究进展,以进一步理解TLR4小分子调节剂与其相互作用的复杂性,帮助靶向TLR4/MD-2的免疫调节剂药物发现。     

Nickel induces secretion of IFN-�� by splenic natural killer cells

Although nickel hypersensitivity is known as a delayed-type hypersensitivity mediated by nickel-specific T cells, it is greatly influenced by other immune cells. Here we show that splenic natural killer cells (NK cells) directly or indirectly respond to nickel by secretion of IFN-γ. Using enzyme-linked immunosorbent spot (ELISPOT) assays, we found that nickel-reactive cells readily secreted IFN-γ when splenocytes were cultured in the presence of varying concentrations of nickel sulfate (NiSO₄) for 24 h. However, nickel-reactive IL-2- or IL- 4-secreting cells were infrequent during the 24-h culture with NiSO₄. Immune responses to nickel were innate, not adaptive, in nature since the frequency of nickel-reactive IFN-γ-secreting cells did not increase upon previous exposure to NiSO₄ and recombination activating gene (RAG)-1-deficient mice contained nickel-reactive IFN-γ-secreting cells. The involvement of NK cells in the innate response to NiSO₄ was confirmed since we could observe a significant reduction of the frequency of nickel-reactive cells in NK cell-depleted mice. Furthermore, the number of IFN-γ secreting cells was significantly reduced in the ELISPOT assays when NKG2D was blocked by anti-NKG2D antibody. These results suggest that there is an early and rapid innate immune response to nickel, which is mediated by NK cells and the NKG2D receptor. The significance of the innate response to nickel is that it may contribute to development of the late T cell-mediated delayed type hypersensitivity against nickel.     

Chemistry of Lipid A: At the Heart of Innate Immunity

In many Gram‐negative bacteria, lipopolysaccharide (LPS) and its lipid A moiety are pivotal for bacterial survival. Depending on its structure, lipid A carries the toxic properties of the LPS and acts as a potent elicitor of the host innate immune system via the Toll‐like receptor 4/myeloid differentiation factor 2 (TLR4/MD‐2) receptor complex. It often causes a wide variety of biological effects ranging from a remarkable enhancement of the resistance to the infection to an uncontrolled and massive immune response resulting in sepsis and septic shock. Since the bioactivity of lipid A is strongly influenced by its primary structure, a broad range of chemical syntheses of lipid A derivatives have made an enormous contribution to the characterization of lipid A bioactivity, providing novel pharmacological targets for the development of new biomedical therapies. Here, we describe and discuss the chemical aspects regarding lipid A and its role in innate immunity, from the (bio)synthesis, isolation and characterization to the molecular recognition at the atomic level.     

Role of lipid rafts in innate immunity and phagocytosis of polystyrene latex microspheres

Understanding of the association of phagocytosis of polymers with signaling of innate immunity of macrophages is the major purpose of this study. Polymer conjugates have been utilized for clinical therapy of cancer and infections, such as Mycobacterium tuberculosis, as effective vectors of drug-delivery systems. They are incorporated through phagocytosis into macrophages and activate innate immunity signaling, which plays a crucial role in its therapeutic and side effects. Macrophage phagocytosis of polystyrene latex microspheres was examined and assayed by treatment of macrophages with the cholesterol depletor methyl-β-cyclodextrin (MβCD) or the sphingolipid depletor n-octyl-β-D-glucopyranoside (OGP). Expressions of various mRNAs during phagocytosis were quantified by real-time PCR. Phagocytosis of polystyrene latex microspheres by various macrophages, such as murine monocyte-derived macrophage J774, rat alveolar macrophage NR8383, and murine Kupffer cell KC13-2, was suppressed by treatment with MβCD or OGP in a concentration-dependent manner. The expression of mRNAs of TNFα, IL-1β, IL-6 and CXCL10 genes induced by lipopolysaccharide (LPS) was not suppressed by treatment with MβCD in J774 cells. Moreover, genes that were induced by LPS were up-regulated even in the absence of LPS by the phagocytosis of polymer conjugates, but such up-regulations were not suppressed by the treatment with MβCD. It was shown that lipid rafts play a significant role in incorporation of polymer conjugates through phagocytosis of macrophages, but their association with signal transduction in innate immunity is very limited.     

Polymorphisms in genes involved in innate immunity and susceptibility to benzene-induced hematotoxicity

Benzene, a recognized hematotoxicant and carcinogen, can damage the human immune system. We studied the association between single nucleotide polymorphisms (SNPs) in genes involved in innate immunity and benzene hematotoxicity in a cross-sectional study of workers exposed to benzene (250 workers and 140 controls). A total of 1,236 tag SNPs in 149 gene regions of six pathways were included in the analysis. Six gene regions were significant for their association with white blood cell (WBC) counts (MBP, VCAM1, ALOX5, MPO, RAC2, and CRP) based on gene-region (P<0.05) and SNP analyses (FDR<0.05). VCAM1 rs3176867, ALOX5 rs7099684, and MPO rs2071409 were the three most significant SNPs. They showed similar effects on WBC subtypes, especially granulocytes, lymphocytes, and monocytes. A 3-SNP block in ALOXE3 (rs7215658, rs9892383, and rs3027208) showed a global association (omnibus P = 0.0008) with WBCs even though the three SNPs were not significant individually. Our study suggests that polymorphisms in innate immunity genes may play a role in benzene-induced hematotoxicity; however, independent replication is necessary.