Edward K. Chow

Toll-like Receptors Induce a Phagocytic Gene Program through p38

Toll-like receptor (TLR) signaling and phagocytosis are hallmarks of macrophage-mediated innate immune responses to bacterial infection. However, the relationship between these two processes is not well established. Our data indicate that TLR ligands specifically promote bacterial phagocytosis, in both murine and human cells, through induction of a phagocytic gene program. Importantly, TLR-induced phagocytosis of bacteria was found to be reliant on myeloid differentiation factor 88–dependent signaling through interleukin-1 receptor–associated kinase-4 and p38 leading to the up-regulation of scavenger receptors. Interestingly, individual TLRs promote phagocytosis to varying degrees with TLR9 being the strongest and TLR3 being the weakest inducer of this process. We also demonstrate that TLR ligands not only amplify the percentage of phagocytes uptaking Escherichia coli, but also increase the number of bacteria phagocytosed by individual macrophages. Taken together, our data describe an evolutionarily conserved mechanism by which TLRs can specifically promote phagocytic clearance of bacteria during infection.

Type I IFNs mediate development of postinfluenza bacterial pneumonia in mice

Influenza-related complications continue to be a major cause of mortality worldwide. Due to unclear mechanisms, a substantial number of influenza-related deaths result from bacterial superinfections, particularly secondary pneumococcal pneumonia. Here, we report what we believe to be a novel mechanism by which influenza-induced type I IFNs sensitize hosts to secondary bacterial infections. Influenza-infected mice deficient for type I IFN-alpha/beta receptor signaling (Ifnar-/- mice) had improved survival and clearance of secondary Streptococcus pneumoniae infection from the lungs and blood, as compared with similarly infected wild-type animals. The less effective response in wild-type mice seemed to be attributable to impaired production of neutrophil chemoattractants KC (also known as Cxcl1) and Mip2 (also known as Cxcl2) following secondary challenge with S. pneumoniae. This resulted in inadequate neutrophil responses during the early phase of host defense against secondary bacterial infection. Indeed, influenza-infected wild-type mice cleared secondary pneumococcal pneumonia after pulmonary administration of exogenous KC and Mip2, whereas neutralization of Cxcr2, the common receptor for KC and Mip2, reversed the protective phenotype observed in Ifnar-/- mice. These data may underscore the importance of the type I IFN inhibitory pathway on CXC chemokine production. Collectively, these findings highlight what we believe to be a novel mechanism by which the antiviral response to influenza sensitizes hosts to secondary bacterial pneumonia.

Differential Requirement for TANK-binding Kinase-1 in Type I Interferon Responses to Toll-like Receptor Activation and Viral Infection

TANK-binding kinase-1 (TBK1) and the inducible IκB kinase (IKK-i) have been shown recently to activate interferon (IFN) regulatory factor-3 (IRF3), the primary transcription factor regulating induction of type I IFNs. Here, we have compared the role and specificity of TBK1 in the type I IFN response to lipopolysaccharide (LPS), polyI:C, and viral challenge by examining IRF3 nuclear translocation, signal transducer and activator of transcription 1 phosphorylation, and induction of IFN-regulated genes. The LPS and polyI:C-induced IFN responses were abolished and delayed, respectively, in macrophages from mice with a targeted disruption of the TBK1 gene. When challenged with Sendai virus, the IFN response was normal in TBK1−/− macrophages, but defective in TBK1−/− embryonic fibroblasts. Although both TBK1 and IKK-i are expressed in macrophages, only TBK1 but not IKK-i was detected in embryonic fibroblasts by Northern blotting analysis. Furthermore, the IFN response in TBK1−/− embryonic fibroblasts can be restored by reconstitution with wild-type IKK-i but not a mutant IKK-i lacking kinase activity. Thus, our studies suggest that TBK1 plays an important role in the Toll-like receptor–mediated IFN response and is redundant with IKK-i in the response of certain cell types to viral infection.

Toll-Like Receptor 3 Mediates a More Potent Antiviral Response Than Toll-Like Receptor 4

We have recently described an IFN regulatory factor 3-mediated antiviral gene program that is induced by both Toll-like receptor (TLR)3 and TLR4 ligands. In our current study, we show that activation of IFN/viral response gene expression in primary macrophage cells is stronger and prolonged with TLR3 stimulation compared with that of TLR4. Our data also reveal that the cytoplasmic tails of both TLR3 and TLR4 can directly interact with myeloid differentiation factor 88 (MyD88). However, although Toll/IL-1 receptor homology domain-containing adaptor protein/MyD88 adaptor-like is able to associate with TLR4, we were unable to detect any interaction between Toll/IL-1 receptor homology domain-containing adaptor protein/MyD88 adaptor-like and TLR3. By using quantitative real-time PCR assays, we found that TLR3 expression is inducible by both TLR3 and TLR4 ligands, while TLR4 expression is not inducible by these same stimuli. Furthermore, using cells derived from mice deficient in the IFN-αβR, we show that both TLR3 and TLR4 require IFN-β autocrine/paracrine feedback to induce TLR3 expression and activate/enhance genes required for antiviral activity. More specifically, a subset of antiviral genes is initially induced independent of IFN-β, yet the cytokine further enhances expression at later time points. This was in contrast to a second set of genes (including TLR3) that is induced only after IFN-β production. Taken together, our data argue that, despite both TLR3 and TLR4 being able to use IFN-β to activate/enhance antiviral gene expression, TLR3 uses multiple mechanisms to enhance and sustain the antiviral response more strongly than TLR4.

Multimodal nanodiamond drug delivery carriers for selective targeting, imaging, and enhanced chemotherapeutic efficacy.

The advancement of next-generation nanocarriers as drug delivery platforms will require the incorporation of these useful properties, through which adverse side effects of chemotherapy drugs can be avoided and overall treatment and diagnosis improved. As such, a variety of nanoparticle-based delivery systems have already been widely investigated and provided interesting avenues of research for improving cancer treatments through therapy and targeted delivery. [ 2–7 ]

Diamond‐Lipid Hybrids Enhance Chemotherapeutic Tolerance and Mediate Tumor Regression

Self-assembled nanodiamond-lipid hybrid particles (NDLPs) harness the potent interaction between the nanodiamond (ND)-surface and small molecules, while providing a mechanism for cell-targeted imaging and therapy of triple negative breast cancers. Epidermal growth factor receptor-targeted NDLPs are highly biocompatible particles that provide cell-specific imaging, promote tumor retention of ND-complexes, prevent epirubicin toxicities and mediate regression of triple negative breast cancers.

Oncogene‐specific formation of chemoresistant murine hepatic cancer stem cells

At least some cancer stem cells (CSCs) display intrinsic drug resistance that may thwart eradication of a malignancy by chemotherapy. We explored the genesis of such resistance by studying mouse models of liver cancer driven by either MYC or the combination of oncogenic forms of activation of v‐akt murine thymoma viral oncogene homolog (AKT) and NRAS. A common manifestation of chemoresistance in CSCs is efflux of the DNA‐binding dye Hoechst 33342. We found that only the MYC‐driven tumors contained a subset of cells that efflux Hoechst 33342. This “side population” (SP) was enriched for CSCs when compared to non‐SP tumor cells and exhibited markers of hepatic progenitor cells. The SP cells could differentiate into non‐SP tumor cells, with coordinate loss of chemoresistance, progenitor markers, and the enrichment for CSCs. In contrast, non‐SP cells did not give rise to SP cells. Exclusion of Hoechst 33342 is mediated by ATP binding cassette drug transporter proteins that also contribute to chemoresistance in cancer. We found that the multidrug resistance gene 1 (MDR1) transporter was responsible for the efflux of Hoechst from SP cells in our MYC‐driven model. Accordingly, SP cells and their tumor‐initiating subset were more resistant than non‐SP cells to chemotherapeutics that are effluxed by MDR1. Conclusion: The oncogenotype of a tumor can promote a specific mechanism of chemoresistance that can contribute to the survival of hepatic CSCs. Under circumstances that promote differentiation of CSCs into more mature tumor cells, the chemoresistance can be quickly lost. Elucidation of the mechanisms that govern chemoresistance in these mouse models may illuminate the genesis of chemoresistance in human liver cancer. (HEPATOLOGY 2012)

TLR agonists regulate PDGF‐B production and cell proliferation through TGF‐β/type I IFN crosstalk

Transforming growth factor‐β (TGF‐β) and type I interferon (IFN) autocrine/paracrine loops are recognized as key mediators of signaling cascades that control a variety of cellular functions. Here, we describe a novel mechanism by which Toll‐like receptor (TLR) agonists utilize these two autocrine/paracrine loops to differentially regulate the induction of PDGF‐B, a growth factor implicated in a number of diseases ranging from tumor metastasis to glomerulonephritis. We demonstrate that CpG‐specific induction of PDGF‐B requires activation of Smads through TGFβ1 autocrine/paracrine signaling. In contrast, polyinosinic:polycytidylic acid strongly represses CpGs as well as its own intrinsic ability to induce PDGF‐B mRNA through type I IFN‐mediated induction of Smad7, a negative regulator of Smad3/4. Furthermore, we have shown that this crosstalk mechanism translates into similar regulation of mesangial cell proliferation. Thus, our results demonstrate the importance of crosstalk between TGF‐β and type I IFNs in determining the specificity of TLR‐mediated gene induction.

A role for IRF3-dependent RXRα repression in hepatotoxicity associated with viral infections

Viral infections and antiviral responses have been linked to several metabolic diseases, including Reyes syndrome, which is aspirin-induced hepatotoxicity in the context of a viral infection. We identify an interferon regulatory factor 3 (IRF3)–dependent but type I interferon–independent pathway that strongly inhibits the expression of retinoid X receptor α (RXRα) and suppresses the induction of its downstream target genes, including those involved in hepatic detoxification. Activation of IRF3 by viral infection in vivo greatly enhances bile acid– and aspirin-induced hepatotoxicity. Our results provide a critical link between the innate immune response and host metabolism, identifying IRF3-mediated down-regulation of RXRα as a molecular mechanism for pathogen-associated metabolic diseases.

Innate immune system regulation of nuclear hormone receptors in metabolic diseases

The immune system modulates a number of biological processes to properly defend against pathogens. Here, we review how crosstalk between nuclear hormone receptors and the innate immune system may influence multiple biological functions during an immune response. Although nuclear hormone receptor repression of innate immune responses and inflammation has been well studied, a number of new studies have identified repression of nuclear hormone receptor signaling by various innate immune responses. IFN regulatory factor 3, a key transcription factor involved in the induction of antiviral genes, may play a role in mediating such crosstalk between the innate immune response and nuclear receptor‐regulated metabolism. This crosstalk mechanism is now implicated in the pathogenesis of atherosclerosis and Reye’s syndrome and could provide an explanation for other pathogen‐associated metabolic and developmental disorders.