Poly(I:C)

Poly r(C) binding protein (PCBP) 1 expression is regulated at the post-translation level in thyroid carcinoma.[Pubmed:28337299]

Am J Transl Res. 2017 Feb 15;9(2):708-714. eCollection 2017.

Poly r(C) binding protein (PCBP) 1 or heterogeneous ribonucleoprotein (hnRNP) E1 is a RNA binding protein that plays a vital role in a wide variety of biological processes. PCBP1 has been shown to function as a tumor suppressor by negatively regulating translation of pro-metastatic proteins in different cancers. Loss of PCBP1 expression or its Akt2-mediated phosphorylation at serine 43 residue has both been indicated to de-repress its regulation of EMT inducer proteins. Our previous work has established that PCBP1 functions as a tumor suppressor in thyroid cancer, where its translation is inhibited by microRNA-490-3p. Here we show that thyroid cancer patients can be divided into 2 cohorts based on miR-490-3p expression and PCBP1 mRNA expression-one cohort with high PCBP1 mRNA expression and basal miR-490-3p expression and a second cohort with low PCBP1 mRNA expression and high miR-490-3p expression. However, PCBP1 protein expression is also downregulated in the cohort with high PCBP1 mRNA expression, with expression levels similar to what is observed in patients with the low PCBP1 mRNA expression. Our analysis shows that PCBP1 mRNA is actively translated in patients with high PCBP1 mRNA expression, but that the protein is post translationally degraded by the proteasome machinery. Our results thus elucidate a novel mechanism responsible for down regulation of PCBP1 expression in thyroid cancer. It will be important in future to identify the mechanism that causes degradation of PCBP1 protein and to identify if similar mechanisms are active in other tumors characterized by low PCBP1 protein expression.

Synthesis of Poly(vitamin C) through ADMET.[Pubmed:28321953]

Macromol Rapid Commun. 2017 Jun;38(11).

l-Ascorbic acid, commonly known as vitamin C and one of the most important biological compounds, is converted to a alpha,omega-diene monomer and subsequently polymerized for the first time by acyclic diene metathesis. Various experimental conditions such as polymerization medium, catalyst type, temperature, and monomer/catalyst ratio are studied. The moderate molecular weight polymers are achieved when the polymerizations are conducted under bulk conditions employing the Grubbs first generation (G1) or Hoveyda-Grubbs second generation catalyst (HG-2). In the solution case, on the other hand, low molecular weight polymers are obtained regardless of the catalyst type. Moreover, when the catalyst performances are compared, it is found that G1 produces the higher molecular weight as well as higher yield polymers with respect to the HG-2.

Transactivation Domain of Human c-Myc Is Essential to Alleviate Poly(Q)-Mediated Neurotoxicity in Drosophila Disease Models.[Pubmed:28316031]

J Mol Neurosci. 2017 May;62(1):55-66.

Polyglutamine (poly(Q)) disorders, such as Huntington's disease (HD) and spinocerebellar ataxias, represent a group of neurological disorders which arise due to an atypically expanded poly(Q) tract in the coding region of the affected gene. Pathogenesis of these disorders inside the cells begins with the assembly of these mutant proteins in the form of insoluble inclusion bodies (IBs), which progressively sequester several vital cellular transcription factors and other essential proteins, and finally leads to neuronal dysfunction and apoptosis. We have shown earlier that targeted upregulation of Drosophila myc (dmyc) dominantly suppresses the poly(Q) toxicity in Drosophila. The present study examines the ability of the human c-myc proto-oncogene and also identifies the specific c-Myc isoform which drives the mitigation of poly(Q)-mediated neurotoxicity, so that it could be further substantiated as a potential drug target. We report for the first time that similar to dmyc, tissue-specific induced expression of human c-myc also suppresses poly(Q)-mediated neurotoxicity by an analogous mechanism. Among the three isoforms of c-Myc, the rescue potential was maximally manifested by the full-length c-Myc2 protein, followed by c-Myc1, but not by c-MycS which lacks the transactivation domain. Our study suggests that strategies focussing on the transactivation domain of c-Myc could be a very useful approach to design novel drug molecules against poly(Q) disorders.

Anti-inflammatory mechanism of lonchocarpine in LPS- or poly(I:C)-induced neuroinflammation.[Pubmed:28288940]

Pharmacol Res. 2017 May;119:431-442.

Neuroinflammation plays an important role in the progression of various neurodegenerative diseases. In this study, we investigated the anti-inflammatory effects of lonchocarpine, a natural compound isolated from Abrus precatorius, under in vitro and in vivo neuroinflammatory conditions induced by challenge with lipopolysaccharide (LPS)- or polyinosinic-polycytidylic acid (Poly(I:C)). Lonchocarpine suppressed the expression of iNOS and proinflammatory cytokines in LPS or Poly(I:C)-stimulated BV2 microglial cells. These anti-inflammatory effects were verified in brains of mice with systemic inflammation induced by administration of LPS or Poly(I:C). Lonchocarpine reduced the number of Iba-1-positive activated microglia, and suppressed the mRNA expression of various proinflammatory markers in the cortex of LPS- or Poly(I:C)-injected mice. Molecular mechanistic experiments showed that lonchocarpine inhibited NF-kappaB activity by reducing the phosphorylation and degradation of IkappaBalpha in LPS- or Poly(I:C)-stimulated BV2 cells. Analysis of further upstream signaling pathways in LPS-stimulated microglia showed that lonchocarpine inhibited the phosphorylation of IkappaB kinase and TGFbeta-activated kinase 1 (TAK1). Moreover, lonchocarpine suppressed the interaction of myeloid differentiation factor 88 (MyD88) and intereleukin-1 receptor-associated kinase 4 (IRAK4). These data suggest that toll-like receptor 4 downstream signals such as MyD88/IRAK4-TAK1-NF-kappaB are at least partly involved in the anti-inflammatory mechanism of lonchocarpine in LPS-stimulated microglia. Its strong anti-inflammatory effects may make lonchocarpine an effective preventative drug for neuroinflammatory disorders that are associated with systemic inflammation.

TLR signalling augments macrophage bactericidal activity through mitochondrial ROS.[Pubmed:21525932]

Nature. 2011 Apr 28;472(7344):476-80.

Reactive oxygen species (ROS) are essential components of the innate immune response against intracellular bacteria and it is thought that professional phagocytes generate ROS primarily via the phagosomal NADPH oxidase machinery. However, recent studies have suggested that mitochondrial ROS (mROS) also contribute to mouse macrophage bactericidal activity, although the mechanisms linking innate immune signalling to mitochondria for mROS generation remain unclear. Here we demonstrate that engagement of a subset of Toll-like receptors (TLR1, TLR2 and TLR4) results in the recruitment of mitochondria to macrophage phagosomes and augments mROS production. This response involves translocation of a TLR signalling adaptor, tumour necrosis factor receptor-associated factor 6 (TRAF6), to mitochondria, where it engages the protein ECSIT (evolutionarily conserved signalling intermediate in Toll pathways), which is implicated in mitochondrial respiratory chain assembly. Interaction with TRAF6 leads to ECSIT ubiquitination and enrichment at the mitochondrial periphery, resulting in increased mitochondrial and cellular ROS generation. ECSIT- and TRAF6-depleted macrophages have decreased levels of TLR-induced ROS and are significantly impaired in their ability to kill intracellular bacteria. Additionally, reducing macrophage mROS levels by expressing catalase in mitochondria results in defective bacterial killing, confirming the role of mROS in bactericidal activity. These results reveal a novel pathway linking innate immune signalling to mitochondria, implicate mROS as an important component of antibacterial responses and further establish mitochondria as hubs for innate immune signalling.

Poly(I:C) used for human dendritic cell maturation preserves their ability to secondarily secrete bioactive IL-12.[Pubmed:15096480]

Int Immunol. 2004 May;16(5):767-73.

Dendritic cells (DC) are professional antigen-presenting cells that play a central role in the control of immunity. Mature DC are characterized by high expression levels of MHC and co-stimulatory molecules, and by the secretion of IL-12, a key cytokine for the priming of cytotoxic T lymphocytes. Here, we have compared different maturation stimuli to reproducibly generate stable mature DC secreting high amounts of bioactive IL-12p70. We have compared soluble human trimeric CD40 ligand (sCD40L) combined with IFN-gamma, Poly(I:C), a cocktail of cytokines (IL-1beta, IL-6 and tumor necrosis factor-alpha) with prostaglandin E(2) and lipopolysaccharide. A major concern, however, is whether DC, that have already produced high amounts of IL-12p70 during the maturation step, are still capable of secreting IL-12p70 after in vivo administration at the time of interaction with the targeted T cells. To mimic that situation, mature DC generated by those methods were compared for their ability to secrete IL-12p70 in the absence of IFN-gamma, using sCD40L. We observed a second consistent secretion of bioactive IL-12p70 upon subsequent sCD40L stimulation only when Poly(I:C) was used as the maturating agent. Our data suggest that, for clinical use, Poly(I:C) may be one of the most appropriate agents to generate stable mature DC. These mature DC might generate in vivo effective immune responses after injection, because they retain the ability to secrete bioactive IL-12 after CD40 ligation.

Nuclear factor-kappaB translocation mediates double-stranded ribonucleic acid-induced NIT-1 beta-cell apoptosis and up-regulates caspase-12 and tumor necrosis factor receptor-associated ligand (TRAIL).[Pubmed:12960048]

Endocrinology. 2003 Oct;144(10):4616-25.

The mechanism of induction of apoptosis by double-stranded RNA (dsRNA) is not fully characterized. The dsRNA is normally present in extremely low quantities in cells, but following infection with RNA viruses, large quantities of the dsRNA viral replicative intermediate may be produced triggering the antiviral response as well as cell death. In this report, transfection of polyinosinic-polycytidylic acid [Poly(I:C)] into NIT 1 cells has been used as a model of intracellular dsRNA-induced beta-cell apoptosis. At 18 h post transfection, 45% of the cells were apoptotic as indicated by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling (TUNEL) staining, and this was accompanied by an increase in nuclear factor kappaB (NF-kappaB) p50/p65 nuclear translocation and cleavage of caspases 3 and 8. The NF-kappaB inhibitor peptide, SN50, significantly reduced caspase-3 activity and the percentage of TUNEL-positive cells, substantiating a role for NF-kappaB in inducing intracellular dsRNA-mediated apoptosis. Concomitantly, RNA-dependent protein kinase activity was observed at 3 h post transfection along with phosphorylation and degradation of inhibitory kappaB-alpha. Expression of TRAIL (TNF-related apoptosis-inducing ligand), Fas, IL-15, and caspase-12 mRNAs was up-regulated in the presence of Poly(I:C) but not when SN50 was also added. In contrast, there was no change detected in Fas, Fas-associated death domain, Bcl-2, Bcl-xl, Bax, p53, or XIAP(X-linked inhibitor of apoptosis protein) expression up to 12 h after Poly(I:C) transfection. In addition, caspase-12 was cleaved, and phosphorylation of eukaryotic initiation factor 2alpha occurred, suggesting that an endoplasmic reticulum stress pathway was involved in addition to NF-kappaB induction of an extrinsic pathway, possibly mediated by TNF-related apoptosis-inducing ligand.