Detection of viruses by host pattern recognition receptors induces the expression of type I interferon (IFN) and IFN-stimulated genes (ISGs), which suppress viral replication. Numerous studies have ...described HIV-1 as a poor activator of innate immunity in vitro. The exact role that the viral capsid plays in this immune evasion is not fully understood.
To better understand the role of the HIV-1 capsid in sensing we tested the effect of making HIV-1 by co-expressing a truncated Gag that encodes the first 107 amino acids of capsid fused with luciferase or GFP, alongside wild type Gag-pol. We found that unlike wild type HIV-1, viral particles produced with a mixture of wild type and truncated Gag fused to luciferase or GFP induced a potent IFN response in THP-1 cells and macrophages. Innate immune activation by Gag-fusion HIV-1 was dependent on reverse transcription and DNA sensor cGAS, suggesting activation of an IFN response by viral DNA. Further investigation revealed incorporation of the Gag-luciferase/GFP fusion proteins into viral particles that correlated with subtle defects in wild type Gag cleavage and a diminished capacity to saturate restriction factor TRIM5α, likely due to aberrant particle formation. We propose that expression of the Gag fusion protein disturbs the correct cleavage and maturation of wild type Gag, yielding viral particles that are unable to effectively shield viral DNA from detection by innate sensors including cGAS.
These data highlight the crucial role of capsid in innate evasion and support growing literature that disruption of Gag cleavage and capsid formation induces a viral DNA- and cGAS-dependent innate immune response. Together these data demonstrate a protective role for capsid and suggest that antiviral activity of capsid-targeting antivirals may benefit from enhanced innate and adaptive immunity in vivo.
Memory T cells induced by previous pathogens can shape susceptibility to, and the clinical severity of, subsequent infections
. Little is known about the presence in humans of pre-existing memory T ...cells that have the potential to recognize severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here we studied T cell responses against the structural (nucleocapsid (N) protein) and non-structural (NSP7 and NSP13 of ORF1) regions of SARS-CoV-2 in individuals convalescing from coronavirus disease 2019 (COVID-19) (n = 36). In all of these individuals, we found CD4 and CD8 T cells that recognized multiple regions of the N protein. Next, we showed that patients (n = 23) who recovered from SARS (the disease associated with SARS-CoV infection) possess long-lasting memory T cells that are reactive to the N protein of SARS-CoV 17 years after the outbreak of SARS in 2003; these T cells displayed robust cross-reactivity to the N protein of SARS-CoV-2. We also detected SARS-CoV-2-specific T cells in individuals with no history of SARS, COVID-19 or contact with individuals who had SARS and/or COVID-19 (n = 37). SARS-CoV-2-specific T cells in uninfected donors exhibited a different pattern of immunodominance, and frequently targeted NSP7 and NSP13 as well as the N protein. Epitope characterization of NSP7-specific T cells showed the recognition of protein fragments that are conserved among animal betacoronaviruses but have low homology to 'common cold' human-associated coronaviruses. Thus, infection with betacoronaviruses induces multi-specific and long-lasting T cell immunity against the structural N protein. Understanding how pre-existing N- and ORF1-specific T cells that are present in the general population affect the susceptibility to and pathogenesis of SARS-CoV-2 infection is important for the management of the current COVID-19 pandemic.
Background and Aims
HBV‐specific T‐cell receptor (HBV‐TCR) engineered T cells have the potential for treating HCC relapses after liver transplantation, but their efficacy can be hampered by the ...concomitant immunosuppressive treatment required to prevent graft rejection. Our aim is to molecularly engineer TCR‐T cells that could retain their polyfunctionality in such patients while minimizing the associated risk of organ rejection.
Approach and Results
We first analyzed how immunosuppressive drugs can interfere with the in vivo function of TCR‐T cells in liver transplanted patients with HBV‐HCC recurrence receiving HBV‐TCR T cells and in vitro in the presence of clinically relevant concentrations of immunosuppressive tacrolimus (TAC) and mycophenolate mofetil (MMF). Immunosuppressive Drug Resistant Armored TCR‐T cells of desired specificity (HBV or Epstein‐Barr virus) were then engineered by concomitantly electroporating mRNA encoding specific TCRs and mutated variants of calcineurin B (CnB) and inosine‐5′‐monophosphate dehydrogenase (IMPDH), and their function was assessed through intracellular cytokine staining and cytotoxicity assays in the presence of TAC and MMF. Liver transplanted HBV‐HCC patients receiving different immunosuppressant drugs exhibited varying levels of activated (CD39+ Ki67+) peripheral blood mononuclear cells after HBV‐TCR T‐cell infusions that positively correlate with clinical efficacy. In vitro experiments with TAC and MMF showed a potent inhibition of TCR‐T cell polyfunctionality. This inhibition can be effectively negated by the transient overexpression of mutated variants of CnB and IMPDH. Importantly, the resistance only lasted for 3‐5 days, after which sensitivity was restored.
Conclusions
We engineered TCR‐T cells of desired specificities that transiently escape the immunosuppressive effects of TAC and MMF. This finding has important clinical applications for the treatment of HBV‐HCC relapses and other pathologies occurring in organ transplanted patients.
Small molecule drugs bind to a pocket in disease causing target proteins based on complementarity in shape and physicochemical properties. There is a likelihood that other proteins could have binding ...sites that are structurally similar to the target protein. Binding to these other proteins could alter their activities leading to off target effects of the drug. One such small molecule drug Nutlin binds the protein MDM2, which is upregulated in several types of cancer and is a negative regulator of the tumor suppressor protein p53. To investigate the off target effects of Nutlin, we present here a shape-based data mining effort. We extracted the binding pocket of Nutlin from the crystal structure of Nutlin bound MDM2. We next mined the protein structural database (PDB) for putative binding pockets in other human protein structures that were similar in shape to the Nutlin pocket in MDM2 using our topology-independent structural superimposition tool CLICK. We detected 49 proteins which have binding pockets that were structurally similar to the Nutlin binding site of MDM2. All of the potential complexes were evaluated using molecular mechanics and AutoDock Vina docking scores. Further, molecular dynamics simulations were carried out on four of the predicted Nutlin–protein complexes. The binding of Nutlin to one of these proteins, gamma glutamyl hydrolase, was also experimentally validated by a thermal shift assay. These findings provide a platform for identifying potential off-target effects of existing/new drugs and also opens the possibilities for repurposing drugs/ligands.
Summary
Recurrence of hepatitis B virus-related hepatocellular carcinoma (HBV-HCC) after liver transplant (LT) is mediated by circulating tumour cells (CTCs) and exacerbated by the immunosuppressants ...required to prevent graft rejection. To circumvent the effects of immunosuppressants, we developed immunosuppressive drug-resistant armoured HBV-specific T-cell receptor-redirected T cells (IDRA HBV-TCR). However, their ability to eliminate HBV-HCC circulating in the whole blood has never been tested, and whether their lytic efficacy is compatible with the number of adoptively transferred T cells in vivo has never been measured. Hence, we developed a microscopy-based assay to quantify CTCs in whole blood. The assay was then used to quantify the efficacy of IDRA HBV-TCRs to lyse free-floating HBV-HCC cells in the presence of Tacrolimus and Mycophenolate Mofetil (MMF). We demonstrated that a panel of antibodies (AFP, GPC3, Vimentin, pan-Cytokeratin, and CD45) specific for HCC tumour antigens and immune cells can effectively differentiate HCC-CTCs in whole blood. Through dose-titration experiments, we observed that in the presence of immunosuppressive drugs, a minimum of 20 000 IDRA HBV-TCR T cells/ml of whole blood is necessary to lyse ~63.5% of free-floating HBV-HCC cells within 16 hours. In conclusion, IDRA HBV-TCR T cells can lyse free-floating HBV-HCC cells in whole blood in the presence of Tacrolimus and MMF. The quantity of IDRA-HBV TCR T cells required can be achieved by the adoptive transfer of 5 × 106 IDRA-HBV TCR-T cells/kg, supporting the utilisation of IDRA HBV-TCR T cells to eliminate CTCs as prophylaxis against recurrence after LT.
Da-Dao-Cheng district, with various kinds of historic shop-houses, is located in Metropolis of Taipei. The conservation movement by the citizens, NPOs, and experts began with the road-widening urban ...plan on the main street, Di-Hwa Street, and led to the change of policies to conservation. The conservation plan was officially announced in 2000 after various discussions, by designated as a special district with the use of system of Transfer of Development Rights. This paper discusses the achievement and issues of the conservation plan, by analyzing the process of pran making, changing of conservation proposals, and the current operations.
碩士
東南科技大學
防災科技研究所
100
This research takes the method of questionnaire survey. 202 of Tungnan University students who had had auto-bike traffic accidents were interviewed as the research objects. The ...data was analyzed with descriptive statistics. The main finding reveals that improper auto-bike riding habits definitely results to the increase of traffic accidents. Accordingly, the researcher suggested that the authorized department should aggressively educate the traffic rules knowledge and reinforce safety awareness to students to improve their riding habits so that the student auto-bike accident rate can be effectively decreased.
The purpose of this paper is to analyze the characteristics and issues of the preservation system for historic environments after Shu-Shu earthquake happened in Taiwan in 1999, by inspecting the ...amendment of the related laws on Cultural property Preservation. It explains by the following aspects. (1) The transition of preservation system for historic environments before Shu-shu earthquake, (2) The response to the survey and renovation of damaged historic buildings and streets after the earthquake, (3) The amendments of the cultural property related laws, (4) The characteristics and issues of the cultural property related laws after Shu-Shu earthquake, (5) The current status. It could be said that the preservation system of historic environment is facing the following issues after earthquake:(1) Urgent and uncompleted amendments, (2) Introduction of historic building registration system, (3) The separated government departments in charge, etc. Consequently, it will remain to be discussed until the accomplishment of the latest amendment of cultural property law.
PDF
