Dendritic Cell-Mediated, DNA-Based Vaccination Against Hepatitis C Induces the Multi-Epitope-Specific Response of Humanized, HLA Transgenic Mice

Hepatitis C virus (HCV) is the etiologic agent of chronic liver disease, hepatitis C. Spontaneous resolution of viral infection is associated with vigorous HLA class I- and class II-restricted T cell responses to multiple viral epitopes. Unfortunately, only 20% of patients clear infection spontaneously, most develop chronic disease and require therapy. The response to chemotherapy varies, however; therapeutic vaccination offers an additional treatment strategy. To date, therapeutic vaccines have demonstrated only limited success. Vector-mediated vaccination with multi-epitope-expressing DNA constructs alone or in combination with chemotherapy offers an additional treatment approach. Gene sequences encoding validated HLA-A2- and HLA-DRB1-restricted epitopes were synthesized and cloned into an expression vector. Dendritic cells (DCs) derived from humanized, HLA-A2/DRB1 transgenic (donor) mice were transfected with these multi-epitope-expressing DNA constructs. Recipient HLA-A2/DRB1 mice were vaccinated s.c. with transfected DCs; control mice received non-transfected DCs. Peptide-specific IFN-γ production by splenic T cells obtained at 5 weeks post-immunization was quantified by ELISpot assay; additionally, the production of IL-4, IL-10 and TNF-α were quantified by cytokine bead array. Splenocytes derived from vaccinated HLA-A2/DRB1 transgenic mice exhibited peptide-specific cytokine production to the vast majority of the vaccine-encoded HLA class I- and class II-restricted T cell epitopes. A multi-epitope-based HCV vaccine that targets DCs offers an effective approach to inducing a broad immune response and viral clearance in chronic, HCV-infected patients

Coupling sensitive \u3cem\u3ein vitro\u3c/em\u3e and in silico techniques to assess cross-reactive CD4\u3csup\u3e+\u3c/sup\u3e T cells against the swine-origin H1N1 influenza virus

The outbreak of the novel swine-origin H1N1 influenza in the spring of 2009 took epidemiologists, immunologists, and vaccinologists by surprise and galvanized a massive worldwide effort to produce millions of vaccine doses to protect against this single virus strain. Of particular concern was the apparent lack of pre-existing antibody capable of eliciting cross-protective immunity against this novel virus, which fueled fears this strain would trigger a particularly far-reaching and lethal pandemic. Given that disease caused by the swine-origin virus was far less severe than expected, we hypothesized cellular immunity to cross-conserved T cell epitopes might have played a significant role in protecting against the pandemic H1N1 in the absence of cross-reactive humoral immunity. In a published study, we used an immunoinformatics approach to predict a number of CD4+ T cell epitopes are conserved between the 2008–2009 seasonal H1N1 vaccine strain and pandemic H1N1 (A/California/04/2009) hemagglutinin proteins. Here, we provide results from biological studies using PBMCs from human donors not exposed to the pandemic virus to demonstrate that pre-existing CD4+ T cells can elicit cross-reactive effector responses against the pandemic H1N1 virus. As well, we show our computational tools were 80–90% accurate in predicting CD4+ T cell epitopes and their HLA-DRB1-dependent response profiles in donors that were chosen at random for HLA haplotype. Combined, these results confirm the power of coupling immunoinformatics to define broadly reactive CD4+ T cell epitopes with highly sensitive in vitro biological assays to verify these in silico predictions as a means to understand human cellular immunity, including cross-protective responses, and to define CD4+ T cell epitopes for potential vaccination efforts against future influenza viruses and other pathogens

HCV Epitope, Homologous to Multiple Human Protein Sequences, Induces a Regulatory T Cell Response in Infected Patients

Background & Aims: Spontaneous resolution of hepatitis C virus (HCV) infections depends upon a broad T cell response to multiple viral epitopes. Most patients fail to clear infections spontaneously, however, and develop chronic disease. The elevated number and function of CD3+CD4+CD25+FoxP3+ regulatory T(reg) cells in HCV-infected patients suggest the role of Treg cells in impaired viral clearance. The factors contributing to increased Treg cell activity in chronic hepatitis C cases remain to be delineated. Methods: Immunoinformatics tools were used to predict promiscuous, highly-conserved HLA-DRB1- restricted immunogenic consensus sequences (ICS), each composed of multiple T cell epitopes. These sequences were synthesized and added to cultures of peripheral blood mononuclear cells (PBMCs) derived from patients who resolved HCV infection spontaneously, patients with persistent infection, and non-infected individuals. The cells were collected following 5 days incubation, quantified and characterized by flow cytometry. Results: One ICS, HCV_G1_p7_794, induced a marked increase in Treg cells in PBMC cultures derived from infected patients, but not patients who spontaneously cleared HCV or non-infected individuals. An analogous human peptide (p7_794), on the other hand, induced a significant increase in Treg cells among PBMCs derived from both HCV infected and non-infected individuals. JanusMatrix analyses determined that HCV_G1_p7_794 is comprised of Treg cell epitopes that exhibit extensive cross-reactivity with the human proteome. Conclusion: A virus-encoded peptide (HCV_G1_p7_794) with extensive human homology activates cross-reactive CD3+CD4+CD25+FoxP3+ nTreg cells, which potentially contribute to immunosuppression and chronic hepatitis C

Mortality and pulmonary complications in patients undergoing surgery with perioperative SARS-CoV-2 infection: an international cohort study

Background: The impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on postoperative recovery needs to be understood to inform clinical decision making during and after the COVID-19 pandemic. This study reports 30-day mortality and pulmonary complication rates in patients with perioperative SARS-CoV-2 infection. Methods: This international, multicentre, cohort study at 235 hospitals in 24 countries included all patients undergoing surgery who had SARS-CoV-2 infection confirmed within 7 days before or 30 days after surgery. The primary outcome measure was 30-day postoperative mortality and was assessed in all enrolled patients. The main secondary outcome measure was pulmonary complications, defined as pneumonia, acute respiratory distress syndrome, or unexpected postoperative ventilation. Findings: This analysis includes 1128 patients who had surgery between Jan 1 and March 31, 2020, of whom 835 (74·0%) had emergency surgery and 280 (24·8%) had elective surgery. SARS-CoV-2 infection was confirmed preoperatively in 294 (26·1%) patients. 30-day mortality was 23·8% (268 of 1128). Pulmonary complications occurred in 577 (51·2%) of 1128 patients; 30-day mortality in these patients was 38·0% (219 of 577), accounting for 81·7% (219 of 268) of all deaths. In adjusted analyses, 30-day mortality was associated with male sex (odds ratio 1·75 [95% CI 1·28–2·40], p\textless0·0001), age 70 years or older versus younger than 70 years (2·30 [1·65–3·22], p\textless0·0001), American Society of Anesthesiologists grades 3–5 versus grades 1–2 (2·35 [1·57–3·53], p\textless0·0001), malignant versus benign or obstetric diagnosis (1·55 [1·01–2·39], p=0·046), emergency versus elective surgery (1·67 [1·06–2·63], p=0·026), and major versus minor surgery (1·52 [1·01–2·31], p=0·047). Interpretation: Postoperative pulmonary complications occur in half of patients with perioperative SARS-CoV-2 infection and are associated with high mortality. Thresholds for surgery during the COVID-19 pandemic should be higher than during normal practice, particularly in men aged 70 years and older. Consideration should be given for postponing non-urgent procedures and promoting non-operative treatment to delay or avoid the need for surgery. Funding: National Institute for Health Research (NIHR), Association of Coloproctology of Great Britain and Ireland, Bowel and Cancer Research, Bowel Disease Research Foundation, Association of Upper Gastrointestinal Surgeons, British Association of Surgical Oncology, British Gynaecological Cancer Society, European Society of Coloproctology, NIHR Academy, Sarcoma UK, Vascular Society for Great Britain and Ireland, and Yorkshire Cancer Research

Dawn Orbit Determination Team: Modeling and Fitting of Optical Data at Vesta

No abstract availabl

Dawn Orbit Determination Team: Modeling and Fitting of Optical Data at Vesta

The Dawn spacecraft was launched on September 27th, 2007. Its mission is to consecutively rendezvous with and observe the two largest bodies in the main asteroid belt, Vesta and Ceres. It has already completed over a year's worth of direct observations of Vesta (spanning from early 2011 through late 2012) and is currently on a cruise trajectory to Ceres, where it will begin scientific observations in mid-2015. Achieving this data collection required careful planning and execution from all Dawn operations teams. Dawn's Orbit Determination (OD) team was tasked with reconstruction of the as-flown trajectory as well as determination of the Vesta rotational rate, pole orientation and ephemeris, among other Vesta parameters. Improved knowledge of the Vesta pole orientation, specifically, was needed to target the final maneuvers that inserted Dawn into the first science orbit at Vesta. To solve for these parameters, the OD team used radiometric data from the Deep Space Network (DSN) along with optical data reduced from Dawn's Framing Camera (FC) images. This paper will de-scribe the initial determination of the Vesta ephemeris and pole using a combination of radiometric and optical data, and also the progress the OD team has made since then to further refine the knowledge of Vesta's body frame orientation and rate with these data

Dawn Orbit Determination Team: Trajectory and Gravity Prediction Performance During Vesta Science Phases

The Dawn spacecraft was launched on September 27th, 2007. Its mission is to consecutively rendezvous with and observe the two largest bodies in the asteroid belt, Vesta and Ceres. It has already completed over a year's worth of direct observations of Vesta (spanning from early 2011 through late 2012) and is currently on a cruise trajectory to Ceres, where it will begin scientific observations in mid-2015. Achieving this data collection required careful planning and execution from all spacecraft teams. Dawn's Orbit Determination (OD) team was tasked with accurately predicting the trajectory of the Dawn spacecraft during the Vesta science phases, and also determining the parameters of Vesta to support future science orbit design. The future orbits included the upcoming science phase orbits as well as the transfer orbits between science phases. In all, five science phases were executed at Vesta, and this paper will describe some of the OD team contributions to the planning and execution of those phases

DCs derived from Flt3L-treated, HLA-A2/DRB1 mice exhibit an immature phenotype^a.

a<p>CD11c⁺PDCA-1⁺ DCs, purified from 2.55×10⁸ splenocytes/HLA-A2/DRB1 transgenic mouse inoculated s.c. with Flt3L-secreting B16 myeloma cells 12 days previously, were quantified and characterized by flow cytometry.</p>b<p>Not determined.</p

IFN-γ ELISpot assays.

<p>The spleens were dissected and pooled on day 35 from groups of 4 mice immunized with vaccine construct-transfected, and IFN-γ ELISpot assays were performed in triplicate. The data, expressed as the means ± SD ELISpots/10⁶ splenocytes minus the average negative control, 0.1% DMSO+2 SD (3,346 ELISpots/10⁶ splenocytes), were obtained in a single experiment representative of duplicate experiments. The average number of ELISpots/10⁶ splenocytes derived from mice immunized with non-transfected DCs was not significantly different from the DMSO control (not shown). *Splenocytes derived from mice immunized with transfected DCs and incubated with the HLA-A2- and -DRB1-restricted peptides indicated did not yield values that were significantly different from the DMSO control (ANOVA).</p