Auditory brainstem responses in the nine-banded armadillo (Dasypus novemcinctus).

The auditory brainstem response (ABR) to tone burst stimuli of thirteen frequencies ranging from 0.5 to 48 kHz was recorded in the nine-banded armadillo (Dasypus novemcinctus), the only extant member of the placental mammal superorder Xenarthra in North America. The armadillo ABR consisted of five main peaks that were visible within the first 10 ms when stimuli were presented at high intensities. The latency of peak I of the armadillo ABR increased as stimulus intensity decreased by an average of 20 µs/dB. Estimated frequency-specific thresholds identified by the ABR were used to construct an estimate of the armadillo audiogram describing the mean thresholds of the eight animals tested. The majority of animals tested (six out of eight) exhibited clear responses to stimuli from 0.5 to 38 kHz, and two animals exhibited responses to stimuli of 48 kHz. Across all cases, the lowest thresholds were observed for frequencies from 8 to 12 kHz. Overall, we observed that the armadillo estimated audiogram bears a similar pattern as those observed using ABR in members of other mammalian clades, including marsupials and later-derived placental mammals.

Transapical Cannulation With a Dual Lumen Cannula for Mechanical Circulatory Support in Cardiogenic Shock.

Temporary mechanical circulatory support can be delivered through a variety of techniques, including percutaneous left ventricular assist devices, surgically implanted rotary pumps, and veno-arterial extracorporeal membrane oxygenation. However, limitations include the effects of high afterload, intravascular hemolysis, patient vascular anatomy, surgical morbidity, and limited patient mobility which can hinder patient recovery. We describe a series of patients managed with transapical left ventricular mechanical circulatory support using a dual lumen cannula for the management of cardiogenic shock as a bridge to recovery or definitive decision. This support strategy may represent an additional option in the care for patients with cardiogenic shock that can provide full temporary anterograde mechanical circulatory support while potentially improving patient mobility and minimizing device-related complications.

IFNΛ3/4 locus polymorphisms and IFNΛ3 circulating levels are associated with COPD severity and outcomes.

BACKGROUND: Interferon lambdas (IFNLs) have important anti-viral/bacterial and immunomodulatory functions in the respiratory tract. How do IFNLs impact COPD and its exacerbations? METHODS: Five hundred twenty eight patients were recruited in a prospective observational multicentre cohort (PROMISE) study. The genetic polymorphisms (rs8099917 and rs12979860) within the IFNL3/4 gene region and circulating levels of IFNL3 in COPD patients were determined and associated with disease activity and outcome during a median follow-up of 24 months. RESULTS: The GG genotype significantly influenced severe exacerbation rate (42 vs. 23%; p = 0.032) and time to severe exacerbation (HR = 2.260; p = 0.012). Compared to the TT or TG genotypes, the GG genotype was associated with severe dyspnoea (modified medical research council score ≥ median 3; 22 vs 42%, p = 0.030). The CC genotype of the rs12979860 SNP was associated with a poorer prognosis (body mass index, airflow obstruction, dyspnea and exercise capacity index ≥ median 4; 46 vs. 36% TC vs. 20.5% TT; p = 0.031). Patients with stable COPD and at exacerbation had significantly lower circulating IFNL3 compared to healthy controls (p < 0.001 and p < 0.001, respectively). Circulating IFNL3 correlated to post-bronchodilator FEV1%predicted and the tissue maturation biomarker Pro-collagen 3. CONCLUSION: IFNL3/4 polymorphisms and circulating IFNL3 may be associated with disease activity and outcomes in COPD. TRIAL REGISTRATION: Clinical Trial registration http://www.isrctn.com/ identifier ISRCTN99586989 on 16 April 2008.

IL-28B is a key regulator of B- and T-cell vaccine responses against influenza.

Influenza is a major cause of morbidity and mortality in immunosuppressed persons, and vaccination often confers insufficient protection. IL-28B, a member of the interferon (IFN)-λ family, has variable expression due to single nucleotide polymorphisms (SNPs). While type-I IFNs are well known to modulate adaptive immunity, the impact of IL-28B on B- and T-cell vaccine responses is unclear. Here we demonstrate that the presence of the IL-28B TG/GG genotype (rs8099917, minor-allele) was associated with increased seroconversion following influenza vaccination (OR 1.99 p = 0.038). Also, influenza A (H1N1)-stimulated T- and B-cells from minor-allele carriers showed increased IL-4 production (4-fold) and HLA-DR expression, respectively. In vitro, recombinant IL-28B increased Th1-cytokines (e.g. IFN-γ), and suppressed Th2-cytokines (e.g. IL-4, IL-5, and IL-13), H1N1-stimulated B-cell proliferation (reduced 70%), and IgG-production (reduced>70%). Since IL-28B inhibited B-cell responses, we designed antagonistic peptides to block the IL-28 receptor α-subunit (IL28RA). In vitro, these peptides significantly suppressed binding of IFN-λs to IL28RA, increased H1N1-stimulated B-cell activation and IgG-production in samples from healthy volunteers (2-fold) and from transplant patients previously unresponsive to vaccination (1.4-fold). Together, these findings identify IL-28B as a key regulator of the Th1/Th2 balance during influenza vaccination. Blockade of IL28RA offers a novel strategy to augment vaccine responses.

Validation of TaqMan® SNP genotyping specificity for rs12979860 of IL-28B: modeling primer specificity in vitro.

Members of the type III interferon gene family arose by gene duplication events and have retained a high percent identity both in their coding and non-coding regions. In this study, the specificity of a widely used TaqMan(®) SNP genotyping assay for rs12979860 is validated. The 66 bp template for SNP genotyping has only 3 bp at one 5' end that vary between IL-28B and IL-28A; excluding the rs12979860 SNP itself. Conflicting annealing temperatures were found for the mismatched 19 bp primer to IL-28B and IL-28A with in silico melting temperature algorithms, or with in vitro dissociation curves. In order to prove specificity for IL-28B, an in vitro competition assay was setup with genomic DNA and synthetic oligonucleotides. When genomic DNA, containing equimolar concentrations of rs12979860 and the homologous region of IL-28A are present, no off-target amplification was observed. This SNP genotyping assay is therefore specific for rs12979860 and all previously reported results are valid. Finally, using a completely synthetic in vitro competition assay it was possible to calculate the amount of off-target template that will produce 1/2 the maximum on-target (VIC) fluorescent signal, a value that is between a C/C genotype and a T/T genotype. This value is defined in the manuscript as the half maximum positive value, KHPV, and in the present assay KHPV is 15.75±0.0721, represented as the relative fold increase in the amount of IL-28A over rs12979860. This method will be of interest to those performing genotyping on highly conserved gene families.

Immunomodulatory Function of Interleukin 28B during primary infection with cytomegalovirus.

BACKGROUND: Feedback mechanisms between interferons α and λ (IFNs) may be affected by single nucleotide polymorphisms (SNP) in interleukin 28B (IL-28B; IFN-λ3) promoter region and may influence cytomegalovirus (CMV) replication. METHODS: We associated IL-28B SNPs with the risk of CMV replication after transplantation. Next, we examined the effect of IL-28B genotypes on IL-28B, and IFN-stimulated gene (ISG) expression, and CMV replication in human foreskin fibroblast (HFF) and peripheral blood mononuclear cells (PBMCs). RESULTS: Transplant recipients with an IL-28B SNP (rs8099917) had significantly less CMV replication (P = .036). Both HFF-cells and PBMCs with a SNP showed lower IL-28B expression during infection with CMV, but higher "antiviral" ISG expression (eg, OAS1). Fibroblasts with a SNP had a 3-log reduction of CMV replication at day 4 (P = .004). IL-28B pretreatment induced ISG expression in noninfected fibroblasts, but a relative decrease of ISG expression could be observed in CMV-infected fibroblasts. The inhibitory effects of IL-28B could be abolished by siRNA or antagonistic peptides against the IL-28 receptor. In fibroblasts, inhibition of IL-28 signaling resulted in an increase of ISG expression and 3-log reduction of CMV-replication (P = .01). CONCLUSIONS: We postulate that IL-28B may act as a key regulator of ISG expression during primary CMV infection. IL-28B SNPs may be associated with higher antiviral ISG expression, which results in better replication control.

Peptide vaccination is superior to genetic vaccination using a recombineered bacteriophage λ subunit vaccine.

Genetic immunization holds promise as a vaccination method, but has so far proven ineffective in large primate and human trials. Herein, we examined the relative merits of genetic immunization and peptide immunization using bacteriophage λ. Bacteriophage λ has proven effective in immune challenge models using both immunization methods, but there has never been a direct comparison of efficacy and of the quality of immune response. In the current study, this vector was produced using a combination of cis and trans phage display. When antibody titers were measured from immunized animals together with IL-2, IL-4 and IFNγ production from splenocytes in vitro, we found that proteins displayed on λ were superior at eliciting an immune response in comparison to genetic immunization with λ. We also found that the antibodies produced in response to immunization with λ displayed proteins bound more epitopes than those produced in response to genetic immunization. Finally, the general immune response to λ inoculation, whether peptide or genetic, was dominated by a Th1 response, as determined by IFNγ and IL-4 concentration, or by a higher concentration of IgG2a antibodies.