Chaperones Ameliorate Beta Cell Dysfunction Associated with Human Islet Amyloid Polypeptide Overexpression

In type 2 diabetes, beta-cell dysfunction is thought to be due to several causes, one being the formation of toxic protein aggregates called islet amyloid, formed by accumulations of misfolded human islet amyloid polypeptide (hIAPP). The process of hIAPP misfolding and aggregation is one of the factors that may activate the unfolded protein response (UPR), perturbing endoplasmic reticulum (ER) homeostasis. Molecular chaperones have been described to be important in regulating ER response to ER stress. In the present work, we evaluate the role of chaperones in a stressed cellular model of hIAPP overexpression. A rat pancreatic beta-cell line expressing hIAPP exposed to thapsigargin or treated with high glucose and palmitic acid, both of which are known ER stress inducers, showed an increase in ER stress genes when compared to INS1E cells expressing rat IAPP or INS1E control cells. Treatment with molecular chaperone glucose-regulated protein 78 kDa (GRP78, also known as BiP) or protein disulfite isomerase (PDI), and chemical chaperones taurine-conjugated ursodeoxycholic acid (TUDCA) or 4-phenylbutyrate (PBA), alleviated ER stress and increased insulin secretion in hIAPP-expressing cells. Our results suggest that the overexpression of hIAPP induces a stronger response of ER stress markers. Moreover, endogenous and chemical chaperones are able to ameliorate induced ER stress and increase insulin secretion, suggesting that improving chaperone capacity can play an important role in improving beta-cell function in type 2 diabetes.     

A microsatellite genetic map of the turbot (Scophthalmus maximus)

A consensus microsatellite-based linkage map of the turbot (Scophthalmus maximus) was constructed from two unrelated families. The mapping panel was derived from a gynogenetic family of 96 haploid embryos and a biparental diploid family of 85 full-sib progeny with known linkage phase. A total of 242 microsatellites were mapped in 26 linkage groups, six markers remaining unlinked. The consensus map length was 1343.2 cM, with an average distance between markers of 6.5 +/- 0.5 cM. Similar length of female and male maps was evidenced. However, the mean recombination at common intervals throughout the genome revealed significant differences between sexes, approximately 1.6 times higher in the female than in the male. The comparison of turbot microsatellite flanking sequences against the Tetraodon nigroviridis genome revealed 55 significant matches, with a mean length of 102 bp and high sequence similarity (81-100%). The comparative mapping revealed significant syntenic regions among fish species. This study represents the first linkage map in the turbot, one of the most important flatfish in European aquaculture. This map will be suitable for QTL identification of productive traits in this species and for further evolutionary studies in fish and vertebrate species.     

Prognostic value of X-chromosome inactivation in symptomatic female carriers of dystrophinopathy

Background

Neonatal screening for Pompe disease has been introduced in Taiwan and a few U.S. states, while other jurisdictions including some European countries are piloting or considering this screening. First-tier screening flags both classic infantile and late-onset Pompe disease, which challenges current screening criteria. Previously, advocacy groups have sometimes supported expanded neonatal screening more than professional experts, while neutral citizens' views were unknown. This study aimed to measure support for neonatal screening for Pompe disease in the general public and to compare it to support among (parents of) patients with this condition. The study was done in the Netherlands, where newborns are not currently screened for Pompe disease. Newborn screening is not mandatory in the Netherlands but current uptake is almost universal.

Methods

A consumer panel (neutral group) and (parents of) patients with Pompe disease (Pompe group) were sent information and a questionnaire. Responses were analyzed of 555 neutral and 58 Pompe-experienced informants who had demonstrated sufficient understanding.

Results

87% of the neutral group and 88% of the Pompe group supported the introduction of screening (95% CI of difference -10 to 7%). The groups were similar in their moral reasoning about screening and acceptance of false positives, but the Pompe-experienced group expected greater benefit from neonatal detection of late-onset disease. Multivariate regression analysis controlling for demographics confirmed that approval of the introduction of screening was independent of having (a child with) Pompe disease. Furthermore, respondents with university education, regardless of whether they have (a child with) Pompe disease, were more likely to be reluctant about the introduction of screening than those with less education, OR for approval 0.29 (95% CI 0.18 to 0.49, p < 0.001).

Conclusions

This survey suggests a rather high level of support for newborn screening for Pompe disease, not only among those who have personal experience of the disease but also among the general public in the Netherlands. Optional screening on the basis of informed parental consent is probably unrealistic, underlining the need for new guidelines to help policymakers in their consideration of newborn screening for broad phenotype conditions.     

CTL responses of high functional avidity and broad variant cross-reactivity are associated with HIV control

Cytotoxic T lymphocyte (CTL) responses targeting specific HIV proteins, in particular Gag, have been associated with relative control of viral replication in vivo. However, Gag-specific CTL can also be detected in individuals who do not control the virus and it remains thus unclear how Gag-specific CTL may mediate the beneficial effects in some individuals but not in others. Here, we used a 10mer peptide set spanning HIV Gag-p24 to determine immunogen-specific T-cell responses and to assess functional properties including functional avidity and cross-reactivity in 25 HIV-1 controllers and 25 non-controllers without protective HLA class I alleles. Our data challenge the common belief that Gag-specific T cell responses dominate the virus-specific immunity exclusively in HIV-1 controllers as both groups mounted responses of comparable breadths and magnitudes against the p24 sequence. However, responses in controllers reacted to lower antigen concentrations and recognized more epitope variants than responses in non-controllers. These cross-sectional data, largely independent of particular HLA genetics and generated using direct ex-vivo samples thus identify T cell responses of high functional avidity and with broad variant reactivity as potential functional immune correlates of relative HIV control.     

Impact of Chiral Bioanalytical Methods on the Bioequivalence of Ibuprofen Products Containing Ibuprofen Lysinate and Ibuprofen Base

The purpose was to assess the impact of the use of a chiral bioanalytical method on the conclusions of a bioequivalence study that compared two ibuprofen suspensions with different rates of absorption. A comparison of the conclusion of bioequivalence between a chiral method and an achiral approach was made. Plasma concentrations of R‐ibuprofen and S‐ibuprofen were determined using a chiral bioanalytical method; bioequivalence was tested for R‐ibuprofen and for S‐ibuprofen separately and for the sum of both enantiomers as an approach for an achiral bioanalytical method. The 90% confidence interval (90% CI) that would have been obtained with an achiral bioanalytical method (90% CI: C_max: 117.69–134.46; AUC₀^t: 104.75–114.45) would have precluded the conclusion of bioequivalence. This conclusion cannot be generalized to the active enantiomer (90% CI: C_max: 103.36–118.38; AUC₀^t: 96.52–103.12), for which bioequivalence can be concluded, and/or the distomer (90% CI: C_max: 132.97–151.33; AUC₀^t: 115.91–135.77) for which a larger difference was observed. Chiral bioanalytical methods should be required when 1) the enantiomers exhibit different pharmacodynamics and 2) the exposure (AUC or C_max) ratio of enantiomers is modified by a difference in the rate of absorption. Furthermore, the bioequivalence conclusion should be based on all enantiomers, since the distomer(s) might not be completely inert, in contrast to what is required in the current regulatory guidelines. In those cases where it is unknown if the ratio between enantiomers is modified by changing the rate of absorption, chiral bioanalytical methods should be employed unless enantiomers exhibit the same pharmacodynamics. Chirality 28:429–433, 2016. © 2016 Wiley Periodicals, Inc.     

Safety, immunogenicity and duration of protection of the RTS,S/AS02(D) malaria vaccine: one year follow-up of a randomized controlled phase I/IIb trial

Background

The RTS,S/AS02_D vaccine has been shown to have a promising safety profile, to be immunogenic and to confer protection against malaria in children and infants.

Methods and Findings

We did a randomized, controlled, phase I/IIb trial of RTS,S/AS02_D given at 10, 14 and 18 weeks of age staggered with routine immunization vaccines in 214 Mozambican infants. The study was double-blind until the young child completed 6 months of follow-up over which period vaccine efficacy against new Plasmodium falciparum infections was estimated at 65.9% (95% CI 42.6–79.8, p<0.0001). We now report safety, immunogenicity and estimated efficacy against clinical malaria up to 14 months after study start. Vaccine efficacy was assessed using Cox regression models. The frequency of serious adverse events was 32.7% in the RTS,S/AS02_D and 31.8% in the control group. The geometric mean titers of anti-circumsporozoite antibodies declined from 199.9 to 7.3 EU/mL from one to 12 months post dose three of RTS,S/AS02_D, remaining 15-fold higher than in the control group. Vaccine efficacy against clinical malaria was 33% (95% CI: −4.3–56.9, p = 0.076) over 14 months of follow-up. The hazard rate of disease per 2-fold increase in anti-CS titters was reduced by 84% (95% CI 35.1–88.2, p = 0.003).

Conclusion

The RTS,S/AS02_D malaria vaccine administered to young infants has a good safety profile and remains efficacious over 14 months. A strong association between anti-CS antibodies and risk of clinical malaria has been described for the first time. The results also suggest a decrease of both anti-CS antibodies and vaccine efficacy over time.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00197028","term_id":"NCT00197028"}}NCT00197028     

Immunoregulatory Actions of Epithelial Cell PPAR γ at the Colonic Mucosa of Mice with Experimental Inflammatory Bowel Disease

Background

Peroxisome proliferator-activated receptors are nuclear receptors highly expressed in intestinal epithelial cells (IEC) and immune cells within the gut mucosa and are implicated in modulating inflammation and immune responses. The objective of this study was to investigate the effect of targeted deletion of PPAR γ in IEC on progression of experimental inflammatory bowel disease (IBD).

Methodology/Principal Findings

In the first phase, PPAR γ flfl; Villin Cre- (VC-) and PPAR γ flfl; Villin Cre+ (VC+) mice in a mixed FVB/C57BL/6 background were challenged with 2.5% dextran sodium sulfate (DSS) in drinking water for 0, 2, or 7 days. VC+ mice express a transgenic recombinase under the control of the Villin-Cre promoter that causes an IEC-specific deletion of PPAR γ. In the second phase, we generated VC- and VC+ mice in a C57BL/6 background that were challenged with 2.5% DSS. Mice were scored on disease severity both clinically and histopathologically. Flow cytometry was used to phenotypically characterize lymphocyte and macrophage populations in blood, spleen and mesenteric lymph nodes. Global gene expression analysis was profiled using Affymetrix microarrays. The IEC-specific deficiency of PPAR γ in mice with a mixed background worsened colonic inflammatory lesions, but had no effect on disease activity (DAI) or weight loss. In contrast, the IEC-specific PPAR γ null mice in C57BL/6 background exhibited more severe inflammatory lesions, DAI and weight loss in comparison to their littermates expressing PPAR γ in IEC. Global gene expression profiling revealed significantly down-regulated expression of lysosomal pathway genes and flow cytometry results demonstrated suppressed production of IL-10 by CD4+ T cells in mesenteric lymph nodes (MLN) of IEC-specific PPAR γ null mice.

Conclusions/Significance

Our results demonstrate that adequate expression of PPAR γ in IEC is required for the regulation of mucosal immune responses and prevention of experimental IBD, possibly by modulation of lysosomal and antigen presentation pathways.