ATF6 is essential for human cone photoreceptor development

Endoplasmic reticulum (ER) stress and Unfolded Protein Response (UPR) signaling promote the pathology of many human diseases. Loss-of-function variants of the UPR regulator Activating Transcription Factor 6 (ATF6) cause severe congenital vision loss diseases such as achromatopsia by unclear pathomechanisms. To investigate this, we generated retinal organoids from achromatopsia patient induced pluripotent stem cells carrying ATF6 disease variants and from gene-edited ATF6 null hESCs. We found that achromatopsia patient and ATF6 null retinal organoids failed to form cone structures concomitant with loss of cone phototransduction gene expression, while rod photoreceptors developed normally. Adaptive optics retinal imaging of achromatopsia patients carrying ATF6 variants also showed absence of cone inner/outer segment structures but preserved rod structures, mirroring the defect in cone formation observed in our retinal organoids. These results establish that ATF6 is essential for human cone development. Interestingly, we find that a selective small molecule ATF6 signaling agonist restores the transcriptional activity of some ATF6 disease-causing variants and stimulates cone growth and gene expression in patient retinal organoids carrying these variants. These findings support that pharmacologic targeting of the ATF6 pathway can promote human cone development and should be further explored for blinding retinal diseases

Unforeseen Costs of Cutting Mosquito Surveillance Budgets

A budget proposal to stop the U.S. Centers for Disease Control and Prevention (CDC) funding in surveillance and research for mosquito-borne diseases such as dengue and West Nile virus has the potential to leave the country ill-prepared to handle new emerging diseases and manage existing ones. In order to demonstrate the consequences of such a measure, if implemented, we evaluated the impact of delayed control responses to dengue epidemics (a likely scenario emerging from the proposed CDC budget cut) in an economically developed urban environment. We used a mathematical model to generate hypothetical scenarios of delayed response to a dengue introduction (a consequence of halted mosquito surveillance) in the City of Cairns, Queensland, Australia. We then coupled the results of such a model with mosquito surveillance and case management costs to estimate the cumulative costs of each response scenario. Our study shows that halting mosquito surveillance can increase the management costs of epidemics by up to an order of magnitude in comparison to a strategy with sustained surveillance and early case detection. Our analysis shows that the total costs of preparedness through surveillance are far lower than the ones needed to respond to the introduction of vector-borne pathogens, even without consideration of the cost in human lives and well-being. More specifically, our findings provide a science-based justification for the re-assessment of the current proposal to slash the budget of the CDC vector-borne diseases program, and emphasize the need for improved and sustainable systems for vector-borne disease surveillance

Searching the protein structure database for ligand-binding site similarities using CPASS v.2

<p>Abstract</p> <p>Background</p> <p>A recent analysis of protein sequences deposited in the NCBI RefSeq database indicates that ~8.5 million protein sequences are encoded in prokaryotic and eukaryotic genomes, where ~30% are explicitly annotated as "hypothetical" or "uncharacterized" protein. Our Comparison of Protein Active-Site Structures (CPASS v.2) database and software compares the sequence and structural characteristics of experimentally determined ligand binding sites to infer a functional relationship in the absence of global sequence or structure similarity. CPASS is an important component of our Functional Annotation Screening Technology by NMR (FAST-NMR) protocol and has been successfully applied to aid the annotation of a number of proteins of unknown function.</p> <p>Findings</p> <p>We report a major upgrade to our CPASS software and database that significantly improves its broad utility. CPASS v.2 is designed with a layered architecture to increase flexibility and portability that also enables job distribution over the Open Science Grid (OSG) to increase speed. Similarly, the CPASS interface was enhanced to provide more user flexibility in submitting a CPASS query. CPASS v.2 now allows for both automatic and manual definition of ligand-binding sites and permits pair-wise, one versus all, one versus list, or list versus list comparisons. Solvent accessible surface area, ligand root-mean square difference, and Cβ distances have been incorporated into the CPASS similarity function to improve the quality of the results. The CPASS database has also been updated.</p> <p>Conclusions</p> <p>CPASS v.2 is more than an order of magnitude faster than the original implementation, and allows for multiple simultaneous job submissions. Similarly, the CPASS database of ligand-defined binding sites has increased in size by ~ 38%, dramatically increasing the likelihood of a positive search result. The modification to the CPASS similarity function is effective in reducing CPASS similarity scores for false positives by ~30%, while leaving true positives unaffected. Importantly, receiver operating characteristics (ROC) curves demonstrate the high correlation between CPASS similarity scores and an accurate functional assignment. As indicated by distribution curves, scores ≥ 30% infer a functional similarity. Software URL: <url>http://cpass.unl.edu</url>.</p

Soluble fms-Like Tyrosine Kinase 1 (sFlt1), Endoglin and Placental Growth Factor (PlGF) in Preeclampsia among High Risk Pregnancies

Background: Differences in circulating concentrations of antiangiogenic factors sFlt1 and soluble endoglin (sEng) and the pro-angiogenic growth factor PlGF are reported to precede the onset of preeclampsia weeks to months in low-risk pregnant women. The objective of this study was to investigate whether similar changes can be detected in pregnant women at high-risk to develop the syndrome. Methods: This study is a secondary analysis of the NICHD MFMU trial of aspirin to prevent preeclampsia in high-risk pregnancies. Serum samples were available from 194 women with pre-existing diabetes, 313 with chronic hypertension, 234 with multifetal gestation, and 252 with a history of preeclampsia in a previous pregnancy. Samples collected across pregnancy were analyzed in a blinded fashion for sFlt1, sEng and PlGF. Results: The odds of developing preeclampsia were significantly increased among women with multiple fetuses for each 2- fold elevation in sFlt1, sEng and the ratio of angiogenic factors (e.g. OR 2.18, 95% CI 1.46-3.32), and significantly decreased for each 2-fold elevation in circulating PlGF (OR 0.50, 95% CI 0.30-0.82) between 7 and 26 weeks' gestation. Cross-sectional analysis of the angiogenic factors across gestation showed significant differences during the third trimester in women who develop preeclampsia compared with appropriate controls in all high-risk groups. However, when data were examined in relation to the gestational week when preeclampsia was diagnosed only sFlt1 was significantly higher 2 to 5 weeks before the clinical onset of preeclampsia and only in women with previous preeclampsia. Conclusions: The pattern of elevated concentrations of sFlt1 and sEng, and low PlGF in high-risk pregnant subjects who develop preeclampsia is similar to that reported in low-risk pregnant women. However, differences in these factors among high-risk women who do and do not develop preeclampsia are modest, and do not appear to be clinically useful predictors in these high-risk pregnant women

Comparison of an ordinal endpoint to time-to-event, longitudinal, and binary endpoints for use in evaluating treatments for severe influenza requiring hospitalization

Background/aims: The Food and Drug Administration recommends research into developing well-defined and reliable endpoints to evaluate treatments for severe influenza requiring hospitalization. A novel 6-category ordinal endpoint of patient health status after 7 days that ranges from death to hospital discharge with resumption of normal activities is being used in a randomized placebo-controlled trial of intravenous immunoglobulin (IVIG) for severe influenza (FLU-IVIG). We compare the power of the ordinal endpoint under a proportional odds model to other types of endpoints as a function of various trial parameters. // Methods: We used closed-form analysis and empirical simulation to compare the power of the ordinal endpoint to time-to-event, longitudinal, and binary endpoints. In the simulation setting, we varied the treatment effect and the distribution of the placebo group across the follow-up period with consideration of adjustment for baseline health status. // Results: In the analytic setting, ordinal endpoints of high granularity provided greater power than time-to-event endpoints when most patients in the placebo group had either naturally progressed to the category of hospital discharge by day 7 or were far from hospital discharge on day 7. In the simulation setting, adjustment for baseline health status universally raised power for the proportional odds model. Across different placebo group distributions of the ordinal endpoint regardless of adjustment for baseline health status, only time-to-event endpoints yielded higher power than the ordinal endpoint for certain treatment effects. // Conclusions: In this case study, the FLU-IVIG ordinal endpoint provided greater power than time-to-event, binary, and longitudinal endpoints for most scenarios of the treatment effect and placebo group distribution, including the target population studied for FLU-IVIG. The ordinal endpoint was only surpassed by the time-to-event endpoint when many patients in the placebo group were on the cusp of hospital discharge on day 7 and the follow-up period for the time-to-event endpoint was extended to allow for additional events. Our general approach for evaluating the power of several potential endpoints for an influenza trial can be used for designing other influenza trials with different target populations and for other trials in other disease areas

Prior exercise and antioxidant supplementation: effect on oxidative stress and muscle injury

<p>Abstract</p> <p>Background</p> <p>Both acute bouts of prior exercise (preconditioning) and antioxidant nutrients have been used in an attempt to attenuate muscle injury or oxidative stress in response to resistance exercise. However, most studies have focused on untrained participants rather than on athletes. The purpose of this work was to determine the independent and combined effects of antioxidant supplementation (vitamin C + mixed tocopherols/tocotrienols) and prior eccentric exercise in attenuating markers of skeletal muscle injury and oxidative stress in resistance trained men.</p> <p>Methods</p> <p>Thirty-six men were randomly assigned to: no prior exercise + placebo; no prior exercise + antioxidant; prior exercise + placebo; prior exercise + antioxidant. Markers of muscle/cell injury (muscle performance, muscle soreness, C-reactive protein, and creatine kinase activity), as well as oxidative stress (blood protein carbonyls and peroxides), were measured before and through 48 hours of exercise recovery.</p> <p>Results</p> <p>No group by time interactions were noted for any variable (P > 0.05). Time main effects were noted for creatine kinase activity, muscle soreness, maximal isometric force and peak velocity (P < 0.0001). Protein carbonyls and peroxides were relatively unaffected by exercise.</p> <p>Conclusion</p> <p>There appears to be no independent or combined effect of a prior bout of eccentric exercise or antioxidant supplementation as used here on markers of muscle injury in resistance trained men. Moreover, eccentric exercise as used in the present study results in minimal blood oxidative stress in resistance trained men. Hence, antioxidant supplementation for the purpose of minimizing blood oxidative stress in relation to eccentric exercise appears unnecessary in this population.</p

A Fetal-Maternal Shift of Blood Oxygen Affinity in an Australian Viviparous Lizard, Sphenomorphus quoyii (Reptilia, Scincidae)

Compared to adults, the oxygen affinity of blood from fetal Sphenomorphus quoyii is very much higher: P50 is approximately 70 Torr in adults and 30 Torr in nearly full term embryos (PC02= 17 Torr, T=34 degrees C). Following birth, oxygen affinity decreases gradually and adult values are approached after about 15 weeks, with the onset of winter retreat. Electrophoresis revealed a multiple hemoglobin system in both adults and embryos, but there were no apparent differences between them

Characterization of a protozoan Phosducin-like protein-3 (PhLP-3) reveals conserved redox activity

We recently identified three novel thioredoxin-like genes in the genome of the protozoan parasite Plasmodium that belong to the Phosducin-like family of proteins (PhLP). PhLPs are small cytosolic proteins hypothesized to function in G-protein signaling and protein folding. Although PhLPs are highly conserved in eukaryotes from yeast to mammals, only a few representatives have been experimentally characterized to date. In addition, while PhLPs contain a thioredoxin domain, they lack a CXXC motif, a strong indicator for redox activity, and it is unclear whether members of the PhLP family are enzymatically active. Here, we describe PbPhLP-3 as the first phosducin-like protein of a protozoan organism, Plasmodium berghei. Initial transcription analysis revealed continuous low-level expression of pbphlp-3 throughout the complex Plasmodium life cycle. Attempts to knockout pbphlp-3 in P. berghei did not yield live parasites, suggesting an essential role for the gene in Plasmodium. We cloned, expressed and purified PbPhLP-3 and determined that the recombinant protein is redox active in vitro in a thioredoxin-coupled redox assay. It also has the capacity to reduce the organic compound tert-Butyl hydroperoxide (TBHP) in vitro, albeit at low efficiency. Sequence analysis, structural modeling, and site-directed mutagenesis revealed a conserved cysteine in the thioredoxin domain to be the redox active residue. Lastly, we provide evidence that recombinant human PhLP-3 exhibits redox activity similar to that of PbPhLP-3 and suggest that redox activity may be conserved in PhLP-3 homologs of other species. Our data provide new insight into the function of PhLP-3, which is hypothesized to act as co-chaperones in the folding and regulation of cytoskeletal proteins. We discuss the potential implications of PhLP-3 as a thioredoxin-target protein and possible links between the cellular redox network and the eukaryotic protein folding machinery