A new epidermal ciliary photoreceptive cell type in Plagiostomum lemani (Plathelminthes, Prolecithophora)

 Plagiostomum lemani possesses extremely specialized intraepidermal sensory cells. These obvious photoreceptors, which are not visible with the light microscope, are ciliary aggregations located in an intracellular cavity. The numerous spiralled cilia have the classic 9 × 2 + 2 arrangement at their base and a modified pattern of microtubules apically. The discovered differentiations do not show a connection to the surface. Neither mantle cells nor pigment cells have been found. The structural similarities with other epidermal photoreceptors of species among the different taxa of free-living Plathelminthes are outlined. Besides the larval stages of the taxon Polycladida known so far, the same kind of light-sensing photoreceptive cell has never been described in any other species of the Plathelminthes. Accepted: 16 November 1997     

Clinical Features of Reported Ethylene Glycol Exposures in the United States

Background

Ethylene glycol is highly toxic and represents an important cause of poisonings worldwide. Toxicity can result in central nervous system dysfunction, cardiovascular compromise, elevated anion gap metabolic acidosis and acute kidney injury. Many states have passed laws requiring addition of the bittering agent, denatonium benzoate, to ethylene glycol solutions to reduce severity of exposures. The objectives of this study were to identify differences between unintentional and intentional exposures and to evaluate the utility of denatonium benzoate as a deterrent.

Methods and Findings

Using the National Poison Data System, we performed a retrospective analysis of reported cases of ethylene glycol exposures from January 2006 to December 2013. Outcome classification was summed for intentionality and used as a basis for comparison of effect groups. There were 45,097 cases of ethylene glycol exposures resulting in 154 deaths. Individuals more likely to experience major effects or death were older, male, and presented with more severe symptoms requiring higher levels of care. Latitude and season did not correlate with increased exposures; however, there were more exposures in rural areas. Denatonium benzoate use appeared to have no effect on exposure severity or number.

Conclusion

Deaths due to ethylene glycol exposure were uncommon; however, there were major clinical effects and more exposures in rural areas. Addition of denatonium benzoate was not associated with a reduction in exposures. Alternative means to deter ingestion are needed. These findings suggest the need to consider replacing ethylene glycol with alternative and less toxic agents.     

Serological Evidence of Chikungunya Virus among Acute Febrile Patients in Southern Mozambique

Background

In the last two decades, chikungunya virus (CHIKV) has rapidly expanded to several geographical areas, causing frequent outbreaks in sub-Saharan Africa, South East Asia, South America, and Europe. Therefore, the disease remains heavily neglected in Mozambique, and no recent study has been conducted.

Methods

Between January and September 2013, acute febrile patients with no other evident cause of fever and attending a health center in a suburban area of Maputo city, Mozambique, were consecutively invited to participate. Paired acute and convalescent serum samples were requested from each participant. Convalescent samples were initially screened for anti-CHIKV IgG using a commercial indirect immunofluorescence test, and if positive, the corresponding acute sample was screened using the same test.

Results

Four hundred patients were enrolled. The median age of study participants was 26 years (IQR: 21–33 years) and 57.5% (224/391) were female. Paired blood samples were obtained from 209 patients, of which 26.4% (55/208) were presented anti-CHIKV IgG antibodies in the convalescent sample. Seroconversion or a four-fold titer rise was confirmed in 9 (4.3%) patients.

Conclusion

The results of this study strongly suggest that CHIKV is circulating in southern Mozambique. We recommend that CHIKV should be considered in the differential diagnosis of acute febrile illness in Mozambique and that systematic surveillance for CHIKV should be implemented.     

The Calcium-Dependent Interaction between S100B and the Mitochondrial AAA ATPase ATAD3A and the Role of This Complex in the Cytoplasmic Processing of ATAD3A

S100 proteins comprise a multigene family of EF-hand calcium binding proteins that engage in multiple functions in response to cellular stress. In one case, the S100B protein has been implicated in oligodendrocyte progenitor cell (OPC) regeneration in response to demyelinating insult. In this example, we report that the mitochondrial ATAD3A protein is a major, high-affinity, and calcium-dependent S100B target protein in OPC. In OPC, ATAD3A is required for cell growth and differentiation. Molecular characterization of the S100B binding domain on ATAD3A by nuclear magnetic resonance (NMR) spectroscopy techniques defined a consensus calcium-dependent S100B binding motif. This S100B binding motif is conserved in several other S100B target proteins, including the p53 protein. Cellular studies using a truncated ATAD3A mutant that is deficient for mitochondrial import revealed that S100B prevents cytoplasmic ATAD3A mutant aggregation and restored its mitochondrial localization. With these results in mind, we propose that S100B could assist the newly synthesized ATAD3A protein, which harbors the consensus S100B binding domain for proper folding and subcellular localization. Such a function for S100B might also help to explain the rescue of nuclear translocation and activation of the temperature-sensitive p53val135 mutant by S100B at nonpermissive temperatures.The S100 proteins comprise a multigene family of low-molecular-weight EF-hand calcium binding and zinc binding proteins (5, 13, 16, 24, 33). To date, 19 different S100 proteins have been assigned to this protein family, and they show different degrees of similarity, ranging from 25 to 56% identity at the amino acid level. With S100B, S100P, and S100Z being the exceptions, the majority of the S100 genes are clustered on human chromosome 1q21 (33). Most S100 proteins serve as calcium sensor proteins that, upon activation, regulate the function and/or subcellular distribution of specific target proteins (13, 33, 47), and they are characterized by common structural motifs, including two low-affinity (K_D [equilibrium dissociation constant] of ∼10 μM to 100 μM) helix-loop-helix calcium binding domains (EF hands) that are separated by a hinge region and flanked by amino- and carboxy-terminal domains. The carboxy-terminal domain is variable among S100 proteins, and it typically is the site that is responsible for the selective interaction of each individual S100 protein with specific target proteins (30). S100 proteins are often upregulated in cancers, in inflammation, and in response to cellular stress (14, 16), suggesting that they function in cell responses to stress situations. Consistent with this hypothesis, stress situations were necessary to reveal phenotypes associated with the S100 knockout in mice (11, 14, 33, 56). Moreover, recent observations revealed a new function for the S100 protein family that included their ability to assist and regulate multichaperone complex-ligand interactions (41, 50, 51).One member of the S100 protein family, S100B, has attracted much interest in the past few years because, like other proteins implicated in neurodegeneration (e.g., amyloid, superoxide dismutase, and dual-specificity tyrosine phosphorylation-regulated kinase 1A), its gene is located within a segment of chromosome 21, which is trisomic in Down''s syndrome (DS). Its expression in the brain of mammals coincides with defined periods of central nervous system (CNS) maturation and cell differentiation (43). In oligodendrocyte progenitor cells (OPC), S100B expression is associated with differentiation, and S100B contributes to OPC differentiation in response to demyelinating insult (11). To understand the contribution of S100B to OPC differentiation, we searched for high-affinity S100B target proteins in this cell type by using far-Western analysis. A major and highly specific S100B target protein was identified, the mitochondrial ATAD3A protein.ATAD3A belongs to a new family of eukaryote-specific mitochondrial AAA⁺ ATPase proteins (17). In the human genome, two genes, Atad3A and Atad3B, are located in tandem on chromosome 1p36.33. The Atad3A gene is ubiquitous among multicellular organisms but absent in yeast. The Atad3B gene is specific to the human genome (27). ATAD3A is a mitochondrial protein anchored into the mitochondrial inner membrane (IM) at contact sites with the outer membrane (OM). Thanks to its simultaneous interaction with the two membranes, ATAD3A regulates mitochondrial dynamics at the interface between the inner and outer membranes and controls diverse cell responses ranging from mitochondrial metabolism, cell growth, and mitochondrial fission 20a, 25). The ATAD3A protein has also been identified as a mitochondrial DNA binding protein (23) and as a cell surface antigen in some human tumors (20, 21). The plasma membrane localization of ATAD3A in tumor cells is suggestive that ATAD3A mitochondrial routing can be compromised in pathological situations such as cancer. To understand the functional response resulting from the interaction between S100B and ATAD3A, we first characterized the minimal interaction domain on ATAD3A for S100B binding using thermodynamic studies of wild-type and ATAD3A variants as well as via nuclear magnetic resonance (NMR) spectroscopy techniques. These studies allowed us to further refine the consensus S100B binding motif, which is conserved in several other S100B target proteins, including the p53 protein and several newly discovered target proteins associated with the cell translational machinery. We next analyzed the cellular interaction of S100B with truncated ATAD3A mutants that harbor the S100B binding domain but that are deficient for mitochondrial import. These studies revealed that S100B could assist ATAD3A mutant proteins during cytoplasmic processing by preventing dysfunctional aggregation events. Our results are discussed in light of the possible function of S100B in assisting the cytoplasmic processing of proteins for proper folding and subcellular localization.     

Grip force variability and its effects on children's handwriting legibility, form, and strokes

A comprehensive understanding of the underlying biomechanical processes during handwriting is needed to accurately guide clinical interventions. To date, quantitative measurement of such biomechanical processes has largely excluded measurements of the forces exerted radially on the barrel of the writing utensil (grip forces) and how they vary over time during a handwriting task. An instrumented writing utensil was deployed for a direct measurement of kinematic and temporal information during a writing task, as well as forces exerted on the writing surface and on the barrel of the pen. The writing utensil was used by a cohort of 35 students (19 males), 16 in first grade and 19 in second grade, as they performed the Minnesota Handwriting Assessment (MHA) test. Quantitative grip force variability measures were computed and tested as correlates of handwriting legibility, form, and strokes. Grip force variability was shown to correlate strongly with handwriting quality, in particular for students classified by the MHA as nonproficient writers. More specifically, static grip force patterns were shown to result in poor handwriting quality and in greater variation in handwriting stroke durations. Grip force variability throughout the writing task was shown to be significantly lower for nonproficient writers (t-test, p<0.01) while the number of strokes and per-stroke durations were shown to be higher (p<0.03). The results suggest that grip force dynamics play a key role in determining handwriting quality and stroke characteristics. In particular, students with writing difficulties exhibited more static grip force patterns, lower legibility and form scores, as well as increased variation in stroke durations. These findings shed light on the underlying processes of handwriting and grip force modulation and may help to improve intervention planning.     

Application of Modeling to Scale-up Dissolution in Pharmaceutical Manufacturing

Liquid mixing scale-up in pharmaceutical industry has often been based on empirical approach in spite of tremendous understanding of liquid mixing scale-up in engineering fields. In this work, we attempt to provide a model-based approach to scale-up dissolution process from a 2 l lab-scale vessel to a 4,000 l scale vessel used in manufacturing. Propylparaben was used as a model compound to verify the model predictions for operating conditions at commercial scale that would result in similar dissolution profile as observed in lab scale. Geometric similarity was maintained between both of the scales to ensure similar mixing characteristics. We utilized computational fluid dynamics (CFD) to ensure that the operating conditions at laboratory and commercial scale will result in similar power per unit volume (P/V). Utilizing this simple scale-up criterion of similar P/V across different scales, results obtained indicate fairly good reproducibility of the dissolution profiles between the two scales. Utilization of concepts of design of experiments enabled summarizing scale-up results in statistically meaningful parameters, for example −90% dissolution in lab scale at a given time under certain operating conditions will result in 75–88% at commercial scale with 95% confidence interval when P/V is maintained constant across the two scales. In this work, we have successfully demonstrated that scale-up of solid dissolution can be done using a systematic process of lab-scale experiments followed by simple CFD modeling to predict commercial-scale experimental conditions.     

Synthesis of linear and comb-like peptide constructs containing up to four copies of a T cell epitope and their capacity to stimulate T cells.

Polypeptide constructs containing up to four copies of the T cell epitope 306-318 of influenza virus haemagglutinin have been synthesized on solid phase. Between the copies, a non-natural PEG-based spacer amino acid has been introduced. The oligomeric epitopes were analysed by RP-HPLC and ES-MS. The arrangement of the epitopes within the peptide constructs was either linear or comb-like. The proliferative response in a T helper cell assay induced by these oligomerized epitopes has been tested, showing that the linearly arranged epitopes are more effective than the comb-like oligomers.     

Synthesis of structured phospholipids by immobilized phospholipase A2 catalyzed acidolysis

Acyl modification of the sn-2 position in phospholipids (PLs) was conducted by acidolysis reaction using immobilized phospholipase A(2) (PLA(2)) as the catalyst. In the first stage we screened different carriers for their ability to immobilize PLA(2). Several carriers were able to fix the enzyme and maintain catalytic activity; however, the final choice of carrier for the continued work was a non-ionic weakly polar macroreticular resin. Response surface methodology was applied to evaluate the influence of substrate ratio, reaction temperature, and water addition during acidolysis reaction between caprylic acid and soybean phosphatidylcholine (PC). Reaction temperature and water addition had significant effect on acidolysis reaction, however no effect was observed for substrate ratio (mol caprylic acid/mol PC) in range tested. In general an inverse relationship between incorporation of caprylic acid and PC recovery was observed. Highest incorporation obtained during acidolysis reactions was 36%. Such incorporation could be obtained under reaction temperature, 45 degrees C; substrate ratio, 9mol/mol caprylic acid/PC; water addition of 2%; 30wt.% immobilized enzyme; and reaction time, 48h. The yield under these conditions was however only 29%. Lysophosphatidylcholine (LPC) was the major by-product formed during the reaction. Incorporation of acyl donor into LPC was very low (<4%), which indicates that acyl migration is only a minor problem for PLA(2) catalyzed synthesis reaction. Conjugated linoleic acid and docosahexaenoic acid were also tested as acyl donors, and were able to be incorporated into PC with 30 and 20%, respectively.