Presentation of available CTL epitopes that induction of cell-mediated immune response against HIV-1 Koran clade B strain using computational technology.

OBJECTIVE: Theoretical predicting cytotoxic T lymphocyte (CTL) epitopes are an important tool in vaccine design and CTL therapy for enhancing our understanding of the cellular immune system. We would like to identify available CTL epitopes against HIV-1 Korean clade B. CTL activity was assessed in freshly isolated peripheral blood mononuclear cells from Korean HIV patients in order to assess whether these CTL epitopes induce a cell-mediated immune response (CMI). METHODS: NetCTLpan1.1 software, which is the most popular prediction computer software package, and full atom-based simulation (FABS), which is a 3D modelling system for binding activity between epitopes and human leucocyte antigen (HLA) molecules, were used to predict the peptide-spanning Env region binding to HLA-A*24:02, HLA-A*02:01 and HLA-B*15:01, which are frequently found in the Korean population. Granzyme B and interferon-γ ELISPOT assays were used to determine whether identified CTL epitopes induce CMI. RESULTS: Three HIV-1 Korean clade B-specific Env CTL epitopes were identified: Gp41-RYL and Gp41-RQG are localized within gp41, and Gp120-LLQ within gp120. In in vitro assays using granzyme B ELISPOT, Gp120-LLQ and Gp41-RQG induced epitope-specific CTL responses in HLA-restricted cells. In ex vivo assay using IFN-γ ELISPOT, cell-mediated immune responses to Gp41-RYL were present in 50% of HLA-matched patients, and responses to Gp120-LLQ and Gp41-RQG were found in 33% of HLA-matched patients. CONCLUSION: In this study, we found that a prediction pipeline for CTL epitopes might be based on the most popular computer prediction software and FABS methods. Our results suggest that these CTL epitopes may provide useful tools and information for the development of a therapeutic vaccine against HIV-1 Korean clade B.

Spatial propagation of protein polymerization.

We consider the spatial dependence of filamentous protein self-assembly. Through studying the cases where the spreading of aggregated material is dominated either by diffusion or by growth, we derive analytical results for the spatial evolution of filamentous protein aggregation, which we validate against Monte Carlo simulations. Moreover, we compare the predictions of our theory with experimental measurements of two systems for which we identify the propagation as either growth or diffusion controlled. Our results connect the macroscopic observables that characterize the spatial propagation of protein self-assembly with the underlying microscopic processes and provide physical limits on spatial propagation and prionlike behavior associated with protein aggregation.

The effects of outdoor air supply rate on work performance during 8-h work period.

UNLABELLED: The purpose of this study is to evaluate the effect of ventilation rate on work performance and perceived air quality through short-term laboratory experiments. The experiment was designed to simulate office work, and a laboratory space was modified using new finish materials to become a typical office space. High levels of volatile organic compounds (VOCs) were found in the exposure chamber, most probably originating from the new finishing materials that were present. Twenty-four subjects were divided into six groups that were randomly exposed to the three ventilation rates, 5, 10, and 20 l/s per person. The subjects performed work tasks three separate times for each ventilation rate over an 8-h exposure period. The work performance of the subjects logarithmically improved with increased ventilation rates, which was similar to the previous research findings. Statistical significance was found for addition task, text-typing task, and memorization task. Increased work performance in this experiment was slightly lower than the results of previous short-term laboratory experiments, yet remained higher than results of previous long-term field experiments. However, it was difficult to directly compare the results of this experiment with those of previous experiments, because of the higher concentration of VOC present in the office rooms and the learning effect associated with the repeated tasks. PRACTICAL IMPLICATIONS: The results of this experiment show that ventilation had positive impacts on perceived air quality and work performance for the subjects tested. Work performance logarithmically increased by approximately 2.5-5% as ventilation rates were increased from 5 to 20 l/s per person. The positive effect of ventilation rate on work performance was shown to be limited at the low ventilation rate. The positive effects on work performance were at lower ventilation rates. The learning effect in repeated work performance tasks could increase the uncertainty of the work performance analysis in 8-h exposure period.

Comparison of saline wastewater treatment performance of SBR with repeated starvation under aerobic and non-aerobic conditions.

This study investigated the effects of repeated starvation and feeding on the performance of a sequencing batch reactor (SBR) used for treating saline wastewater. The effects of aerobic and non-aerobic conditions on the sludge during starvation were evaluated to recover the performance of the SBR in terms of floc size and pollutant removal after resuming wastewater feeding. The floc size, fractal dimension, sludge volume index (SVI), specific oxygen uptake rate (SOUR), and pollutant removal efficiency were monitored. Experiment results revealed that the floc size and fractal dimensions decreased during starvation under both aerobic and non-aerobic conditions and increased after re-feeding wastewater. However, the difference in floc physical characteristics and performance depended on the starvation condition and was pronounced as starvation and re-feeding were repeated. The floc size and fractal dimensions decreased from 152.7 to 72.2 and 1.98 to 1.79 at the end of the fourth starvation period, resulting in deterioration of the sludge settleability and effluent quality. On the other hand, the floc size and fractal dimensions decreased from 158.7 to 135.7 and 1.95 to 1.81 at the end of the fourth starvation period but remained relatively constant after sludge adaptation. Some correlations were observed between the parameters monitored in this study. The results showed that maintaining the sludge under non-aerobic conditions was an effective strategy for reducing the effects of repeated starvation.

Absolute contrast estimation for soft X-ray photon fluctuation spectroscopy using a variational droplet model.

X-ray photon fluctuation spectroscopy using a two-pulse mode at the Linac Coherent Light Source has great potential for the study of quantum fluctuations in materials as it allows for exploration of low-energy physics. However, the complexity of the data analysis and interpretation still prevent recovering real-time results during an experiment, and can even complicate post-analysis processes. This is particularly true for high-spatial resolution applications using CCDs with small pixels, which can decrease the photon mapping accuracy resulting from the large electron cloud generation at the detector. Droplet algorithms endeavor to restore accurate photon maps, but the results can be altered by their hyper-parameters. We present numerical modeling tools through extensive simulations that mimic previous x-ray photon fluctuation spectroscopy experiments. By modification of a fast droplet algorithm, our results demonstrate how to optimize the precise parameters that lift the intrinsic counting degeneracy impeding accuracy in extracting the speckle contrast. These results allow for an absolute determination of the summed contrast from multi-pulse x-ray speckle diffraction, the modus operandi by which the correlation time for spontaneous fluctuations can be measured.

In cellulo crystallization of Trypanosoma brucei IMP dehydrogenase enables the identification of genuine co-factors.

Sleeping sickness is a fatal disease caused by the protozoan parasite Trypanosoma brucei (Tb). Inosine-5'-monophosphate dehydrogenase (IMPDH) has been proposed as a potential drug target, since it maintains the balance between guanylate deoxynucleotide and ribonucleotide levels that is pivotal for the parasite. Here we report the structure of TbIMPDH at room temperature utilizing free-electron laser radiation on crystals grown in living insect cells. The 2.80 Å resolution structure reveals the presence of ATP and GMP at the canonical sites of the Bateman domains, the latter in a so far unknown coordination mode. Consistent with previously reported IMPDH complexes harboring guanosine nucleotides at the second canonical site, TbIMPDH forms a compact oligomer structure, supporting a nucleotide-controlled conformational switch that allosterically modulates the catalytic activity. The oligomeric TbIMPDH structure we present here reveals the potential of in cellulo crystallization to identify genuine allosteric co-factors from a natural reservoir of specific compounds.

Similarity of sli-1, a regulator of vulval development in C. elegans, to the mammalian proto-oncogene c-cbl.

Vulval induction during Caenorhabditis elegans development is mediated by LET-23, a homolog of the mammalian epidermal growth factor receptor tyrosine kinase. The sli-1 gene is a negative regulator of LET-23 and is shown here to encode a protein similar to c-Cbl, a mammalian proto-oncoprotein. SLI-1 and c-Cbl share approximately 55 percent amino acid identity over a stretch of 390 residues, which includes a C3HC4 zinc-binding motif known as the RING finger, and multiple consensus binding sites for Src homology 3 (SH3) domains. SLI-1 and c-Cbl may define a new class of proteins that modify receptor tyrosine kinase-mediated signal transduction.

Requirements of multiple domains of SLI-1, a Caenorhabditis elegans homologue of c-Cbl, and an inhibitory tyrosine in LET-23 in regulating vulval differentiation.

SLI-1, a Caenorhabditis elegans homologue of the proto-oncogene product c-Cbl, is a negative regulator of LET-23-mediated vulval differentiation. Lack of SLI-1 activity can compensate for decreased function of the LET-23 epidermal growth factor receptor, the SEM-5 adaptor, but not the LET-60 RAS, suggesting that SLI-1 acts before RAS activation. SLI-1 and c-Cbl comprise an N-terminal region (termed SLI-1:N/Cbl-N, containing a four-helix bundle, an EF hand calcium-binding domain, and a divergent SH2 domain) followed by a RING finger domain and a proline-rich C-terminus. In a transgenic functional assay, the proline-rich C-terminal domain is not essential for sli-1(+) function. A protein lacking the SH2 and RING finger domains has no activity, but a chimeric protein with the SH2 and RING finger domains of SLI-1 replaced by the equivalent domains of c-Cbl has activity. The RING finger domain of c-Cbl has been shown recently to enhance ubiquitination of active RTKs by acting as an E3 ubiquitin-protein ligase. We find that the RING finger domain of SLI-1 is partially dispensable. Further, we identify an inhibitory tyrosine of LET-23 requiring sli-1(+) for its effects: removal of this tyrosine closely mimics the loss of sli-1 but not of another negative regulator, ark-1. Thus, we suggest that this inhibitory tyrosine mediates its effects through SLI-1, which in turn inhibits signaling upstream of LET-60 RAS in a manner not wholly dependent on the ubiquitin-ligase domain.

Data systems for the Linac coherent light source.

The data systems for X-ray free-electron laser (FEL) experiments at the Linac coherent light source (LCLS) are described. These systems are designed to acquire and to reliably transport shot-by-shot data at a peak throughput of 5 GB/s to the offline data storage where experimental data and the relevant metadata are archived and made available for user analysis. The analysis and monitoring implementation (AMI) and Photon Science ANAlysis (psana) software packages are described. Psana is open source and freely available.

Correlation of sonographic severity with biochemical markers of synovium and cartilage in knee osteoarthritis patients.

OBJECTIVE: Ultrasonography can be used to detect soft tissue abnormalities within the joints that cannot be assessed using conventional X-rays. This study investigated the relationship between soft tissue and/or bony abnormalities on ultrasonography and the biochemical markers of the synovium and cartilage in the knee of osteoarthritis (OA) patients. METHODS: The knees from 51 OA patients who fulfilled the ACR criteria were enrolled in this study. Knee ultrasonography was performed in the affected knee joints using a 12 MHz linear probe to assess the presence of effusion, synovial proliferation, capsular distention, the length of osteophytes and the cartilage thickness. At the same time, the serum hyaluronic acid (HA) and the cartilage oligomeric protein (COMP) levels were measured by ELISA, and RIA was used to determine the serum osteocalcin levels. RESULTS: The patients with a longer medial osteophyte showed higher serum HA and COMP levels than those with a shorter one. The serum HA levels were significantly higher in those patients with a larger amount of effusion and/or synovial proliferation, which indicated inflammatory changes, than in those without. In addition, the severity of the capsular distention also correlated well with the serum HA and COMP levels. However, the length of the lateral osteophytes and the thickness of the femoral cartilage showed no correlation with the serum HA or COMP levels. In addition, the serum osteocalcin levels did not show any association with the above ultrasonographic parameters. CONCLUSION: This study demonstrated that the serum HA and COMP levels were elevated in the more severe OA patients by knee ultrasonography than in the less severe patients. This suggests that the detailed pathological changes in the soft tissue and/or bone of the OA joints on ultrasonography are directly reflected by the biochemical markers measured in the peripheral blood.

Effects of addition sequence and rapid mixing conditions on use of dual coagulants.

The performance of dual coagulants in clay suspension was investigated in this study using aluminium chloride and the cationic polymer as coagulants. According to the study results, the performance of dual coagulants was affected by dosage of aluminium chloride. Beneficial effect by use of dual coagulants were only noted when aluminium chloride was underdosed. The addition sequence of coagulants was important for the performance of dual coagulants. Simultaneous addition resulted in the best performance, while addition of the polymer first resulted in the worst performance. Addition of aluminium chloride first resulted in the similar performance as single use of aluminium chloride. Although sulphate ion improved the floc characteristics, similar results were obtained. The effectiveness of rapid mixing depended on dosage of aluminium chloride. Extending rapid mixing (6 min) was beneficial when aluminium chloride was underdosed so that coagulation occurred at the combination region. However, such benefit was not observed at the optimum condition, which belonged to the sweep coagulation region. Different floc formation caused the difference. Extended rapid mixing would be beneficial when collision between clay particles and Al(III) was necessary. However, such benefit would disappear at the optimum condition because rapid mixing could break up the floc already formed.

The fouling characterization and control in the high concentration PAC membrane bioreactor HCPAC-MBR.

This study focuses on the experimental investigation to identify the effect of PAC at high concentrations on the fouling of membranes. A pilot-scale experimental apparatus was installed at a water treatment plant located downstream of Nakdong river basin, Korea. Effluent of rapid sand filter was used as influent of the system, which consists of PAC bio-reactor, submerged membrane module (hollow fiber with pore size 0.1 m) and air supply facility. PAC was dosed at 40 g/L initially and it was not replaced during the operation period. Suction type filtration was carried out at intervals of 12 min. suction and 3 min. idling. At the initial flux 0.36 m/d, the system could be operated stably for around 90 days at target trans-membrane pressure (TMP) of 40 kPa. Among total resistance of membrane filtration, cake and gel layer resistance, Rc+Rg, was the dominant fraction (more than 90% of the total) to increase the filtration pressure, which means that the filtration resistance could be controlled by the PAC cake layer and then irreversible membrane fouling could be prevented. Three minutes air backwashing every 3 days could extend the operation period to 127 days. Organics were analyzed in terms of molecular weight structure. The influent of the system consists of 15.0% and 74.4% of hydrophobic and hydrophilic natural organic matter (NOM), respectively. Hydrophobic and hydrophilic (electrostatic) interaction was the main factor on fouling of the membrane in the reactor. Hydrophobic fraction decreased slightly in the effluent, which means hydrophobic NOM removal in the reactor by adsorption. Organics accumulated in the membrane were extracted for analysis after a certain period of operation. The fraction of hydrophobic and hydrophilic organics was 41.4% and 38.9%, respectively. On the basis of the experimental results, the hydrophobic organics were the major materials causing the fouling of the membrane, which should be changed to other types of material.

Post-infectious bronchiolitis obliterans in children: CT features that predict responsiveness to pulse methylprednisolone.

OBJECTIVE: Intravenous pulse methylprednisolone therapy (IPMT) is an important treatment option for post-infectious obliterative bronchiolitis (OB), although it must be used carefully and only in selected patients because of its drawbacks. This study evaluated whether CT and clinical features of children with post-infectious OB can predict their responsiveness to IPMT. METHODS: We searched the medical records for patients (less than 18 years of age) who were diagnosed with post-infectious OB between January 2000 and December 2011. 17 children who received IPMT were included in this study. All underwent chest CT before and after IPMT. The radiological features seen on pre-treatment CT were recorded. The air-trapping area percentages on pre- and post-treatment CT images were determined. The nine patients who exhibited decreased air trapping on post-treatment CT scans relative to pre-treatment scans were classed as responders. The patient ages and time from initial pneumonia to IPMT were recorded. RESULTS: All responders and only four non-responders had thickened bronchial walls before treatment (p = 0.029). The two groups did not differ significantly in terms of bronchiolitis, bronchiectasis or the extent of air trapping, although the responders had a significantly shorter median interval between initial pneumonia and IPMT (4 vs 50 months; p = 0.005) and were significantly younger (median, 2.0 vs 7.5 years; p = 0.048). CONCLUSION: Immediate IPMT may improve the degree of air trapping in children with post-infectious OB if they show a thickened bronchial wall on CT. ADVANCES IN KNOWLEDGE: Children with post-infectious OB may respond favourably to IPMT when pre-treatment CT indicates bronchial-wall thickening.

Electronic damage in S atoms in a native protein crystal induced by an intense X-ray free-electron laser pulse.

Current hard X-ray free-electron laser (XFEL) sources can deliver doses to biological macromolecules well exceeding 1 GGy, in timescales of a few tens of femtoseconds. During the pulse, photoionization can reach the point of saturation in which certain atomic species in the sample lose most of their electrons. This electronic radiation damage causes the atomic scattering factors to change, affecting, in particular, the heavy atoms, due to their higher photoabsorption cross sections. Here, it is shown that experimental serial femtosecond crystallography data collected with an extremely bright XFEL source exhibit a reduction of the effective scattering power of the sulfur atoms in a native protein. Quantitative methods are developed to retrieve information on the effective ionization of the damaged atomic species from experimental data, and the implications of utilizing new phasing methods which can take advantage of this localized radiation damage are discussed.

Fine structure of the cerebellum of "staggerer-reeler", a double mutant of mice affected by staggerer and reeler conditions. I. The premature disappearance of the external granular layer and ensuing cerebellar disorganization.

It was reasoned that cerebella of mice carrying both staggerer and reeler conditions, called double mutant, might accentuate some of the anomalies observed in single mutants, thus revealing further details otherwise undetectable. As expected, the cerebellar hypoplasia, the paucity of fissures, granule cells and interneurons of the molecular layer, and the external granular layer hypoplasia, common in both staggerer and reeler, were much more pronounced in double mutant. However, the most striking feature of the double mutant cerebellum was the premature disappearance of the external granular layer. The external granular layer is smaller both in staggerer and reeler as compared to the normal but it persists beyond the normal time (around day 15). In double mutant, however, the external granular layer disappeared by day 7 or 8, reducing the density of granule cells even further. This observation was interpreted as indicating that both the staggerer and reeler genes effect the premature disappearance of the external granular layer. Most of the gross cerebellar anomalies in double mutant such as cerebellar hypoplasia, paucity of fissures and Purkinje and Golgi epithelial cell disalignment could be explained as the result of this premature disappearance of the external granular layer. On the other hand, anomalies of Purkinje cells and Bergmann fibers, which are also present both in staggerer and reeler, did not follow the same additive change.

Fine structure of the cerebellum of "staggerer-reeler", a double mutant of mice affected by staggerer and reeler conditions. II. Purkinje cell anomalies.

Purkinje cells in double mutant were very similar to those in staggerer both at days 7 and 15, and could not be distinguished from staggerer Purkinje cells by morphological criteria. They were small and immature looking, and their nuclei were invariably eccentric in position both at days 7 and 15. The typical funnel stage was not observed at any time. Somatic spines and their synapses were extermely rare at day 7 when they are abundant in the normal, but they were present at day 15 with typical synapses with climbing fibers. Dendritic spines were also present at day 15 although extremely infrequent, and they showed synapses both with parallel and climbing fibers. More often, however, parallel and climbing fibers were found in synapsis with the smooth surgace of these dentrites. These observations were in close agreement with those of staggerer Purkinje cells. The persistence of staggerer Purkinje cell soma anomalies in double mutant was interpreted as indicating that these anomalies are specific effects of the staggerer gene, being relatively free from environmental influences.

Fine structure of the cerebellum of "staggerer-reeler", a double mutant of mice affected by staggerer and reeler conditions. III. bergmann fiber anomalies.

Bergmann fibers and the distribution of Golgi epithelial cells were significantly altered in staggerer, reeler and double mutant (affected by both staggerer and reeler conditions). However, the Bergmann fiber anomalies in double mutant were not identical to those of either staggerer or reeler. A comparison of these anomalies with the anomalies described in weaver revealed that the developmental patterns of Bergmann fibers are distinct in all four cerebellar mutants just as are the cytoarchitectural anomalies of these cerebella. The results of the present electronmicroscopic examinations of Bergmann fibers in staggerer and reeler were in good agreement with the findings made with the immunofluorescence technique. There was, however, one important discrepancy between these two sets of observations. Immunofluorescent astrocytic fibers were nearly absent in 21 and 26 day old staggerers while Bergmann fibers were structurally present in nearly normal numbers both at days 21 and 31. A possible reason for this discrepancy was discussed.

Changes in voltage-gated calcium channel alpha(1) gene expression in rat dorsal root ganglia following peripheral nerve injury.

Although an increase in the excitability and ectopic spontaneous discharge (ESD) of primary sensory neurons can lead to abnormal burst activity, which is associated with neuropathic pain, the underlying molecular mechanisms are not fully understood. To investigate the relationship between these electrical abnormalities in injured neurons and voltage-gated calcium channel (VGCC) gene expression, reverse transcription-polymerase chain reaction (RT-PCR) was used to monitor the expression of the VGCC alpha(1) gene in the dorsal root ganglion (DRG) following chronic constriction injury (CCI) and axotomy of the rat sciatic nerve. Electrophoresis of the RT-PCR products showed the presence of multiple types of VGCC alpha(1) transcripts with various levels of basal expression in lumbar 4, 5, and 6 DRGs. CCI decreased alpha(1C), alpha(1D), alpha(1H), and alpha(1I) mRNA expression at 7 days in the ipsilateral DRG, to approximately 34-50% of the contralateral side. The same transcripts were repressed 7 days after sciatic axotomy and their reduction levels proved similar to those of CCI. Considering that changes of the intracellular calcium concentration modify the maintenance of ESD in injured DRG, these results suggest that the downregulation of alpha(1C), alpha(1D), alpha(1H) and alpha(1I) subunit gene expression in the rat DRG following peripheral nerve injury may contribute to the production of ESD associated with damaged nerves.

Regio- and stereocontrol elements in Rh(II)-catalyzed intramolecular C-H insertion of alpha-diazo-alpha-(phenylsulfonyl)acetamides.

[reaction: see text]. Intramolecular C-H insertion reaction of alpha-diazo-alpha-(phenylsulfonyl)acetamides proceeded with high regio- and stereoselectivities to afford highly functionalized gamma-lactams predominantly or exclusively. The high regioselectivity was attributed to the use of the phenylsulfonyl moiety, which altered electron density at the carbenoid center and exerted a steric effect during the insertion reaction. Also described herein are three control elements to determine regioselectivity, which are amide conformational, stereoelectronic, and substituent effects.

Interaction of cerebellar mutant genes. II. Mice doubly affected by 'reeler' and 'weaver' conditions.

An an extension of a study of the mode of ineraction of mutant genes that affect motor activities and cerebellar cytoarchitectonics, mice doubly affected by both the 'reeler' and 'weaver' conditions were produced. The cerebellum of the double mutant showed both reeler and weaver characteristics. Anomalies present in both mutants were greatly accentuated in the double mutant. In addition to the paucity of fissures and general cerebellar disorganization observed in reeler and the extreme scarcity of granule cells seen in weaver, double mutants showed an additional decrease in the cerebellar size and the molecular layer, and a further loss of external granule cells. These observations were interpreted as indicating that the effects of the reeler and weaver genes upon altering the cytoarchitectonics of the cerebellum were additive.