Relations between thermodynamics, structures, and dynamics for modified water models in their supercooled regimes.

We use molecular dynamics simulations to study relations between thermodymamic, structural, and dynamical properties of TIP4P/2005 water models with systematically reduced partial charges and, thus, weaker hydrogen bonds. Observing a crossing of isochores in the P-T diagram, we show that these water-like models have a readily accessible liquid-liquid critical point (LLCP) associated with a transition between high-density liquid (HDL) and low-density liquid (LDL) forms and determine the dependence of the critical temperature Tc, pressure Pc, and density ρc on the charge-scaling factor from fits to a two-structure equation of states. The results indicate that the water-like models exhibit liquid polyamorphism in a wide range of interaction parameters. Considering elongated systems, we observe a decomposition into extended and stable HDL-like and LDL-like regions at appropriate pressures and low temperatures and analyze the respective structural and dynamical properties. We show that the diverse local order results in very different correlation times of local dynamics, while the fragility is hardly changed. The results yield insights into the origin of a dynamical crossover, which is observed when lowering the temperature along isobars and was previously interpreted in terms of a fragile-to-strong transition. Our findings imply that the effect does not involve two liquid phases with an exceptionally large difference of the fragility but rather a high temperature dependence near the LLCP results from a rapid conversion from HDL-like environments with faster dynamics to LDL-like ones with slower dynamics.

MD simulations of charged binary mixtures reveal a generic relation between high- and low-temperature behavior.

Experimental studies of the glassy slowdown in molecular liquids indicate that the high-temperature activation energy E∞ of glass-forming liquids is directly related to their glass transition temperature Tg. To further investigate such a possible relation between high- and low-temperature dynamics in glass-forming liquids, we analyze the glassy dynamics of binary mixtures using molecular dynamics simulations. We consider a binary mixture of charged Lennard-Jones particles and vary the partial charges of the particles and, thus, the high-temperature activation energy and the glass transition temperature of the system. Based on previous results, we introduce a phenomenological model describing relaxation times over the whole temperature regime from high temperatures to temperatures well inside the supercooled regime. By investigating the dynamics of both particle species on molecular and diffusive length scales along isochoric and isobaric pathways, we find a quadratic charge dependence of both E∞ and Tg, resulting in an approximately constant ratio of both quantities independent of the underlying observable, the thermodynamic ensemble, and the particle species, and this result is robust against the actual definition of Tg. This generic relation between the activation energy and the glass transition temperature indicates that high-temperature dynamics and the glassy slowdown are related phenomena, and the knowledge of E∞ may allow us to approximately predict Tg.

Effects of confinement on supercooled tetrahedral liquids.

We use molecular dynamics simulations to ascertain the effects of geometrical restriction on glass-forming tetrahedral liquids. Striving for a broad approach, we study families of waterlike and silicalike liquids, for which we systematically scale the partial charges and, hence, the relevance of the tetrahedral networks. The confined liquids and the confining matrices consist of the same type of particles to avoid disruptive interactions and distorted structures at the interfaces. Spatially resolved analyses show that these neutral confinements still impose static mobility gradients and density correlations on the liquids. We quantify the increasing degree and range of the altered properties upon cooling. For both families of models, common relations describe the confinement effects of all systems with tetrahedral order, while deviations occur for systems with lower polarities and different structures. The observations are rationalized by considering the fact that a pinned wall imprints a static energy landscape to a neighboring liquid. We explore the properties of this landscape based on changes in vibrational motion and structural relaxation and find that typical barrier heights amount to two to three times the activation energy of bulk dynamics. Combining the present and previous results, we predict the evolution of confinement effects down to the glass transition temperature for liquids without fragile-to-strong crossover. In addition, it is found for silicalike liquids that the temperature dependence of dynamic and static correlation lengths from confinement studies is not affected when cooling through fragile-to-strong transitions of the bulk materials, casting doubt on the relevance of these length scales for the glassy slowdown.

Common behaviors associated with the glass transitions of water-like models.

We perform molecular dynamics simulations to ascertain effects of the molecular polarity on structural and dynamical properties of water-like systems, in particular, on their glassy slowdown. To systematically vary the molecular dipole moments, we scale the partial charges of the established SPC/E and TIP4P/2005 models. In broad ranges of the molecular polarity, the studied SPC/E and TIP4P/2005 descendants show a density anomaly, which can be attributed to the removal of water molecules interstitial between the first and the second neighbor shells upon cooling. While all considered modified water models behave as typical glass formers, the structural relaxation time τ heavily depends on the molecular dipole moment. This large dynamical diversity is exploited to systematically ascertain characteristic properties of glass-forming liquids. For all studied water-like systems, we observe a close relation between the activation energy E∞ describing the Arrhenius behavior of the regular liquid and the glass transition temperature Tg characterizing the supercooled liquid, explicitly, E∞/Tg≈10. Moreover, decomposing the activation energy of the structural relaxation according to E(T)=E∞+Ec(T), we show that the glassy slowdown of all modified water molecules can fully be traced back to an exponential temperature dependence of the contribution Ec(T) related to cooperative dynamics. Extrapolation of this behavior suggests a common value at the glass transition temperature, Ec(Tg)/Tg≈25. Finally, we discuss links between the structural relaxation and the vibrational displacement, as proposed in various theoretical approaches to the glass transition.

Lacosamide cardiac safety: a thorough QT/QTc trial in healthy volunteers.

OBJECTIVE: To determine whether lacosamide prolongs the corrected QT interval (QTc). MATERIALS AND METHODS: In this randomized, double-blind, positive- and placebo-controlled, parallel-design trial, healthy volunteers were randomized to lacosamide 400 mg/day (maximum-recommended daily dose, 6 days), lacosamide 800 mg/day (supratherapeutic dose, 6 days), placebo (6 days), or moxifloxacin 400 mg/day (3 days). Variables included maximum time-matched change from baseline in QT interval individually corrected for heart rate ([HR] QTcI), other ECG parameters, pharmacokinetics (PK), and safety/tolerability. RESULTS: The QTcI mean maximum difference from placebo was -4.3 ms and -6.3 ms for lacosamide 400 and 800 mg/day; upper limits of the 2-sided 90% confidence interval were below the 10 ms non-inferiority margin (-0.5 and -2.5 ms, respectively). Placebo-corrected QTcI for moxifloxacin was +10.4 ms (lower 90% confidence bound >0 [6.6 ms]), which established assay sensitivity for this trial. As lacosamide did not increase QTcI, the trial is considered a negative QTc trial. There was no dose-related or clinically relevant effect on QRS duration. HR increased from baseline by ~5 bpm with lacosamide 800 mg/day versus placebo. Placebo-subtracted mean increases in PR interval at tmax were 7.3 ms (400 mg/day) and 11.9 ms (800 mg/day). There were no findings of second-degree or higher atrioventricular block. Adverse events (AEs) were dose related and most commonly involved the nervous and gastrointestinal systems. CONCLUSIONS: Lacosamide (≤ 800 mg/day) did not prolong the QTc interval. Lacosamide caused a small, dose-related increase in mean PR interval that was not associated with AEs. Cardiac, overall safety, and PK profiles for lacosamide in healthy volunteers were consistent with those observed in patients with partial-onset seizures.

Lacosamide cardiac safety: clinical trials in patients with partial-onset seizures.

OBJECTIVE: To evaluate the cardiac safety of adjunctive lacosamide in a large pool of adults with partial-onset seizures (POS). METHODS: Post-randomization changes from baseline for electrocardiographic (ECG) measurements, diagnostic findings, and relevant adverse events (AEs) were compared for pooled data from three randomized, placebo-controlled trials of adjunctive lacosamide for the treatment of POS. RESULTS: Lacosamide did not prolong the QTc interval or affect heart rate as determined by an analysis of data from patients randomized to lacosamide 200, 400, or 600 mg/day (n = 944) compared with placebo (n = 364). After 12-week maintenance treatment, mean changes from baseline for QRS duration were similar between the placebo and lacosamide 200 and 400 mg/day groups (0.0, -0.2, and 0.4 ms), but slightly increased for lacosamide 600 mg/day (2.3 ms). A small, dose-related mean increase in PR interval was observed (-0.3, 1.4, 4.4, and 6.6 ms for the placebo and lacosamide 200, 400, and 600 mg/day groups, respectively). First-degree atrioventricular (AV) block was reported as a non-serious AE in 0.0%, 0.7%, 0.2%, and 0.5% of patients in the same respective groups. Second- or higher degree AV block was not observed. There was no evidence of a PR-interval-related pharmacodynamic interaction of lacosamide with either carbamazepine or lamotrigine. CONCLUSIONS: Evaluation of the pooled cardiac safety data from patients with POS showed that adjunctive lacosamide at the maximum recommended dose (400 mg/day) was not clearly associated with any cardiac effect other than a small, dose-related increase in PR interval that had no evident symptomatic consequence.

A susceptibility locus for Parkinson's disease maps to chromosome 2p13.

Parkinson's disease (PD) is a common degenerative neurologic disorder, which is pathologically characterized by a selective degeneration of dopaminergic neurons of the substantia nigra pars compacta, and the presence of characteristic eosinophilic inclusions, known as Lewy-bodies in affected brain areas. The cause of PD is unknown but, in recent years, genetic factors have been implicated in the aetiology of the disease. Firstly, clinico-genetic, epidemiologic and twin studies revealed inheritable effects and questioned earlier studies which had denied such influences. Secondly, several family studies suggested autosomal-dominant inheritance of syndromes which, to variable degrees, resembled sporadic PD clinically and in some cases also neuropathologically. Recently, a disease locus has been mapped to chromosome 4q21-22 in a large Mediterranean pedigree, in which disease expression is clinically and pathologically within the spectrum of sporadic PD; being atypical only for a relatively young mean age at onset of 46 years and rapid course of 10 years from onset to death. In affected individuals of this family and of three unrelated Greek kindreds, a putative disease-causing mutation has been identified in the gene encoding alpha-synuclein. With the first variant being defined, genetic heterogeneity has become apparent, as in other families parkinsonism was not linked to the 4q-locus and was not associated with the alpha-synuclein mutation (unpublished data). We describe a different genetic locus that appears to be involved in the development of parkinsonism closely resembling sporadic PD including a similar mean age of onset (59 years in the families, 59.7 years in sporadic PD; ref. 12). This locus was detected in a group of families of European origin. In two of these families, there is genetic evidence for a common founder. The penetrance of the mutation appears to be low, most likely below 40%. This is compatible with a possible role of this locus not only in familial, but also in typical (sporadic) PD.

A structural variant of the human interferon-gamma gene.

The first structural IFNG variant, G54D (c.287G>A, ss105106770), located in the second exon, was identified.

[Molecular diagnostics of hereditary fever syndromes. Familial Mediterranean fever (FMF), hyperimmunoglobulin D syndrome (HIDS), tumor necrosis factor receptor-associated periodic syndrome (TRAPS), cryopyrin-associated periodic syndrome (CAPS: FCAS, MWS, NOMID/CINCA)]. / Molekulargenetische Diagnostik erblicher Fiebersyndrome. Familiäres Mittelmeerfieber (FMF), Hyperimmunglobulin-D-Syndrom (HIDS), TNF-rezeptorassoziiertes periodisches Syndrom (TRAPS), Cryopyrin-assoziiertes periodisches Syndrom (CAPS: FCAS, MWS, NOMID/CINCA).

Periodic episodes of fever and inflammation can have a genetic origin. Nowadays, the identification of the causative genetic variants in the majority of cases allows molecular genetic confirmation of the clinical diagnosis, which enables approaches with specific drug treatment and improves patient compliance as well as genetic counseling. Besides a detailed clinical examination a medical history including family history and an assessment of the ethnic origin are required. In order to make genetic testing straightforward and cost effective an iterative procedure should be followed which should include, in addition to clinical data, the frequencies of causative mutations in the various gene segments involved.

No associations of human pulmonary tuberculosis with Sp110 variants.

BACKGROUND: After a recent report on the role of the Ipr1 gene in mediating innate immunity in a mouse model of Mycobacterium tuberculosis infection, the human Ipr1 homologue, Sp110, was considered a promising candidate for an association study in human tuberculosis. METHODS: In a sample of >1000 sputum positive, HIV negative West African patients with pulmonary tuberculosis and >1000 exposed, apparently healthy controls, we have genotyped 21 Sp110 gene variants that were either available from public databases, including HapMap data, or identified by DNA re-sequencing. RESULTS: No significant differences in the frequencies of any of the 21 variants were observed between patients and controls. This applied also for HapMap tagging variants and the corresponding haplotypes, when including sliding window analyses with three adjacent variants, and when stratifying controls for positivity and negativity according to the results of intradermal tuberculin (purified protein derivative, PPD) skin tests. DNA re-sequencing revealed 13 novel Sp110 variants in the 5'-UTR, exons, and adjacent intronic regions. CONCLUSIONS: Based on the results obtained in this case-control study, the hypothesis that Sp110 variants and haplotypes might be associated with distinct phenotypes of human M tuberculosis infection is doubtful.

YAC/BAC contig spanning the MHC class III region of cattle.

A contig of the class III region of the bovine major histocompatibility complex (MHC) was established from bacterial and yeast artificial chromosomes using PCR and BAC-end sequencing. The marker content of individual clones was determined by gene and BAC-end specific PCR, and the location of genes and BAC-ends was confirmed analyzing somatic hybrid cells. A comparative analysis indicated that the content and order of MHC class III genes is strongly conserved between cattle and other mammalian species. Fluorescence in situ hybridization localized the bovine class III region to BTA23q21-->q22. The results show that the collection of sequenced BAC-ends is a powerful resource for generating high-resolution comparative chromosome maps.

Pre-treatment with chloroquine and parasite chloroquine resistance in Ghanaian children with severe malaria.

BACKGROUND: Self-medication with anti-malarial drugs is widespread, and chloroquine (CQ) resistance is increasing. The effect of these factors on the incidence and presentation of severe malaria is uncertain. AIM: To investigate subtype of severe malaria, duration of illness, previous CQ treatment and prevalence of Plasmodium falciparum CQ-resistance markers among children presenting with severe malaria to a teaching hospital in Ghana. DESIGN: Prospective clinical study. METHODS: Consecutive patients (n = 189) presenting with severe malaria were examined clinically, and blood was taken for routine haematology and malaria films. Plasma and blood cells were stored and subsequently analysed by ELISA for CQ levels (n = 168) and by PCR and restriction digest for P. falciparum chloroquine resistance transporter gene (pfcrt) mutations (n = 139). RESULTS: Of 47 presenting with cerebral malaria, 21 had severe anaemia and 13 respiratory distress (RDS). Twenty-nine had prostration or RDS alone, 41 severe anaemia with prostration or RDS, and 72 severe anaemia not associated with coma, prostration or RDS. Of the patients studied, 77% had CQ in their plasma, and 88% were carrying P. falciparum with a CQ-resistance genotype. Significant associations were found (i) between the CQ-resistance genotype of parasites and plasma CQ levels, (ii) between the presence of CQ in plasma and the reported duration of illness, and (iii) between the reported duration of illness and the occurrence of severe but otherwise uncomplicated anaemia. DISCUSSION: There was extensive prior CQ use in our patients presenting with severe malaria, and a high prevalence of parasites with the CQ-resistance genotype. CQ resistance in P. falciparum may contribute to the development of severe but otherwise uncomplicated anaemia in this setting.

Variability of the CD36 gene in West Africa.

Studying 12 selected individuals from a malaria-endemic area in West Africa, 24 variants of the CD36 gene were found, 21 of them novel ones. These included three single-nucleotide substitutions causing non-conservative amino acid exchanges E123K, T174A, and I271T as well as a three base pair (bp) insertion resulting in the addition of an asparagine residue (N232-233ins). The E123K variant was located within the putative ligand-binding domain for oxidized low density lipoprotein, while the other substitutions resided outside any of the binding sites for reaction partners mapped on CD36 so far. Twelve single-nucleotide polymorphisms (SNPs) were identified in untranslated parts of the exons and in introns. Five additional SNPs were located in the promoter region whereby -144G-->T, -53G-->T, and -2A-->G alter putative binding sites for the transcription factors purine factor (PuF), phorbol ester-responsive element AP-2, and CCAAT/enhancer-binding protein. A G-->T exchange at position -50 appears to introduce a new recognition site for PuF. Calculations of nucleotide diversity revealed extraordinarily high numbers for all parts of the gene, which may, however, to some extent be due to the selection of individuals studied.

Pattern of connexin 26 (GJB2) mutations causing sensorineural hearing impairment in Ghana.

Mutations of the connexin 26 gene (GJB2) were studied in 365 apparently unrelated individuals with profound nonsyndromic, sensorineural hearing impairment from Ghana, West Africa. Among 121 mutated chromosomes found, 110 carried the previously described R143W mutation. A total of 6 novel mutations: L79P, V178A, R184Q, A197S, I203K, and L214P, were identified, whereby I203K was based on a dinucleotide exchange and R184Q appeared to be dominant. The GJB2 variants found in Ghana tend to comprise less nonsense and frameshift mutations and more mutations located in the C-terminal half of the molecule than the variants found in other parts of the world.

Direct sequencing of variable HLA gene segments after in vitro amplification and allele separation by temperature-gradient gel electrophoresis.

Previously unrecognized variants of human leukocyte antigens (HLA) are currently being analyzed by in vitro amplification and sequencing of the variable gene segments. In heterozygous individuals, molecular cloning is required to separate the two concomitantly amplified haplotypic gene segments. A method is presented which facilitates the procedure of separating the two haplotypic gene segments by using a temperature-gradient gel electrophoresis (TGGE). The procedure comprises PCR amplification of the variable HLA gene segments, allele separation by TGGE, re-amplification of each of the separated allelic segments, and direct DNA sequencing using the PCR primers.

Comparison of pore-forming peptides from pathogenic and nonpathogenic Entamoeba histolytica.

Similar to the findings obtained with pathogenic Entamoeba histolytica, nonpathogenic isolates were found to kill mammalian cells in vitro, and cell extract caused pore formation in liposome membranes. A pore-forming peptide termed APnp was isolated from a nonpathogenic isolate using the schedule developed for the purification of APp or amoebapore, the homologous peptide of the pathogenic isolate HM-1:IMSS. Compared to APp, the specific activity of APnp in pore formation was 60% lower. cDNA sequencing indicated 95% identity of the primary structures of APnp and APp, and secondary structure predictions revealed a high degree of similarity. Notably, a glutamic acid residue at position 2 of APp is in APnp replaced by proline, which shortens one of the two amphipathic alpha-helices considered crucial for the pore-forming function. This structural divergence of the two peptides might explain the difference in their pore-forming activities.

Putative serine/threonine protein kinase expressed in complement-resistant forms of Entamoeba histolytica.

Entamoeba histolytica is susceptible to complement attack in its lumen-dwelling state and develops complement resistance during pathogenic tissue invasion. As experimental evidence suggests that this change in phenotype is accompanied by a change in gene expression, we constructed a subtractive cDNA library to identify genes involved. Poly(A) + RNA from complement-sensitive trophozoites was subtracted from single stranded cDNA derived from complement-resistant ones. Transcripts enriched in the library were found to code for a putative polypeptide comprising all sequence elements characteristic for serine/threonine protein kinases. The gene contains an intron of 46 nucleotides and two polyadenylation sites. Northern-blot analyses confirmed that the gene is expressed in both tissue-derived and laboratory-grown forms of complement-resistant E. histolytica.

Pathogenic and nonpathogenic Entamoeba histolytica: identification and molecular cloning of an iron-containing superoxide dismutase.

Superoxide dismutase (SOD) activity was determined in the cell lysate of the axenically cultured Entamoeba histolytica isolate HM-1:IMSS. Under anaerobic culture conditions, 18.7 (+/- 4.9) units SOD activity (mg protein)-1 were found. By inhibition studies the activity was attributed to an iron-containing type of SOD (FeSOD). Using degenerate oligonucleotide primers derived from regions highly conserved in prokaryotic FeSOD sequences, a genomic DNA fragment was amplified by the polymerase chain reaction. The fragment was used to isolate FeSOD specific cDNA clones from a pathogenic and a nonpathogenic E. histolytica isolate. A comparison of the 2 sequences revealed 5% nucleotide differences resulting in a single amino acid exchange. The primary structure showed the characteristics of an iron-containing type of SOD with a homology of approximately 55% with other FeSOD sequences. The enzyme was found to be encoded by single copy genes in both the pathogenic and the nonpathogenic E. histolytica, but restriction fragment lengths differed between the 2 groups. In 5 isolates studied, no correlation was found between pathogenic behavior of the amebae and the expression of FeSOD-related mRNA.

High mortality of infant bacteraemia clinically indistinguishable from severe malaria.

BACKGROUND: Early recognition of children at highest risk of dying and the targeting of appropriate drug therapy are vital to the improvement of paediatric care in developing countries. This will rely upon the development of simple clinically-based algorithms and treatment guidelines. AIM: To determine the role of bacteraemia in children presenting with clinical signs and symptoms of severe malaria. DESIGN: Retrospective analysis of blood culture results following prospective data collection. METHODS: We studied 251 children presenting with symptoms and signs of severe malaria to a tertiary referral centre in Ghana. Blood was taken for malaria blood films, bacterial culture and haemograms. RESULTS: On the basis of clinical signs alone, malaria-film-positive (n = 182) and -negative (n = 69) patients were indistinguishable. Some 40% of film-negative patients were bacteraemic, vs. 12% of film-positive patients. Severe malaria and bacteraemia were not positively associated. Film-negative bacteraemic patients had a mortality of 39%, primarily affecting the age group <30 months. DISCUSSION: Infants presenting with symptoms and signs of severe malaria but a negative malaria film require immediate antibiotic treatment.

Tolerance of mefloquine alone and in combination with sulfadoxine-pyrimethamine in the prophylaxis of malaria.

A randomized double blind study was performed to evaluate the tolerance and the acceptance of mefloquine alone (Lariam) compared to a combined drug regimen consisting of mefloquine, sulfadoxine and pyrimethamine (MSP; Fansimef) in the prophylaxis of malaria. 175 Europeans travelling to different malaria endemic areas received either mefloquine alone (250 mg/week) or its combination with sulfadoxine (500 mg/week) plus pyrimethamine (25 mg/week). One person taking mefloquine and two taking MSP discontinued the drug intake because of moderate clinical side effects. Mild and moderate adverse clinical reactions predominantly concerning the gastro-intestinal tract and the autonomous nervous system were reported with a significantly higher occurrence in the MSP group. With both prophylactic regimens, reversibly elevated liver enzyme activities (glutamate oxalate transaminase and glutamate pyruvate transaminase [GPT]) were observed after prophylaxis. The increase of GPT serum activity correlated significantly with relatively high GPT levels before prophylaxis in both groups. This finding suggests a limited use of both regimens in cases of liver dysfunction. One case of mefloquine-resistant Plasmodium falciparum malaria was observed from West Africa; this patient was cured by a standard regimen of chloroquine.