Determining the neurocognitive profile of children with tuberous sclerosis complex within the Western Cape region of South Africa.

BACKGROUND: Tuberous sclerosis complex (TSC) is a multisystem genetic disorder associated with a wide spectrum of cognitive impairments that can often result in impaired academic, social and adaptive functioning. However, studies investigating TSC have found it difficult to determine whether TSC is associated with a distinct cognitive phenotype and more specifically which aspects of functioning are impaired. Furthermore, children with TSC living in low-income and middle-income countries, like South Africa, experience additional burdens due to low socio-economic status, high mortality rates and poor access to health care and education. Hence, the clinical population of South Africa may vary considerably from those populations from high-income countries discussed in the literature. METHODS: A comprehensive neuropsychological battery composed of internationally recognised measures examining attention, working memory, language comprehension, learning and memory, areas of executive function and general intellectual functioning was administered to 17 children clinically diagnosed with TSC. RESULTS: The exploration of descriptive data indicated generalised cognitive difficulties in most cognitive domains, aside from memory. With only two participants performing in the average to above-average ranges, the rest of the sample showed poor verbal comprehension, perceptual reasoning, working memory, processing speed, disinhibition, and problems with spatial planning, problem solving, frustration tolerance, set shifting and maintaining a set of rules. Furthermore, correlational findings indicated several associations between socio-demographic and cognitive variables. CONCLUSIONS: Importantly, this is the first study to comprehensively examine multiple domains of neurocognitive functioning in a low-resource setting sample of children with TSC. Current study findings showed that children with TSC have generalised impairments across several cognitive domains, rather than domain-specific impairments. Therefore, although examining individual aspects of cognition, such as those found in previous literature, is important, this approach is limiting. With a comprehensive assessment, including understanding the associations between domains, appropriate and directed support can be provided to ensure all aspects of development are addressed and considered.

BRCA1 protein is linked to the RNA polymerase II holoenzyme complex via RNA helicase A.

The breast cancer specific tumour suppressor protein, BRCA1 (refs 1,2), activates transcription when linked with a DNA-binding domain and is a component of the RNA polymerase II (Pol II) holoenzyme. We show here that RNA helicase A (RHA) protein links BRCA1 to the holoenzyme complex. The region of BRCA1 which interacts with RHA and, thus, the holoenzyme complex, corresponds to subregions of the BRCT domain of BRCA1 (ref. 9). This interaction was shown to occur in yeast nuclei, and expression in human cells of a truncated RHA molecule which retains binding to BRCA1 inhibited transcriptional activation mediated by the BRCA1 carboxy terminus. These data are the first to identify a specific protein interaction with the BRCA1 C-terminal domain and are consistent with the model that BRCA1 functions as a transcriptional coactivator.

Tuberous sclerosis complex in the Western Cape, South Africa: The clinical presentation features.

Tuberous sclerosis complex (TSC) is a genetic neurocutaneous condition, which affects multiple organ systems. This study aimed to determine the presenting features of children with TSC in Cape Town, South Africa. A cross-sectional study was conducted at a TSC clinic, and clinical features at presentation were prospectively collected. Thirty-nine children (23 boys; median age 10 (range 1 - 26) years; median diagnosis age 16 (0 - 153) months) were recruited. Twenty-one (54%) children presented with focal seizures. Seven (18%) children had epileptic spasms. Skin manifestations led to a diagnosis in 13 (33%) and neuroimaging in 22 (56%) children. Antenatal screening detected cardiac rhabdomyomas in 3 children. One child had a positive family history. In the paediatric service, TSC diagnosis usually followed neuroimaging, whereas at the neurology service skin manifestations indicated TSC. In conclusion, most children with TSC presented as emergency cases with seizures. Health practitioner awareness of the common TSC clinical signs was lacking, with the diagnosis often delayed.

An European survey of structure and organisation of nutrition support teams in Germany, Austria and Switzerland.

BACKGROUND & AIMS: Nutritional support teams (NST) have been demonstrated to be an excellent mechanism for identifying patients in need of nutrition support, improving the efficacy of nutrition support in a variety of hospital environments. Focus of this study was the investigation of function, structure and organisation of NST in Germany (D), Austria (A) and Switzerland (CH). METHODS: Prospective investigation of the function, structure and organisation of NST in D, A and CH, using standardised questionnaires. RESULTS: From a total of 3071 hospitals in D, A and CH, NST have been established at 98 hospitals (3.2%). Their main activities were creating nutritional regimes (100%), education (87%) and monitoring nutrition therapy (92%). In general, the NST are not independently operating units but are affiliated to a special discipline. Seventy-one per cent of the physicians, 40% of the nurses and 69% of the dieticians in the NST held a nutrition-specific additional qualification. A total of 12% of the physicians, 37% of the nurses and 46% of the dieticians are exclusively responsible for the NST. A reduction of complications (88%) and cost saving (98%) were indicated since their establishment. The NST received in 32% funding support. CONCLUSION: In D, A, CH neither a uniform nor comprehensive patient care by NST existed in 2004. Standards of practice, development of guidelines in clinical nutrition and better documentation in NSTs are necessary. Special efforts should be aimed at education of NST members and financing of teams.

Expression and characterization of recombinant type 2 3 alpha-hydroxysteroid dehydrogenase (HSD) from human prostate: demonstration of bifunctional 3 alpha/17 beta-HSD activity and cellular distribution.

In androgen target tissues, 3alpha-hydroxysteroid dehydrogenase (3alpha-HSD) may regulate occupancy of the androgen receptor (AR) by catalyzing the interconversion of 5alpha-dihydrotestosterone (5alpha-DHT) (a potent androgen) and 3alpha-androstanediol (a weak androgen). In this study, a 3alpha-HSD cDNA (1170 bp) was isolated from a human prostate cDNA library. The human prostatic 3alpha-HSD cDNA encodes a 323-amino acid protein with 69.9%, 84.1%, 99.4%, and 87.9% sequence identity to rat liver 3alpha-HSD and human type 1, type 2, and type 3 3alpha-HSDs, respectively, and is a member of the aldo-keto reductase superfamily. The close homology with human type 2 3alpha-HSD suggests that it is either identical to this enzyme or a structural allele. Surprisingly, when the recombinant protein was expressed and purified from Escherichia coli, the enzyme did not oxidize androsterone when measured spectrophotometrically, an activity previously assigned to recombinant type 2 3alpha-HSD using this assay. Complete kinetic characterization of the purified protein using spectrophotometric, fluorometric, and radiometric assays showed that the catalytic efficiency favored 3alpha-androstanediol oxidation over 5alpha-DHT reduction. Using [14C]-5alpha-DHT as substrate, TLC analysis confirmed that the reaction product was [14C]-3alpha-androstanediol. However, in the reverse reaction, [3H]-3alpha-androstanediol was oxidized first to [3H]-androsterone and then to [3H]-androstanedione, revealing that the expressed protein possessed both 3alpha- and 17beta-HSD activities. The 17beta-HSD activity accounted for the higher catalytic efficiency observed with 3alpha-androstanediol. These findings indicate that, in the prostate, type 2 3alpha-HSD does not interconvert 5alpha-DHT and 3alpha-androstanediol but inactivates 5alpha-DHT through its 3-ketosteroid reductase activity. Levels of 3alpha-HSD mRNA were measured in primary cultures of human prostatic cells and were higher in epithelial cells than stromal cells. In addition, elevated levels of 3alpha-HSD mRNA were observed in epithelial cells derived from benign prostatic hyperplasia and prostate carcinoma tissues. Expression of 3alpha-HSD was not prostate specific, since high levels of mRNA were also found in liver, small intestine, colon, lung, and kidney. This study is the first complete characterization of recombinant type 2 3alpha-HSD demonstrating dual activity and cellular distribution in the human prostate.

The effect of beta-carotene on the expression of interleukin-6 and heme oxygenase-1 in UV-irradiated human skin fibroblasts in vitro.

beta-Carotene is discussed as an anti-oxidant micronutrient and singlet oxygen quencher in human skin, protecting against UV light-induced damage. However, we recently demonstrated that beta-carotene has a pro-oxidant potential in cultured human skin fibroblasts because it enhances the UVA induction of heme oxygenase-1 (HO-1). Herein, we further show that beta-carotene also strongly promotes the UVA induction of pro-inflammatory interleukin-6 (IL-6) in skin fibroblasts in vitro. Singlet oxygen quencher sodium azide abrogated up-regulation of IL-6, and likewise also of HO-1. In UVB-irradiated cells, beta-carotene did not modulate levels of IL-6 and HO-1. The observed effects might be relevant for UV-induced inflammatory processes.

Mammalian 3 alpha-hydroxysteroid dehydrogenases.

Mammalian 3 alpha-hydroxysteroid dehydrogenases (3 alpha-HSDs) regulate steroid hormone levels. For example, hepatic 3 alpha-HSDs inactivate circulating androgens, progestins, and glucocorticoids. In target tissues they regulate access of steroid hormones to steroid hormone receptors. For example, in the prostate 3 alpha-HSD acts as a molecular switch and controls the amount of 5 alpha-dihydrotestosterone that can bind to the androgen receptor, while in the brain 3 alpha-HSD can regulate the amount of tetrahydrosteroids that can alter GABAa receptor function. Molecular cloning indicates that these mammalian 3 alpha-HSDs belong to the aldo-keto reductase superfamily and that they are highly homologous proteins. Using the three-dimensional structure of rat liver 3 alpha-HSD as a template for site-directed mutagenesis, details regarding structure function relationships, including catalysis and cofactor and steroid hormone recognition have been elucidated. These details may be relevant to all mammalian 3 alpha-HSDs.

Structure and function of 3 alpha-hydroxysteroid dehydrogenase.

Mammalian 3 alpha-hydroxysteroid dehydrogenases (3 alpha-HSDs) inactivate circulating steroid hormones, and in target tissues regulate the occupancy of steroid hormone receptors. Molecular cloning indicates that 3 alpha-HSDs are members of the aldo-keto reductase (AKR) superfamily and display high sequence identity (> 60%). Of these, the most extensively characterized is rat liver 3 alpha-HSD. X-ray crystal structures of the apoenzyme and the E.NADP+ complex have been determined and serve as structural templates for other 3 alpha-HSDs. These structures reveal that rat liver 3 alpha-HSD adopts an (alpha/beta)8-barrel protein fold. NAD(P)(H) lies perpendicular to the barrel axis in an extended conformation, with the nicotinamide ring at the core of the barrel, and the adenine ring at the periphery of the structure. The nicotinamide ring is stabilized by interaction with Y216, S166, D167, and Q190, so that the A-face points into the vacant active site. The 4-pro-(R) hydrogen transferred in the oxidoreduction of steroids is in close proximity to a catalytic tetrad that consists of D50, Y55, K84, and H117. A water molecule is within hydrogen bond distance of H117 and Y55, and its position may mimic the position of the carbonyl of a 3-ketosteroid substrate. The catalytic tetrad is conserved in members of the AKR superfamily and resides at the base of an apolar cleft implicated in binding steroid hormone. The apolar cleft consists of a side of apolar residues (L54, W86, F128, and F129), and opposing this side is a flexible loop that contains W227. These constraints suggest that the alpha-face of the steroid would orient itself along that side of the cleft containing W86. Site-directed mutagenesis of the catalytic tetrad indicates that Y55 and K84 are essential for catalysis. Y55S and Y55F mutants are catalytically inactive, but still form binary (E.NADPH) and ternary (E.NADH.Testosterone) complexes; by contrast K84R and K84M mutants are catalytically inactive, but do not bind steroid hormone. The reliance on a Tyr/Lys pair is reminiscent of catalytic mechanisms proposed for other AKR members as well as for HSDs that belong to the short-chain dehydrogenase/reductase (SDR) family, in which Tyr is the general acid, with its pKa being lowered by Lys. Superimposition of the nicotinamide rings in the structures of 3 alpha-HSD (an AKR) and 3 alpha, 20 beta-HSD (an SDR) show that the Tyr/Lys pairs are positionally conserved, suggesting convergent evolution across protein families to a common mechanism for HSD catalysis. W86Y and W227Y mutants bind testosterone to the E.NADH complex, with effective increases in Kd of 8- and 20-fold. These data provide the first evidence that the side of the apolar cleft containing W86 and the opposing flexible loop containing W227 are parts of the steroid-binding site. Detailed mutagenesis studies of the apolar cleft and elucidation of a ternary complex structure will ultimately provide details of the determinants that govern steroid hormone recognition. These determinants could provide a rational basis for structure-based inhibitor design.

High-dose intravenous vitamin C is not associated with an increase of pro-oxidative biomarkers.

OBJECTIVE: High-dose vitamin C therapy might mediate beneficial clinical effects by counteracting reactive oxygen species. However, concerns are raised whether this approach might provoke diametrical (ie pro-oxidative) effects. The objective was to determine ascorbyl free radical (AFR) concentrations and potential variables of pro-oxidative damage. DESIGN: Crossover study; six healthy males received daily infusions of 750 or 7500 mg vitamin C for six consecutive days. Fasting concentrations of vitamin C and AFR were determined daily. On day 1, concentrations of vitamin C and AFR were measured at 0.25, 0.5, 1, 2, 4 and 8 h post infusion. Plasma concentrations of thiobarbituric acid-reactive substances (TBARS), tocopherol and urine concentrations of 8-oxoguanosine were determined on days 1 and 6. RESULTS: Kinetic studies on day 1 showed that concentrations of vitamin C and AFR displayed parallel dose- and time-dependent kinetics and elimination was highly efficient. Vitamin C and AFR fasting concentrations on days 2-6 were slightly above the baseline, suggesting new, stable steady states. TBARS decreased in both groups, whereas tocopherol and 8-oxoguanosine concentrations remained unchanged. CONCLUSION: Kinetics of AFR largely depend on plasma vitamin C concentrations and AFR is eliminated efficiently. Our data do not support induction of pro-oxidative effects in healthy volunteers given intravenous high-dose vitamin C. SPONSORSHIP: Pascoe Pharmazeutische Präparate GmbH, Giessen, Germany.

Differences in ion-channel formation by ampullosporins B, C, D and semisynthetic desacetyltryptophanyl ampullosporin A.

Peptaibol-type ampullosporins B (2) and D (4) are capable of forming ion-conducting pores in planar lipid bilayer membrane prepared from soybean phosphatidylcholine in a similar manner as it was shown for ampullosporin A (1). However, the isomeric ampullosporin C (3) was required in 20-fold higher concentration to afford a comparable effect. In contrast to 1, 2, 3 and 4, the desacetyltryptophanyl ampullosporin A (5) failed to form ion channels. The results suggest that the sequence of amino acids especially at positions 8-10, the nitrogen-terminal acetyl residues and tryptophane are major factors determining ion-channel formation within bilayer membranes. The differences in membrane activities were comparable to the observed biological activities.

New anthracycline antibiotics produced by interspecific recombinants of streptomycetes. III. Isolation and structure of iremycin.

The structure of the anthracycline antibiotic iremycin isolated from Streptomyces violaceus subspecies iremyceticus has been elucidated as 10-(alpha-L-rhodosaminyl)-gamma-rhodomycinone (I) on the basis of spectroscopic analyses and chemical reactions.

Chrysospermins, new peptaibol antibiotics from Apiocrea chrysosperma Ap101.

Four new members of peptaibol antibiotics, designated as chrysospermins A, B, C, and D, were isolated from the mycelium of Apiocrea chrysosperma Ap101 by solvent extraction, silica gel chromatography and preparative recycling HPLC. Their structures as new nonadecapeptides were settled by detailed spectroscopic analysis and chemical degradation experiments. The chrysospermins display antibacterial and antifungal activity, and induce pigment formation by the fungus Phoma destructiva.

Ampullosporin, a new peptaibol-type antibiotic from Sepedonium ampullosporum HKI-0053 with neuroleptic activity in mice.

Ampullosporin (I; Ac-Trp-Ala-Aib-Aib-Leu-Aib-Gln-Aib-Aib-Aib-Gln-Leu-Aib-Gln-Leuol) was isolated from the mycelium of Sepedonium ampullosporum as a new 15-membered peptaibol-type antibiotic. The structure was determined by mass spectrometric and two-dimensional NMR experiments. Ampullosporin displays narrow-spectrum antibacterial and antifungal activity, induces pigment formation by Phoma destructiva, causes hypothermia and decreased spontaneous locomotor activity in mice in dosages > 1 mg/kg.

Helioferins; novel antifungal lipopeptides from Mycogone rosea: screening, isolation, structures and biological properties.

Helioferins A and B were detected as novel aminolipopeptides in cultures of Mycogone rosea DSM 8822 in the course of a screening for mediators of helianthate anion transfer from aqueous to toluene phases. Their structures as novel antibiotics and cytotoxic agents were elucidated by mass spectrometry and NMR spectroscopic methods. Antimicrobial activity was estimated against Candida albicans and Gram-positive bacteria including Mycobacterium spp.

Lipohexin, a new inhibitor of prolyl endopeptidase from Moeszia lindtneri (HKI-0054) and Paecilomyces sp. (HKI-0055; HKI-0096). I. Screening, isolation and structure elucidation.

Lipohexin was isolated as a novel lipohexapeptide (I) (C39H68N6O9) from three fungal strains, Moeszia lindtneri HKI-0054, Paecilomyces sp. HKI-0055 and Paecilomyces sp. HKI-0096. The structure was elucidated by detailed mass spectrometric and NMR experiments. The proline-containing peptide displays moderate antibacterial activity against Bacillus subtilis ATCC 6633 and inhibits competitively the prolyl endopeptidase from human placenta.

Anticoccidial efficacy of narasin in floor pen trials.

Narasin is effective against all species of chicken coccidia when tested in short-term battery cage experiments. The efficacy of narasin at concentrations of 20, 40, 60, 80, and 100 ppm was evaluated in ten floor pen trials in which commercial broiler production conditions were simulated. To provide intentional exposure to different levels of coccidia challenge, the litter of some pens was seeded with oocysts of each of the pathogenic species of chicken coccidia, whereas some pens were left nonseeded. Weight gain, feed efficiency, and lesion score data from the ten trials were analyzed as one randomized block experiment. Medication with narasin resulted in a significant reduction in lesion scores and significant improvement in weight gain and feed:gain ratios when compared with scores and gain of nonmedicated controls for both seeded and nonseeded pens. Each increase in narasin concentration up to 100 ppm for the seeded pens and up to 80 ppm for the nonseeded pens resulted in a significant reduction in cecal lesion scores. Although maximum weight gain in the seeded pens was obtained with 40 ppm narasin, concentrations greater than or equal to 60 ppm narasin were significantly better than the 40 ppm concentration in improving feed:gain ratios. These results confirm the effectiveness of narasin in controlling coccidiosis in broilers exposed to oocysts in the litter of floor pens. Furthermore, a clear relationship between the response to narasin and the level of oocyst challenge was demonstrated.

Effect of narasin and roxarsone combinations on Eimeria tenella infections in floor pen-raised broilers.

A series of four floor pen trials was conducted to evaluate the effects of narasin and roxarsone, both alone and in combination, on their capacity to control severe Eimeria tenella infections in broilers. Three levels of narasin (0, 60, and 80 ppm) were fed to chickens receiving either 0, 25, or 50 ppm roxarsone in a factorial design. Cecal coccidiosis was induced by seeding the litter with ionophore-tolerant and ionophore-sensitive strains of E. tenella. After 8 days, 10 birds/pen were killed and their cecal lesions scored. Performance (body weight and feed consumption) and mortality were measured at the termination of the trials. Narasin reduced the severity of cecal coccidiosis as measured by a reduction in cecal lesions and an improvement in bird performance. Roxarsone also reduced cecal lesion scores. The highest level of roxarsone (50 ppm) in combination with 60 or 80 ppm narasin produced additive responses in the control of E. tenella infections. Maximum performance was obtained when narasin alone was fed at 80 ppm; drug combinations improved performance when compared with that of nontreated or roxarsone only-medicated groups.

Field experience trials comparing narasin and monensin.

Narasin is effective against all species of chicken coccidia when tested in battery cage and floor pen studies. To confirm the efficacy of narasin under practical broiler production conditions, the drug was fed at concentrations of 60 ppm or 80 ppm to broiler chickens being raised by six different commercial broiler producers in five different geographic areas. Monensin was fed in each trial at a concentration of 100 ppm or 121 ppm as a reference control. The usual management practices of each of the integrated broiler companies were followed throughout the respective trials. Nine trials were conducted and approximately 100,000 broilers were tested for each treatment. No adverse reactions attributable to treatments were observed in any of the trials, and performance results obtained with narasin-medicated birds were generally comparable with those obtained with monensin-medicated birds in the same trial. These findings support the conclusion that narasin at final feed concentrations in the range of 60 to 80 ppm can be safely and effectively used as an anticoccidial agent in commercial broiler production facilities.

Differentiation between inducers of apoptosis and nonspecific cytotoxic drugs by means of cell analyzer and immunoassay.

Differentiation between specific and nonspecific cytotoxic drugs by means of cell analyzer and immunoassay is described. The determination of antiproliferative, cytotoxic, and apoptotic effects as well as enlargement of cells in response to toxic compounds can be used for the disclosure of compounds with a specific mode of action. The usefulness of the procedure was demonstrated with camptothecin (1) as a cytotoxic drug and established inducer of apoptosis. Aphidicolin (2) and the new compound oxoaphidicolin (3) were shown to display no comparable effect. Isolation and structural data of 3 are reported.

[Anthracycline antibiotics from genetically modified streptomycetes. The isolation, spectroscopic structural elucidation and biologic effects of beta-rhodomycin I]. / Anthracyclinantibiotica genetisch modifizierter Streptomyceten. Zur Isolierung, spektroskopischen Strukturaufklärung und biologischen Wirkung von beta-Rhodomycin-1.

By mutagenic treatment and selection procedures the mutant ZIMET 43678 was obtained from a population of the interspecific recombinant Streptomyces violaceus subsp. iremyceticus ZIMET 43615, which showed a changed spectrum of secondary metabolites. The main component isolated from the fermentation broth was a pure anthracycline evidenced by TLC. By means of acid hydrolysis, identification of the degradation products and also by spectroscopic UV/VIS-, IR-, MS-, 1H/13C-NMR- and CD-investigations with intact anthracycline the structure 7-(alpha-L- rhodosaminyl )-beta- rhodomycinon with the absolute configuration 7S, 9R , 10R was found. The anthracycline called beta- rhodomycin -1 (1) exhibits antimicrobial and cytostatic activity in vitro and is also effective on tumour cells in tumour bearing animals.