Ultrasonography in screening for developmental dysplasia of the hip in newborns: systematic review

Objective To assess the accuracy and effectiveness of the screening of all newborn infants for developmental dysplasia of the hip (DDH) using ultrasound imaging, as is standard practice in some European countries but not in the United Kingdom, the United States, or Scandinavia.Design Systematic review.Data sources Twenty three medical, economic, and grey literature databases (to March 2004), with no limitations of design or language; some references were provided by experts.Selection of studies Only diagnostic accuracy studies and comparative studies conducted in an unselected newborn population were eligible for the review. Two reviewers independently selected the studies and performed the quality assessment.Results The review identified one diagnostic accuracy study, and this was of limited quality. In this study the reference standard was treatment up to age of 8 months or an abnormal ultrasound finding at age 8 months. Ultrasound screening had a sensitivity of 88.5% (95% confidence interval 84.1% to 92.1%), specificity of 96.7% (96.4% to 97.4%), a positive predictive value of 61.6% and a negative predictive value of 99.4%. Ten studies evaluated the impact of ultrasound in screening, but these too had various methodological weaknesses, limiting the reliability of their findings. Compared with clinical screening, general ultrasound screening in newborns may increase overall treatment rates, but ultrasound screening seems to be associated with shorter and less intrusive treatment.Conclusions Clear evidence is lacking either for or against general ultrasound screening of newborn infants for DDH. Studies that investigate the natural course of the disorder, the optimal treatment for DDH, and the best strategy for ultrasound screening are needed.     

APC2 Cullin Protein and APC11 RING Protein Comprise the Minimal Ubiquitin Ligase Module of the Anaphase-promoting Complex

In mitosis, the anaphase-promoting complex (APC) regulates the onset of sister-chromatid separation and exit from mitosis by mediating the ubiquitination and degradation of the securin protein and mitotic cyclins. With the use of a baculoviral expression system, we have reconstituted the ubiquitin ligase activity of human APC. In combination with Ubc4 or UbcH10, a heterodimeric complex of APC2 and APC11 is sufficient to catalyze the ubiquitination of human securin and cyclin B1. However, the minimal APC2/11 ubiquitin ligase module does not possess substrate specificity, because it also ubiquitinates the destruction box deletion mutants of securin and cyclin B1. Both APC11 and UbcH10 bind to the C-terminal cullin homology domain of APC2, whereas Ubc4 interacts with APC11 directly. Zn(2+)-binding and mutagenesis experiments indicate that APC11 binds Zn(2+) at a 1:3 M ratio. Unlike the two Zn(2+) ions of the canonical RING-finger motif, the third Zn(2+) ion of APC11 is not essential for its ligase activity. Surprisingly, with Ubc4 as the E2 enzyme, Zn(2+) ions alone are sufficient to catalyze the ubiquitination of cyclin B1. Therefore, the Zn(2+) ions of the RING finger family of ubiquitin ligases may be directly involved in catalysis.     

Identification and Biological Characterization of Mouse beta-defensin 14, the orthologue of human beta-defensin 3

beta-Defensins are small antimicrobial polypeptides that are mainly expressed by epithelial cells and play an important role in the antimicrobial innate immune response. In addition to the direct microbicidal effects of these polypeptides, it became evident that certain members of the beta-defensin super family have the capacity to promote local innate inflammatory and systemic adaptive immune responses by interacting with the CC-chemokine receptor CCR6. We have identified mouse beta-defensin 14 (mBD14, Defb14) as an orthologue of human beta-defensin 3 (hBD3 or DEFB103). Based on primary structural analysis, mBD14 demonstrates greater (68%) homology to its human orthologue, containing three conserved cystein linkages, characteristic for the beta-defensin super family. mBD14 is expressed in a wide variety of tissues including spleen, colon, and tissues of the upper and lower respiratory tract. Interestingly, we also detected mBD14 expression in immature CD11c+ bone marrow-derived dendritic cells. The expression of mBD14 can be induced by Toll-like receptor agonists such as lipopolysaccharide and poly(I:C) and by pro-inflammatory stimuli e.g. tumor necrosis factor and interferon-gamma. Furthermore, expression of mBD14 seems to be regulated by activation of the intracellular pattern recognition receptor NOD2/CARD15 as revealed by reporter gene analysis. We prepared a recombinant mBD14-Ig fusion protein that retained potent antimicrobial activity against several Escherichia coli strains but not against various Gram-positive Staphylococcus aureus strains. hBD3 and also the newly identified mBD14 were chemotactic for cells expressing the mouse CC-chemokine receptor CCR6. In addition, both hBD3 and mBD14 were chemotactic for freshly isolated mouse resident peritoneal cells. Thus, mBD14, based on structural and functional similarities, appears to be an orthologue of hBD3.     

Reovirus activates human dendritic cells to promote innate antitumor immunity

Oncolytic viruses can exert their antitumor activity via direct oncolysis or activation of antitumor immunity. Although reovirus is currently under clinical investigation for the treatment of localized or disseminated cancer, any potential immune contribution to its efficacy has not been addressed. This is the first study to investigate the ability of reovirus to activate human dendritic cells (DC), key regulators of both innate and adaptive immune responses. Reovirus induced DC maturation and stimulated the production of the proinflammatory cytokines IFN-alpha, TNF-alpha, IL-12p70, and IL-6. Activation of DC by reovirus was not dependent on viral replication, while cytokine production (but not phenotypic maturation) was inhibited by blockade of PKR and NF-kappaB signaling. Upon coculture with autologous NK cells, reovirus-activated DC up-regulated IFN-gamma production and increased NK cytolytic activity. Moreover, short-term coculture of reovirus-activated DC with autologous T cells also enhanced T cell cytokine secretion (IL-2 and IFN-gamma) and induced non-Ag restricted tumor cell killing. These data demonstrate for the first time that reovirus directly activates human DC and that reovirus-activated DC stimulate innate killing by not only NK cells, but also T cells, suggesting a novel potential role for T cells in oncolytic virus-induced local tumor cell death. Hence reovirus recognition by DC may trigger innate effector mechanisms to complement the virus's direct cytotoxicity, potentially enhancing the efficacy of reovirus as a therapeutic agent.     

Effects of high-intensity training on muscle lactate transporters and postexercise recovery of muscle lactate and hydrogen ions in women

The purpose of this study was to investigate the effects of high-intensity interval training (3 days/wk for 5 wk), provoking large changes in muscle lactate and pH, on changes in intracellular buffer capacity (betam(in vitro)), monocarboxylate transporters (MCTs), and the decrease in muscle lactate and hydrogen ions (H+) after exercise in women. Before and after training, biopsies of the vastus lateralis were obtained at rest and immediately after and 60 s after 45 s of exercise at 190% of maximal O2 uptake. Muscle samples were analyzed for ATP, phosphocreatine (PCr), lactate, and H+; MCT1 and MCT4 relative abundance and betam(in vitro) were also determined in resting muscle only. Training provoked a large decrease in postexercise muscle pH (pH 6.81). After training, there was a significant decrease in betam(in vitro) (-11%) and no significant change in relative abundance of MCT1 (96 +/- 12%) or MCT4 (120 +/- 21%). During the 60-s recovery after exercise, training was associated with no change in the decrease in muscle lactate, a significantly smaller decrease in muscle H+, and increased PCr resynthesis. These results suggest that increases in betam(in vitro) and MCT relative abundance are not linked to the degree of muscle lactate and H+ accumulation during training. Furthermore, training that is very intense may actually lead to decreases in betam(in vitro). The smaller postexercise decrease in muscle H+ after training is a further novel finding and suggests that training that results in a decrease in H+ accumulation and an increase in PCr resynthesis can actually reduce the decrease in muscle H+ during the recovery from supramaximal exercise.     

Mycophenolic Acid in Silage

We examined 233 silage samples and found that molds were present in 206 samples with counts between 1 × 10³ and 8.9 × 10⁷ (mean, 4.7 × 10⁶) CFU/g. Mycophenolic acid, a metabolite of Penicillium roqueforti, was detected by liquid chromatography-mass spectrometry in 74 (32%) of these samples at levels ranging from 20 to 35,000 (mean, 1,400) μg/kg. This compound has well-known immunosuppressive properties, so feeding with contaminated silage may promote the development of infectious diseases in livestock.     

Protein-protein interactions of tandem affinity purification-tagged protein kinases in rice

Forty-one rice cDNAs encoding protein kinases were fused to the tandem affinity purification (TAP) tag and expressed in transgenic rice plants. The TAP-tagged kinases and interacting proteins were purified from the T1 progeny of the transgenic rice plants and identified by mass spectrometry. Ninety-five percent of the TAP-tagged kinases were recovered. Fifty-six percent of the TAP-tagged kinases were found to interact with other rice proteins. A number of these interactions were consistent with known protein complexes found in other species, validating the TAP-tag method in rice plants. Phosphorylation sites were identified on four of the kinases that interacted with either 14-3-3 proteins or cyclins.     

Microbubble-enhanced ultrasound to deliver an antisense oligodeoxynucleotide targeting the human androgen receptor into prostate tumours

We have shown recently that downregulation of the androgen receptor (AR), one of the key players in prostate tumor cells, with short antisense oligodeoxynucleotides (ODNs) results in inhibition of prostate tumor growth. Particularly with regard to an application of these antisense drugs in vivo, we now investigated the usefulness of microbubble-enhanced ultrasound to deliver these ODNs into prostate cancer cells.

Our short antisense AR ODNs were loaded onto the lipid surface of cationic gas-filled microbubbles by ion charge binding, and delivered into the cells by bursting the loaded microbubbles with ultrasound. In vitro experiments were initially performed to show that this kind of delivery system works in principle. In fact, transfection of prostate tumor cells with antisense AR ODNs using microbubble-enhanced ultrasound resulted in 49% transfected cells, associated with a decrease in AR expression compared to untreated controls. In vivo, uptake of a digoxigenin-labelled ODN was found in prostate tumour xenografts in nude mice following intratumoral or intravenous injection of loaded microbubbles and subsequent exposure of the tumour to ultrasound, respectively. Our results show that ultrasound seems to be the driving force of this delivery system. Uptake of the ODN was also observed in tumors after treatment with ultrasound alone, with only minor differences compared to the combined use of microbubbles and ultrasound.     

Novel boronated derivatives of 5,10,15,20-tetraphenylporphyrin: synthesis and toxicity for drug-resistant tumor cells

We have developed the synthesis of boronated porphyrins for potential application in cancer treatment, based on the functional derivatives of 5,10,15,20-tetraphenylporphyrin. Boronated amide derivatives starting from 5,10,15,20-tetra(p-aminophenyl)porphyrin and 9-o- and 9-m-carborane carboxylic acid chlorides were prepared. Also, the reaction of 2-formyl-5,10,15,20-tetraphenylporphyrin with closo-C-lithium-o- and m-carboranes, as well as with closo-C-lithium monocarbon carborane, yielded neutral and anionic boronated hydroxy derivatives of 5,10,15,20-tetraphenylporphyrin, respectively. Water-soluble forms of neutral compounds were prepared by deboronation of closo-polyhedra with Bu4NF into nido-7,8- and nido-7,9-dicarbaundecaborate anions. Monocarbon carborane conjugated with copper (II) complex of 5,10,15,20-tetraphenylporphyrin was active for a variety of tumor cell lines (IC50 approximately 5 microM after 48-72 h of exposure) but was inert for non-malignant fibroblasts at up to 100 microM. At low micromolar concentrations, this compound caused the death of cells that express P-glycoprotein and other mechanisms of resistance to conventional anticancer drugs.     

Effects of age and environmental factors on semen production and semen quality of Austrian Simmental bulls

More than 90% of the breeding stock of Austrian dual purpose Simmental cows is artificially inseminated. Knowledge of factors affecting sperm production and semen quality is of importance with regard to reproductive efficiency and thus genetic improvement as well as for the productivity and profitability of AI centers. Hence, semen data from two Austrian AI centres collected in the years 2000 and 2001 were evaluated. In total, 3625 and 3654 ejaculates from 147 and 127 AI bulls, respectively, were analysed regarding ejaculate volume, sperm concentration, percentage of viable spermatozoa in the ejaculate, total spermatozoa per ejaculate and motility. Effects accounted for were the bull (random), age of bull, collection interval, number of collection on collection day, bull handler, semen collector, temperature on day of semen collection, in the course of epididymal maturation (average temperature of days 1-11 before collection) and during spermatogenesis (average temperature of days 12-65 before collection). Age of bull significantly affected all traits (P<0.01 to P<0.001) except motility score in center 2. Ejaculate volume and total number of spermatozoa increased with age of bull while sperm concentration was lower in higher age classes (center 1). The collection team was also found to significantly influence semen quality traits. With increasing collection interval ejaculate volume and total number of spermatozoa increased significantly (P<0.05 to P<0.001) while collection intervals between 4-9 days and 1-6 days were superior with regard to sperm concentration and percentage of viable spermatozoa, respectively (P<0.10 to P<0.001). First ejaculates were superior with respect to ejaculate volumes, sperm concentrations and total number of spermatozoa per ejaculate (P<0.001). Temperature, either on day of semen collection or during epididymal maturation or spermatogenesis, had important but inconsistent effects on semen production and sperm quality. Overall, however, ambient temperatures in the range of 5-15 degrees C were found to be optimal for semen production.