Untreated silicone breast implant rupture

Implant rupture is a well-known complication of breast implant surgery that can pass unnoticed by both patient and physician. To date, no prospective study has addressed the possible health implications of silicone breast implant rupture. The aim of the present study was to evaluate whether untreated ruptures are associated with changes over time in magnetic resonance imaging findings, serologic markers, or self-reported breast symptoms. A baseline magnetic resonance imaging examination was performed in 1999 on 271 women who were randomly chosen from a larger cohort of women having cosmetic breast implants for a median period of 12 years (range, 3 to 25 years). A follow-up magnetic resonance imaging examination was carried out in 2001, excluding women who underwent explantation in the period between the two magnetic resonance imaging examinations (n = 44). On the basis of these examinations, the authors identified 64 women who had at least one ruptured implant at the first magnetic resonance imaging examination and, for comparison, all women who had intact implants at both examinations (n = 98). Magnetic resonance images from the two examinations were compared and changes in rupture configuration were evaluated. Comparisons were also made for self-reported breast symptoms occurring during the study period and for changes in serum values of antinuclear antibodies, rheumatoid factor, and cardiolipin antibodies immunoglobulin G and immunoglobulin M. The majority of the women with implant rupture had no visible magnetic resonance imaging changes of their ruptured implants. For 11 implants (11 percent) in 10 women, the authors observed progression of silicone seepage, either as a conversion from intracapsular into extracapsular rupture (n = 7), as progression of extra-capsular silicone (n = 3), or as increasing herniation of the silicone within the fibrous capsule (n = 1); however, in most cases, these changes were minor. Some changes could be ascribed to trauma, but others seemed spontaneous. There was no increase in levels of autoantibodies during the study period in either study group. Women with untreated implant ruptures reported a significant increase in nonspecific breast changes (odds ratio, 2.1; 95 percent confidence interval, 1.2 to 3.8) compared with women without ruptures. On the basis of this first study of women with untreated silicone breast implant rupture, the authors conclude that implant rupture is a relatively harmless condition, which only rarely progresses and gives rise to notable symptoms. Even so, because of a small risk of silicone spread, the authors suggest that women with implant ruptures be followed clinically, if not operated on. Because implant ruptures often occur asymptomatically, any woman with silicone implants, regardless of rupture status, should be evaluated at regular intervals.     

Transformation of Leuconostoc carnosum 4010 and evidence for natural competence of the organism

Plasmid transformation in Leuconostoc carnosum 4010 was analyzed. A successful transformation protocol for L. carnosum was established by modifying an existing protocol for Lactococcus lactis. Several parameters, including the number of generations that the cells had grown at the time of harvest, glycine concentration, the time of incubation for phenotypic expression, and the electrical field strength, were investigated and proved to have influence on the transformation frequency. Electrocompetence was found to be transient and to peak in the early exponential growth phase. Optimized conditions resulted in transformation frequencies of up to 6.7 x 10(5) transformants per microgram of plasmid DNA. A total of five plasmids in L. carnosum were successfully introduced and maintained. Interestingly, we discovered that DNA uptake was of a frequency of 3 x 10(-6) to 19 x 10(-6) transformants per CFU in the absence of an applied electrical field. We concluded that L. carnosum is naturally competent.     

Elucidation of the antibacterial mechanism of the Curvularia haloperoxidase system by DNA microarray profiling

A novel antimicrobial enzyme system, the Curvularia haloperoxidase system, was examined with the aim of elucidating its mechanism of antibacterial action. Escherichia coli strain MG1655 was stressed with sublethal concentrations of the enzyme system, causing a temporary arrest of growth. The expression of genes altered upon exposure to the Curvularia haloperoxidase system was analyzed by using DNA microarrays. Only a limited number of genes were involved in the response to the Curvularia haloperoxidase system. Among the induced genes were the ibpA and ibpB genes encoding small heat shock proteins, a gene cluster of six genes (b0301-b0306) of unknown function, and finally, cpxP, a member of the Cpx pathway. Knockout mutants were constructed with deletions in b0301-b0306, cpxP, and cpxARP, respectively. Only the mutant lacking cpxARP was significantly more sensitive to the enzyme system than was the wild type. Our results demonstrate that DNA microarray technology cannot be used as the only technique to investigate the mechanisms of action of new antimicrobial compounds. However, by combining DNA microarray analysis with the subsequent creation of knockout mutants, we were able to pinpoint one of the specific responses of E. coli--namely, the Cpx pathway, which is important for managing the stress response from the Curvularia haloperoxidase system.     

Immunolocalization of AQP-5 in rat parotid and submandibular salivary glands after stimulation or inhibition of secretion in vivo

In vitro studies of cultured salivary gland cells and gland slices have indicated that there may be regulated translocation of aquaporin (AQP)-5 between the apical plasma membrane and intracellular compartments of the secretory cells. However, it remains unknown whether AQP-5 in salivary glands is subject to regulated trafficking in vivo. To examine this possibility, we have investigated the subcellular localization of AQP-5 in rat parotid and submandibular glands fixed in vivo under conditions of stimulated or inhibited salivary secretion. Immunofluorescence and immunoelectron microscopy was used to determine the subcellular distribution of AQP-5 in control conditions following the stimulation of secretion with pilocarpine (a muscarinic agonist) or epinephrine (an alpha-adrenoceptor agonist) or during inhibition of basal secretion with atropine (a muscarinic antagonist) or phentolamine (an alpha-adrenoceptor antagonist). Under control conditions, >90% of AQP-5 was associated with the apical plasma membrane of acinar and intercalated duct cells, with only rare gold particles associated with intracellular membrane domains. Pilocarpine treatment dramatically increased saliva production but had no discernible effect on AQP-5 distribution. However, the increased salivary secretion was associated with luminal dilation and the appearance of a markedly punctate AQP-5 labeling pattern due to clustering of AQP-5 at the microvilli (especially evident in the parotid gland) after 10 min of drug injection. No changes in the subcellular localization of AQP-5 were seen in response to epinephrine, atropine, or phentolamine treatment compared with control tissues. Thus AQP-5 is localized predominantly in the apical plasma membrane under control conditions, and neither the onset nor the cessation of secretion is associated in vivo with any significant short-term translocation of AQP-5 between intracellular structures and the apical plasma membrane.     

Molecular classification of liver cirrhosis in a rat model by proteomics and bioinformatics

Liver cirrhosis is a worldwide health problem. Reliable, noninvasive methods for early detection of liver cirrhosis are not available. Using a three-step approach, we classified sera from rats with liver cirrhosis following different treatment insults. The approach consisted of: (i) protein profiling using surface-enhanced laser desorption/ionization (SELDI) technology; (ii) selection of a statistically significant serum biomarker set using machine learning algorithms; and (iii) identification of selected serum biomarkers by peptide sequencing. We generated serum protein profiles from three groups of rats: (i) normal (n=8), (ii) thioacetamide-induced liver cirrhosis (n=22), and (iii) bile duct ligation-induced liver fibrosis (n=5) using a weak cation exchanger surface. Profiling data were further analyzed by a recursive support vector machine algorithm to select a panel of statistically significant biomarkers for class prediction. Sensitivity and specificity of classification using the selected protein marker set were higher than 92%. A consistently down-regulated 3495 Da protein in cirrhosis samples was one of the selected significant biomarkers. This 3495 Da protein was purified on-chip and trypsin digested. Further structural characterization of this biomarkers candidate was done by using cross-platform matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) peptide mass fingerprinting (PMF) and matrix-assisted laser desorption/ionization time of flight/time of flight (MALDI-TOF/TOF) tandem mass spectrometry (MS/MS). Combined data from PMF and MS/MS spectra of two tryptic peptides suggested that this 3495 Da protein shared homology to a histidine-rich glycoprotein. These results demonstrated a novel approach to discovery of new biomarkers for early detection of liver cirrhosis and classification of liver diseases.     

Prediction of post-translational glycosylation and phosphorylation of proteins from the amino acid sequence

Post-translational modifications (PTMs) occur on almost all proteins analyzed to date. The function of a modified protein is often strongly affected by these modifications and therefore increased knowledge about the potential PTMs of a target protein may increase our understanding of the molecular processes in which it takes part. High-throughput methods for the identification of PTMs are being developed, in particular within the fields of proteomics and mass spectrometry. However, these methods are still in their early stages, and it is indeed advantageous to cut down on the number of experimental steps by integrating computational approaches into the validation procedures. Many advanced methods for the prediction of PTMs exist and many are made publicly available. We describe our experiences with the development of prediction methods for phosphorylation and glycosylation sites and the development of PTM-specific databases. In addition, we discuss novel ideas for PTM visualization (exemplified by kinase landscapes) and improvements for prediction specificity (by using ESS--evolutionary stable sites). As an example, we present a new method for kinase-specific prediction of phosphorylation sites, NetPhosK, which extends our earlier and more general tool, NetPhos. The new server, NetPhosK, is made publicly available at the URL http://www.cbs.dtu.dk/services/NetPhosK/. The issues of underestimation, over-prediction and strategies for improving prediction specificity are also discussed.     

Real-time process/quality control for HPC processing

BACKGROUND: JACIE Standards (FACT Standards in the USA) have been implemented in Europe since 1999. An on-site accreditation inspection took place at our center in January 2004. The purpose of this work was to develop a real-time process/quality control system meeting the JACIE Standards for HPC release. METHODS: Data from 194 HPC processing procedures for autologous transplantation performed over a 5-year period were analyzed. The results of different processing methods applied at our facility were compared: (1) cryopreservation without washing cells (n=50), (2) washing cells (n=87), (3) cell-density separation (n=12) and (4) positive CD34 selection (n=45). RESULTS: Four critical control points were set for the validation of HPC processing: (a) number of lost CD34(+) cells during processing, (b) contamination, (c) viability of the cells after thawing and (d) ability to reconstitute hematopoiesis after transplantation. On the basis of statistical analysis, ranges of acceptable values were defined for each critical control point and for each processing method. Those acceptable values were used for cell release and real-time quality control. DISCUSSION: This study describes a model for the validation of HPC processing and for a real-time process/quality control system for HPC release. Optimization of processing techniques, standardization of methods and comparison between facilities will open the way towards external quality controls and quality improvement.     

Identification of a core set of genes that signifies pathways underlying cardiac hypertrophy

Although the molecular signals underlying cardiac hypertrophy have been the subject of intense investigation, the extent of common and distinct gene regulation between different forms of cardiac hypertrophy remains unclear. We hypothesized that a general and comparative analysis of hypertrophic gene expression, using microarray technology in multiple models of cardiac hypertrophy, including aortic banding, myocardial infarction, an arteriovenous shunt and pharmacologically induced hypertrophy, would uncover networks of conserved hypertrophy-specific genes and identify novel genes involved in hypertrophic signalling. From gene expression analyses (8740 probe sets, n = 46) of rat ventricular RNA, we identified a core set of 139 genes with consistent differential expression in all hypertrophy models as compared to their controls, including 78 genes not previously associated with hypertrophy and 61 genes whose altered expression had previously been reported. We identified a single common gene program underlying hypertrophic remodelling, regardless of how the hypertrophy was induced. These genes constitute the molecular basis for the existence of one main form of cardiac hypertrophy and may be useful for prediction of a common therapeutic approach. Supplementary material for this article can be found at: http://www.interscience.wiley.com/jpages/1531-6912/suppmat.     

Influence of triacontanol on somatic embryogenesis in <Emphasis Type="Italic">Coffea arabica</Emphasis> L. and <Emphasis Type="Italic">Coffea canephora</Emphasis> P. ex Fr.

Summary A highly reproducible method for regeneration of Coffea arabica and C. canephora plants via direct somatic embryogenesis from cultured leaf and stem segments of regenerated plants was developed. Embryogenesis was influenced by the presence of triacontanol (TRIA) in the medium. TRIA incorporated at 4.55 and 11.38 μM in half-strength MS basal medium containing 1.1 μM 6-benzyladenine (BA) and 2.28 μM indole-3-acetic acid (IAA) induced direct somatic embryogenesis in both species. A maximum of 260±31.8 and 59.2±12.8 somatic embryos per culture were induced from in vitro leaf explants of C. arabica and C. canephora, respectively. TRIA also induced embryo formation from in vitro stem segment callus tissues along with multiplication of primary embryos into secondary embryos. By using TRIA, it was possible to obtain somatic embryogenesis in C. arabica and C. canephora.