The Histological Composition of Capsular Contracture Focussed on the Inner Layer of the Capsule: An Intra-Donor Baker-I Versus Baker-IV Comparison.

BACKGROUND: Capsular contracture remains one of the major complications after breast implantation surgery. The extent of capsular contraction is scored using the Baker scale. The aim of this study was to compare intra-individual Baker-I with Baker-IV capsules, and in particular the prevalence and histological properties of the inner capsule layer. METHODS: Twenty capsules from ten patients were included after bilateral explantation surgery due to unilateral capsular contracture (Baker-IV) after cosmetic augmentation with textured implants. All capsules underwent (immune-)histochemical analysis: haematoxylin-eosin (morphology), CD68 (macrophages), cytokeratin (epithelial cells) and vimentin (fibroblasts), and were visually scored for cell density and the presence of an inner layer and measured for thickness. RESULTS: Baker-IV (n = 10) capsules were significantly thicker compared to Baker-I (n = 10) capsules (P = 0.004). An inner layer was present in 8 Baker-I capsules. All Baker-I capsules were vimentin and CD68-positive and cytokeratin-negative. Positive vimentin was seen throughout the inner layer, and CD-68 staining was observed adjacent to the intermediate capsule layer. In contrast, only 2 Baker-IV capsules had an inner layer, of which only 1 showed the same profile as Baker-I capsules (P = 0.016). No cytokeratin positivity was seen in any capsule. In Baker-IV capsules, outer layers showed more positivity for both vimentin and CD68. CONCLUSION: The inner layer is morphologically consistent with synovial metaplasia and is more prevalent in healthy, uncontracted Baker-I capsules. This inverse relation between the presence of the inner layer and higher Baker classification or pathological contracture could indicate a protective role of the inner layer against capsular contracture formation. LEVEL OF EVIDENCE III: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Techno-Functional Properties of Crude Extracts from the Green Microalga Tetraselmis suecica.

A mild fractionation process to extract functional biomolecules from green microalgae was implemented. The process includes bead milling, centrifugation, and filtration with several membrane cut-offs. For each fraction, the corresponding composition was measured, and the surface activity and gelation behavior were determined. A maximum protein yield of 12% was obtained in the supernatant after bead milling and between 3.2 and 11.7% after filtration. Compared to whey protein isolate, most of the algae fractions exhibited comparable or enhanced functionality. Surface activity for air-water and oil-water interfaces and gelation activities were notably superior for the retentate fractions compared to the permeates. It is proposed that such functionality in the retentates is due to the presence of hydrophobic compounds and molecular complexes exhibiting a similar behavior as Pickering particles. We demonstrated that excellent functionality can be obtained with crude fractions, requiring minimum processing and, thus, constituting an interesting option for commercial applications.

Energy consumption and water-soluble protein release by cell wall disruption of Nannochloropsis gaditana.

Several cell disruption methods were tested on Nannochloropsis gaditana, to evaluate their efficiency in terms of cell disintegration, energy input and release of soluble proteins. High-pressure homogenization (HPH) and bead milling were the most efficient with >95% cell disintegration, ±50% (w/w) release of total proteins and low energy input (<0.5kWh.kg-1biomass). Enzymatic treatment required low energy input (<0.34kWh.kg-1biomass), but it only released ±35% protein (w/w). Pulsed Electric Field (PEF) was neither energy-efficient (10.44kWh.kg-1biomass) nor successful for protein release (only 10% proteins w/w) and cell disintegration. The release of proteins after applying HPH and bead milling always required less intensive operating conditions for cell disruption. The energy cost per unit of released protein ranged from 0.15-0.25 €.kgProtein-1 in case of HPH, and up to 2-20 €.kgProtein-1 in case of PEF.

Increased epidermal thickness and abnormal epidermal differentiation in keloid scars.

BACKGROUND: The pathogenesis underlying keloid formation is still poorly understood. Research has focused mostly on dermal abnormalities, while the epidermis has not yet been studied. OBJECTIVES: To identify differences within the epidermis of mature keloid scars compared with normal skin and mature normotrophic and hypertrophic scars. METHODS: Rete ridge formation and epidermal thickness were evaluated in tissue sections. Epidermal proliferation was assessed using immunohistochemistry (Ki67, keratins 6, 16 and 17) and with an in vitro proliferation assay. Epidermal differentiation was evaluated using immunohistochemistry (keratin 10, involucrin, loricrin, filaggrin, SPRR2, SKALP), reverse-transcriptase polymerase chain reaction (involucrin) and transmission electron microscopy (stratum corneum). RESULTS: All scars showed flattening of the epidermis. A trend of increasing epidermal thickness correlating to increasing scar abnormality was observed when comparing normal skin, normotrophic scars, hypertrophic scars and keloids. No difference in epidermal proliferation was observed. Only the early differentiation marker involucrin showed abnormal expression in scars. Involucrin was restricted to the granular layer in healthy skin, but showed panepidermal expression in keloids. Normotrophic scars expressed involucrin in the granular and upper spinous layers, while hypertrophic scars resembled normotrophic scars or keloids. Abnormal differentiation was associated with ultrastructural disorganization of the stratum corneum in keloids compared with normal skin. CONCLUSIONS: Keloids showed increased epidermal thickness compared with normal skin and normotrophic and hypertrophic scars. This was not due to hyperproliferation, but possibly caused by abnormal early terminal differentiation, which affects stratum corneum formation. Our findings indicate that the epidermis is associated with keloid pathogenesis and identify involucrin as a potential diagnostic marker for abnormal scarring.

Biorefinery of microalgal soluble proteins by sequential processing and membrane filtration.

A mild biorefinery process was investigated on the microalga Nannochloropsis gaditana, to obtain an enriched fraction of water soluble proteins free from chlorophyll. After harvesting, a 100g.L-1 solution of cells was first subjected to cell disruption by either high-pressure homogenization (HPH) or enzymatic treatment (ENZ). HPH resulted in a larger release of proteins (49%) in the aqueous phase compared to the Alcalase incubation (35%). In both cases, an ultrafiltration/diafiltration (UF/DF) was then performed on the supernatant obtained from cell disruption by testing different membrane cut-off (1000kDa, 500kDa and 300kDa). After optimising the process conditions, the combination of ENZ→UF/DF ended in a larger overall yield of water soluble proteins (24.8%) in the permeate compared to the combination of HPH→UF/DF (17.4%). A gel polarization model was implemented to assess the maximum achievable concentration factor during ultrafiltration and the mass transfer coefficient related to the theoretical permeation flux rate.

Cationic polymers for successful flocculation of marine microalgae.

Flocculation of microalgae is a promising technique to reduce the costs and energy required for harvesting microalgae. Harvesting marine microalgae requires suitable flocculants to induce the flocculation under marine conditions. This study demonstrates that cationic polymeric flocculants can be used to harvest marine microalgae. Different organic flocculants were tested to flocculate Phaeodactylum tricornutum and Neochloris oleoabundans grown under marine conditions. Addition of 10 ppm of the commercial available flocculants Zetag 7557 and Synthofloc 5080H to P. tricornutum showed a recovery of, respectively, 98% ± 2.0 and 94% ± 2.9 after flocculation followed by 2h sedimentation. Using the same flocculants and dosage for harvesting N. oleoabundans resulted in a recovery of 52% ± 1.5 and 36% ± 11.3. This study shows that cationic polymeric flocculants are a viable option to pre-concentrate marine cultivated microalgae via flocculation prior to further dewatering.

[Prevention of perioperative cardiac complications in non-cardiac surgery: an evidence-based guideline]. / De preventie van perioperatieve cardiale complicaties bij niet-cardiale chirurgie: een evidence-based richtlijn.

Approximately 2.5% of the patients undergoing non-cardiac surgery suffer from perioperative cardiac complications. These are associated with a mortality of 20.60%, a longer stay in hospital and higher costs. The risk factors for perioperative cardiac complications are: high-risk surgery, ischaemic heart disease, a history of congestive heart failure, cerebrovascular disease, diabetes, and renal failure. Recently, the scope of medical management has shifted from assessing and treating underlying culprit coronary lesions toward coronary plaque stabilisation and prevention of myocardial oxygen supply demand mismatch. Currently, the prevention of cardiac problems consists of identification of the patients at risk, optimisation of the preoperatieve condition by modification of underlying risk factors, optimisation of the perioperative medication with adrenergic beta-antagonists, statins, and acetylsalicylic acid, adequate perioperative monitoring and measures to prevent myocardial ischaemia. These include adequate sedation and analgesia, adequate oxygenation, oxygen transport, and ventilation, and if necessary additional cardiac medication.

Development of quality indicators for diagnosis and treatment of patients with non-small cell lung cancer: a first step toward implementing a multidisciplinary, evidence-based guideline.

BACKGROUND: While developing and distributing clinical practice guidelines are important in optimising clinical healthcare, insight into actual care is necessary to achieve successful implementation. Developing quality indicators may be the first step to becoming aware of actual care. The Dutch national practice guideline Non-small cell lung cancer: staging and treatment is one of the first clinical, multidisciplinary guidelines for oncology in the Netherlands for which quality indicators were developed systematically. We describe indicator development based on this guideline as a practical experience. METHODS: To develop a set of indicators for diagnosis and treatment of patients with non-small cell lung cancer, we systematically achieved consensus on the basis of a national, multidisciplinary, evidence-based guideline and the opinions of professionals and patients. After the researchers extracted the recommendations from the guideline, we carried out a so-called Rand-modified-Delphi procedure. This consisted of three rounds: a national panel of professionals and representatives of the national patient organization scored all recommendations, the professionals had a consensus meeting, and the final set of indicators was e-mailed for a last check. Subsequently, some clinimetric characteristics of this final set were assessed in a practice test. RESULTS: Thirty-two of 83 recommendations were selected in the first round. After the consensus meeting, 8 recommendations met the final criteria and were incorporated into 15 indicators, which were tested in practice. The most successful indicators for quality improvement are indicators that are measurable, have potential for improvement, have a broad range between practices and are applicable to a large part of the population. CONCLUSIONS: For successful implementation of evidence-based guidelines, each new guideline should be developed and tested with a set of indicators based on the guideline. The procedure we describe can serve as an example for other new guidelines.

Fat digestion in veal calves fed milk replacers low or high in calcium and containing either casein or soy protein isolate.

The hypothesis tested was that the inhibitory effect of dietary soy protein versus casein on fat digestion in veal calves would be smaller when diets were fed with high instead of low calcium content. Male calves, 1 wk of age, were fed 1 of 4 experimental milk replacers in a 2 x 2 factorial design. There were 19 animals per dietary group. The milk replacers contained either casein or soy protein isolate as variable protein source and were either low or high in calcium. Body weight gain was not significantly affected by the experimental diets. Soy protein isolate versus casein significantly reduced apparent fat digestibility. High versus low calcium intake also depressed fat digestion. The protein effect was smaller (2.9% units) for the high than the low calcium diets (3.6% units), but the interaction did not reach statistical significance. Soy protein isolate versus casein raised fecal bile acid excretion and so did high versus low calcium intake. The difference in bile acid excretion between the soy and casein containing diets was significantly greater for the high than low calcium diets. The absorption of phosphorus, calcium, and magnesium was higher for the casein diets than for the soy-containing diets. This study shows for the first time that soy protein isolate versus casein depressed fat digestion and raised fecal bile acid excretion in veal calves.

Overexpression of P70 S6 kinase protein is associated with increased risk of locoregional recurrence in node-negative premenopausal early breast cancer patients.

The RPS6KB1 gene is amplified and overexpressed in approximately 10% of breast carcinomas and has been found associated with poor prognosis. We studied the prognostic significance of P70 S6 kinase protein (PS6K) overexpression in a series of 452 node-negative premenopausal early-stage breast cancer patients (median follow-up: 10.8 years). Immunohistochemistry was used to assess PS6K expression in the primary tumour, which had previously been analysed for a panel of established prognostic factors in breast cancer. In a univariate analysis, PS6K overexpression was associated with worse distant disease-free survival as well as impaired locoregional control (HR 1.80, P 0.025 and HR 2.50, P 0.006, respectively). In a multivariate analysis including other prognostic factors, PS6K overexpression remained an independent predictor for poor locoregional control (RR 2.67, P 0.003). To our knowledge, P70 S6 kinase protein is the first oncogenic marker that has prognostic impact on locoregional control and therefore may have clinical implications in determining the local treatment strategy in early-stage breast cancer patients.

Physico-chemical and transglucosylation properties of recombinant sucrose phosphorylase from Bifidobacterium adolescentis DSM20083.

Clones of a genomic library of Bifidobacterium adolescentis were grown in minimal medium with sucrose as sole carbon source. An enzymatic fructose dehydrogenase assay was used to identify sucrose-degrading enzymes. Plasmids were isolated from the positive colonies and sequence analysis revealed that two types of insert were present, which only differed with respect to their orientation in the plasmid. An open reading frame of 1,515 nucleotides with high homology for sucrose phosphorylases was detected on these inserts. The gene was designated SucP and encoded a protein of 56,189 Da. SucP was heterologously expressed in Escherichia coli, purified, and characterized. The molecular mass of SucP was 58 kDa, as estimated by SDS-PAGE, while 129 kDa was found with gel permeation, suggesting that the native enzyme was a dimer. The enzyme showed high activity towards sucrose and a lower extent towards alpha-glucose-1-phosphate. The transglucosylation properties were investigated using a broad range of monomeric sugars as acceptor substrate for the recombinant enzyme, while alpha-glucose-1-phosphate served as donor. D- and L-arabinose, D- and L-arabitol, and xylitol showed the highest production of transglucosylation products. The investigated disaccharides and trisaccharides were not suitable as acceptors. The structure of the transglucosylation product obtained with D-arabinose as acceptor was elucidated by NMR. The structure of the synthesized non-reducing dimer was alpha-Glcp(1-->1)beta-Araf.

Nuclear beta-catenin is a molecular feature of type I endometrial carcinoma.

Two types of endometrial carcinoma can be distinguished: type I tumours, which are oestrogen-related and are typically low-grade endometrioid carcinomas; and type II tumours, which are unrelated to oestrogen stimulation and are often non-endometrioid carcinomas. The molecular abnormalities involved in carcinogenesis appear to be different for these tumour types. The aim of this study was to test the hypothesis that an abnormality in the Wnt/beta-catenin signalling pathway is a molecular feature of type I endometrial carcinoma. This study investigated nuclear beta-catenin by immunohistochemistry in 233 endometrial carcinomas and analysed its correlation with several immunohistochemical, histological, and clinical parameters, such as proliferation rate (Ki-67), expression of oestrogen and progesterone receptors, and survival. Nuclear beta-catenin expression was observed in 39 cases (16%). All tumours expressing nuclear beta-catenin were endometrioid adenocarcinomas, were significantly better differentiated, and were more often hormone receptor-positive than tumours without nuclear beta-catenin. No correlation with proliferation rate was found. It was found that several features of type I endometrial carcinoma occur significantly more often in tumours expressing nuclear beta-catenin, suggesting that an abnormality in the Wnt/beta-catenin signalling pathway, resulting in nuclear beta-catenin immunopositivity, is a molecular feature of a subset of type I endometrial carcinomas.

Cloning and characterization of two alpha-glucosidases from Bifidobacterium adolescentis DSM20083.

Two alpha-glucosidase encoding genes (aglA and aglB) from Bifidobacterium adolescentis DSM 20083 were isolated and characterized. Both alpha-glucosidases belong to family 13 of the glycosyl hydrolases. Recombinant AglA (EC 3.2.1.10) and AglB (EC 3.2.1.20), expressed in Escherichia coli, showed high hydrolytic activity towards isomaltose and pnp-alpha-glucoside. The K(m) for pnp-alpha-glucoside was 1.05 and 0.47 mM and the V(max) was 228 and 113 U mg(-1) for AglA and AglB, respectively. Using pnp-alpha-glucoside as substrate, the pH optimum for AglA was 6.6 and the temperature optimum was 37 degrees C. For AglB, values of pH 6.8 and 47 degrees C were found. AglA also showed high hydrolytic activity towards isomaltotriose and, to a lesser extent, towards trehalose. AglB has a high preference for maltose and less activity towards sucrose; minor activity was observed towards melizitose, low molecular weight dextrin, maltitol, and maltotriose. The recombinant alpha-glucosidases were tested for their transglucosylation activity. AglA was able to synthesize oligosaccharides from trehalose and sucrose. AglB formed oligosaccharides from sucrose, maltose, and melizitose.

Kv1.1 channels of dorsal root ganglion neurons are inhibited by n-butyl-p-aminobenzoate, a promising anesthetic for the treatment of chronic pain.

In this study, we investigated the effects of the local anesthetic n-butyl-p-aminobenzoate (BAB) on the delayed rectifier potassium current of cultured dorsal root ganglion (DRG) neurons using the patch-clamp technique. The majority of the K(+) current of small DRG neurons rapidly activates and slowly inactivates at depolarized voltages. BAB inhibited the whole-cell K(+) current of these neurons with an IC(50) value of 228 microM. Dendrotoxin K (DTX(K)), a specific inhibitor of Kv1.1, reduced the DRG K(+) current at +20 mV by 34%, consistent with an important contribution of channels incorporating the Kv1.1 subunit to the delayed rectifier current. To further investigate the mechanism of BAB inhibition, we examined its effect on Kv1.1 channels heterologously expressed in mammalian tsA201 cells. BAB inhibits the Kv1.1 channels with an IC(50) value of 238 microM, similar to what was observed for the native DRG current. BAB accelerates the opening and closing of Kv1.1, but does not alter the midpoint of steady-state activation. BAB seems to inhibit Kv1.1 by stabilizing closed conformations of the channel. Coexpression with the Kv beta 1 subunit induces rapid inactivation and reduces the BAB sensitivity of Kv1.1. Comparison of the heterologously expressed Kv1.1 and native DRG currents indicates that the Kv beta 1 subunit does not modulate the gating of the DTX(K)-sensitive Kv1.1 channels of DRG neurons. Inhibition of the delayed rectifier current of these neurons may contribute to the long-duration anesthesia attained during the epidural administration of BAB.

Adamantinoma-like Ewing's sarcoma and Ewing's-like adamantinoma. The t(11; 22), t(21; 22) status.

Adamantinoma of the long bones and Ewing's sarcoma are two malignant tumours between which, at first sight, there seems to be no morphological and clinical relationship. Both tumours, however, are known to express cytokeratins. Adamantinoma is a tumour of true epithelial nature, predominantly expressing cytokeratins 14 and 19. Ewing's sarcoma, believed to be from neuroectodermal origin, like other mesenchymal tumours, can aberrantly express cytokeratin 8 and 18. In the literature there are some reports of tumours showing clinical and/or morphological overlap between adamantinoma and Ewing's sarcoma, suggesting a possible relationship. These studies are mostly based on the epithelioid configuration of these lesions and their cytokeratin expression on immunohistochemistry. This raises the question of whether there is occasionally a morphological similarity between adamantinoma and Ewing's sarcoma, or whether there is a common genetic background. The Ewing's sarcoma/primitive peripheral neuroectodermal tumour (PNET) family is characterized in 90-95% of cases by a t(11; 22) and in 5-10% of cases by t(21; 22). In the few reports in the literature on cytogenetic investigations on adamantinoma, these translocations were never found using classical karyotyping. This study investigated the putative presence of t(11; 22) and t(21; 22) in 14 cases of adamantinoma by RT-PCR. These translocations were not found in any of these cases. The results support the view that these tumours are genetically and clinically distinct, but may eventually show overlapping morphological and immunohistochemical features.

Relative abundance and inhibitory distribution of protease inhibitors in potato juice from cv. Elkana.

Protease inhibitors from potato juice of cv. Elkana were purified and quantified. The protease inhibitors represent ca. 50% of the total soluble proteins in potato juice. The protease inhibitors were classified into seven different families: potato inhibitor I (PI-1), potato inhibitor II (PI-2), potato cysteine protease inhibitor (PCPI), potato aspartate protease inhibitor (PAPI), potato Kunitz-type protease inhibitor (PKPI), potato carboxypeptidase inhibitor (PCI), and "other serine protease inhibitors". The most abundant families were the PI-2 and PCPI families, representing 22 and 12% of all proteins in potato juice, respectively. Potato protease inhibitors show a broad spectrum of enzyme inhibition. All the families (except PCI) inhibited trypsin and/or chymotrypsin. PI-2 isoforms exhibit 82 and 50% of the total trypsin and chymotrypsin inhibiting activity, respectively. A strong variation within the latter activities was shown within one family and between protease inhibitor families.

A new family of rhamnogalacturonan lyases contains an enzyme that binds to cellulose.

Pseudomonas cellulosa is an aerobic bacterium that synthesizes an extensive array of modular cellulases and hemicellulases, which have a modular architecture consisting of catalytic domains and distinct non-catalytic carbohydrate-binding modules (CBMs). To investigate whether the main-chain-cleaving pectinases from this bacterium also have a modular structure, a library of P. cellulosa genomic DNA, constructed in lambdaZAPII, was screened for pectinase-encoding sequences. A recombinant phage that attacked arabinan, galactan and rhamnogalacturonan was isolated. The encoded enzyme, designated Rgl11A, had a modular structure comprising an N-terminal domain that exhibited homology to Bacillus and Streptomyces proteins of unknown function, a middle domain that exhibited sequence identity to fibronectin-3 domains, and a C-terminal domain that was homologous to family 2a CBMs. Expression of the three modules of the Pseudomonas protein in Escherichia coli showed that its C-terminal module was a functional cellulose-binding domain, and the N-terminal module consisted of a catalytic domain that hydrolysed rhamnogalacturonan-containing substrates. The activity of Rgl11A against apple- and potato-derived rhamnogalacturonan substrates indicated that the enzyme had a strong preference for rhamnogalacturonans that contained galactose side chains, and which were not esterified. The enzyme had an absolute requirement for calcium, a high optimum pH, and catalysis was associated with an increase in absorbance at 235 nm, indicating that glycosidic bond cleavage was mediated via a beta-elimination mechanism. These data indicate that Rgl11A is a rhamnogalacturonan lyase and, together with the homologous Bacillus and Streptomyces proteins, comprise a new family of polysaccharide lyases. The presence of a family 2a CBM in Rgl11A, and in a P. cellulosa pectate lyase described in the accompanying paper [Brown, Mallen, Charnock, Davies and Black (2001) Biochem. J. 355, 155-165] suggests that the capacity to bind cellulose plays an important role in the activity of main-chain-cleaving Pseudomonas pectinases, in addition to cellulases and hemicellulases.

Chondrosarcoma is not characterized by detectable telomerase activity.

Reactivation of telomerase, an enzyme which elongates human telomeres, is associated with cell immortilization. In approximately 90% of malignant tumours telomerase activity can be demonstrated, whereas in benign tumours it is mostly absent. Chondrosarcomas are relatively rare malignant cartilaginous neoplasms. A small number of chondrosarcomas located centrally in bone arise secondarily to an enchondroma, while the majority of chondrosarcomas developing from the surface arise within the cartilage cap of an osteochondroma. The histological distinction between a benign lesion and low-grade chondrosarcoma is generally considered difficult. To investigate whether the progression towards chondrosarcoma is characterized by reactivation of telomerase activity, this study determined telomerase activity in ten enchondromas, five osteochondromas, and 37 chondrosarcomas using the TRAP assay. In all tumour samples except one, telomerase activity was absent. By adding tumour lysates to the positive control, an increasing inhibition of telomerase activity was found with an increasing chondroid matrix, suggesting that it may contain inhibitory factors. Inhibition due to endogenous RNAse or Taq-polymerase inhibitors was excluded. The lack of detectable telomerase activity in the high-grade component of a dedifferentiated chondrosarcoma without matrix favours the possibility that telomerase is truly absent. Either its true absence or inhibitory effects disabling telomerase detection exclude the telomerase TRAP assay as a diagnostic tool in the differential diagnosis of benign and low-grade malignant cartilaginous tumours.