BslA is a self-assembling bacterial hydrophobin that coats the Bacillus subtilis biofilm

Biofilms represent the predominant mode of microbial growth in the natural environment. Bacillus subtilis is a ubiquitous Gram-positive soil bacterium that functions as an effective plant growth-promoting agent. The biofilm matrix is composed of an exopolysaccharide and an amyloid fiber-forming protein, TasA, and assembles with the aid of a small secreted protein, BslA. Here we show that natively synthesized and secreted BslA forms surface layers around the biofilm. Biophysical analysis demonstrates that BslA can self-assemble at interfaces, forming an elastic film. Molecular function is revealed from analysis of the crystal structure of BslA, which consists of an Ig-type fold with the addition of an unusual, extremely hydrophobic “cap” region. A combination of in vivo biofilm formation and in vitro biophysical analysis demonstrates that the central hydrophobic residues of the cap are essential to allow a hydrophobic, nonwetting biofilm to form as they control the surface activity of the BslA protein. The hydrophobic cap exhibits physiochemical properties remarkably similar to the hydrophobic surface found in fungal hydrophobins; thus, BslA is a structurally defined bacterial hydrophobin. We suggest that biofilms formed by other species of bacteria may have evolved similar mechanisms to provide protection to the resident bacterial community.     

In situ mantle cell lymphoma: clinical implications of an incidental finding with indolent clinical behavior

Background

Cyclin D1-positive B cells are occasionally found in the mantle zones of reactive lymphoid follicles, a condition that has been called “in situ mantle cell lymphoma”. The clinical significance of this lesion remains uncertain.

Design and Methods

The clinical and pathological characteristics, including SOX11 expression, of 23 cases initially diagnosed as in situ mantle cell lymphoma were studied.

Results

Seventeen of the 23 cases fulfilled the criteria for in situ mantle cell lymphoma. In most cases, the lesions were incidental findings in reactive lymph nodes. The t(11;14) was detected in all eight cases examined. SOX11 was positive in seven of 16 cases (44%). Five cases were associated with other small B-cell lymphomas. In two cases, both SOX11-positive, the in situ mantle cell lymphoma lesions were discovered after the diagnosis of overt lymphoma; one 4 years earlier, and one 3 years later. Twelve of the remaining 15 patients had a follow-up of at least 1 year (median 2 years; range, 1–19.5), of whom 11 showed no evidence of progression, including seven who were not treated. Only one of 12 patients with an in situ mantle cell lymphoma lesion and no diagnosis of mantle cell lymphoma at the time developed an overt lymphoma, 4 years later; this case was also SOX11-positive. The six remaining cases were diagnosed as mantle cell lymphoma with a mantle zone pattern. Five were SOX11-positive and four of them were associated with lymphoma without a mantle zone pattern.

Conclusions

In situ mantle cell lymphoma lesions are usually an incidental finding with a very indolent behavior. These cases must be distinguished from mantle cell lymphoma with a mantle zone pattern and overt mantle cell lymphoma because they may not require therapeutic intervention.     

Plasma Enhanced Chemical Vapour Deposition of Horizontally Aligned Carbon Nanotubes

A plasma-enhanced chemical vapour deposition reactor has been developed to synthesis horizontally aligned carbon nanotubes. The width of the aligning sheath was modelled based on a collisionless, quasi-neutral, Child’s law ion sheath where these estimates were empirically validated by direct Langmuir probe measurements, thereby confirming the proposed reactors ability to extend the existing sheath fields by up to 7 mm. A 7 mbar growth atmosphere combined with a 25 W plasma permitted the concurrent growth and alignment of carbon nanotubes with electric fields of the order of 0.04 V μm⁻¹ with linear packing densities of up to ~5 × 10⁴ cm⁻¹. These results open up the potential for multi-directional in situ alignment of carbon nanotubes providing one viable route to the fabrication of many novel optoelectronic devices.     

Quantum speciation in Aegilops: molecular cytogenetic evidence from rDNA cluster variability in natural populations

Data are presented on quantum speciation in the Sitopsis section of the genus Aegilops (Poaceae, Monocotyledones). Two small, peripheral, isolated, wild populations of annual cross-pollinated Ae. speltoides and annual self-pollinated Ae. sharonensis are located 30 m apart on different soil types. Despite the close proximity of the two species and their close relatedness, no mixed groups are known. Comparative molecular cytogenetic analysis based on the intrapopulation variability of rRNA-encoding DNA (rDNA) chromosomal patterns of individual Ae. speltoides geno-types revealed an ongoing dynamic process of permanent chromosomal rearrangements. Chromosomal mutations can arise de novo and can be eliminated. Analysis of the progeny of the investigated genotypes testifies that inheritance of de novo rDNA sites happens frequently. Heterologous recombination and/or transposable elements-mediated rDNA transfer seem to be the mechanisms for observed chromosomal repatterning. Consequently, several modified genomic forms, intermediate between Ae. speltoides and Ae. sharonensis, permanently arise in the studied wild population of Ae. speltoides, which make it possible to recognize Ae. sharonensis as a derivative species of Ae. speltoides, as well as to propose rapidness and canalization of quantum speciation in Sitopsis species.     

Metabolizing enzyme toxicology assay chip (MetaChip) for high-throughput microscale toxicity analyses

The clinical progression of new chemical entities to pharmaceuticals remains hindered by the relatively slow pace of technology development in toxicology and clinical safety evaluation, particularly in vitro approaches, that can be used in the preclinical and early clinical phases of drug development. To alleviate this bottle-neck, we have developed a metabolizing enzyme toxicology assay chip (MetaChip) that combines high-throughput P450 catalysis with cell-based screening on a microscale platform. The MetaChip concept is demonstrated by using sol-gel encapsulated P450s to activate the prodrug cyclophosphamide, which is the major constituent of the anticancer drug Cytoxan, as well as other compounds that are activated by P450 metabolism. The MetaChip provides a high-throughput microscale alternative to currently used in vitro methods for human metabolism and toxicology screening based on liver slices, cultured human hepatocytes, purified microsomal preparations, or isolated and purified P450s. This technology creates opportunities for rapid and inexpensive assessment of ADME/Tox (absorption, distribution, metabolism, excretion/toxicology) at very early phases of drug development, thereby enabling unsuitable candidates to be eliminated from consideration much earlier in the drug discovery process.     

Condyloma acuminatum of the anal canal,treated with endoscopic submucosal dissection

Condyloma acuminatum(CA) is a common sexually transmitted disease caused by human papilloma virus infection. Not all individuals develop persistent, progressive disease, but careful follow up is required with moderate-to-severe dysplasia to prevent progression to malignancy. Standard therapies include surgical treatments(trans-anal resection and transanal endoscopic microsurgery) and immunotherapeutic and topical methods(topical imiquimod); however, local recurrence remains a considerable problem. Here, we report a case with superficial CA of the anal canal, treated with endoscopic submucosal dissection(ESD). A 28-year-old man presented with a chief complaint of hematochezia. Digital exam did not detect a tumor. Screening colonoscopy revealed 10-mm long, whitish condyles extending from the anal canal to the lower rectum. The lesion covered almost the whole circumference, and only a small amount of normal mucosa remained. Magnifying endoscopy with narrow band imaging showed brownish hairpin-shaped, coiled capillaries. Although histopathological diagnosis by biopsy revealed CA, accurate histological differentiation between CA, papilloma, and squamous cell carcinoma can be difficult with a small specimen. Therefore, weperformed diagnostic ESD, which provides a complete specimen for precise histopathological evaluation. The pathological diagnosis was CA, with moderate dysplasia(anal intraepithelial neoplasia 2). There was no recurrence at 16 mo after the initial ESD. Compared to surgical treatment, endoscopic diagnosis and resection could be performed simultaneously and the tumor margin observed clearly with a magnifying chromocolonoscopy, resulting in less recurrence. These findings suggest that endoscopic resection may be an alternative method for CA that prevents recurrence.     

Using Converter Dust to Produce Low Cost Cementitious Composites by in situ Carbon Nanotube and Nanofiber Synthesis

Carbon nanotubes (CNTs) and nanofibers (CNFs) were synthesized on clinker and silica fume particles in order to create a low cost cementitious nanostructured material. The synthesis was carried out by an in situ chemical vapor deposition (CVD) process using converter dust, an industrial byproduct, as iron precursor. The use of these materials reduces the cost, with the objective of application in large-scale nanostructured cement production. The resulting products were analyzed by scanning electron microscopy (SEM), transmission electron microscopy (TEM) and thermogravimetric analysis (TGA) and were found to be polydisperse in size and to have defective microstructure. Some enhancement in the mechanical behavior of cement mortars was observed due to the addition of these nano-size materials. The contribution of these CNTs/CNFs to the mechanical strength of mortar specimens is similar to that of high quality CNTs incorporated in mortars by physical mixture.     

Three-dimensional cellular microarray for high-throughput toxicology assays

We have developed a miniaturized 3D cell-culture array (the Data Analysis Toxicology Assay Chip or DataChip) for high-throughput toxicity screening of drug candidates and their cytochrome P450-generated metabolites. The DataChip consists of human cells encapsulated in collagen or alginate gels (as small as 20 nl) arrayed on a functionalized glass slide for spatially addressable screening against multiple compounds. A single DataChip containing 1,080 individual cell cultures, used in conjunction with the complementary human P450-containing microarray (the Metabolizing Enzyme Toxicology Assay Chip or MetaChip), simultaneously provided IC(50) values for nine compounds and their metabolites from CYP1A2, CYP2D6, and CYP3A4 and a mixture of the three P450s designed to emulate the human liver. Similar responses were obtained with the DataChip and conventional 96-well plate assays, demonstrating that the near 2,000-fold miniaturization does not influence the cytotoxicity response. The DataChip may therefore enable toxicity analyses of drug candidates and their metabolites at throughputs compatible with the availability of compounds at early-stage drug discovery.     

Mechanical Properties of Titanium Nitride Nanocomposites Produced by Chemical Precursor Synthesis Followed by High-P,T Treatment

We investigated the high-P,T annealing and mechanical properties of nanocomposite materials with a highly nitrided bulk composition close to Ti₃N₄. Amorphous solids were precipitated from solution by ammonolysis of metal dialkylamide precursors followed by heating at 400–700 °C in flowing NH₃ to produce reddish-brown amorphous/nanocrystalline materials. The precursors were then densified at 2 GPa and 200–700 °C to form monolithic ceramics. There was no evidence for N₂ loss during the high-P,T treatment. Micro- and nanoindentation experiments indicate hardness values between 4–20 GPa for loads ranging between 0.005–3 N. Young''s modulus values were measured to lie in the range 200–650 GPa. Palmqvist cracks determined from microindentation experiments indicate fracture toughness values between 2–4 MPa·m^1/2 similar to Si₃N₄, SiC and Al₂O₃. Significant variations in the hardness may be associated with the distribution of amorphous/crystalline regions and the very fine grained nature (~3 nm grain sizes) of the crystalline component in these materials.     

Fatigue Failure Analyses on a Ti-45Al-8Nb-0.2W-0.2B-0.1Y Alloy at Different Temperatures

The fatigue micro crack initiation and propagation tests of a TiAl alloy with 8% Nb content were carried out by using scanning electron microscopy in situ technology at room temperature and at 750 °C. These results indicated that the fatigue micro crack initiation was mainly caused by the stress concentration at room temperature, but at an elevated temperature (750 °C) the multi-cracks were caused by the coupled factors of both lamellar microstructure and stress concentration. Therefore, fatigue micro crack initiation behavior is much more dependent on the lamellar structure at an elevated temperature. One of the reasons is that the elevated temperature degrades the interface strength between the lamellar of the TiAl alloy with 8% Nb content. Therefore, the small fatigue crack propagation behavior of the alloy exhibited a mixture damage model of interlamellar and translamellar at a micro scale. The crack growth path and fracture characteristics provided a proof of crack deflection, branching and/or bridging induced either by interlamellar or by translamellar failure mode.     

Expression of somatostatin mRNA in various differentiated types of gastric carcinoma

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The Microstructural Evolution and Mechanical Properties of Zr-Based Metallic Glass under Different Strain Rate Compressions

In this study, the high strain rate deformation behavior and the microstructure evolution of Zr-Cu-Al-Ni metallic glasses under various strain rates were investigated. The influence of strain and strain rate on the mechanical properties and fracture behavior, as well as microstructural properties was also investigated. Before mechanical testing, the structure and thermal stability of the Zr-Cu-Al-Ni metallic glasses were studied with X-ray diffraction (XRD) and differential scanning calorimeter. The mechanical property experiments and microstructural observations of Zr-Cu-Al-Ni metallic glasses under different strain rates ranging from 10⁻³ to 5.1 × 10³ s⁻¹ and at temperatures of 25 °C were investigated using compressive split-Hopkinson bar (SHPB) and an MTS tester. An in situ transmission electron microscope (TEM) nanoindenter was used to carry out compression tests and investigate the deformation behavior arising at nanopillars of the Zr-based metallic glass. The formation and interaction of shear band during the plastic deformation were investigated. Moreover, it was clearly apparent that the mechanical strength and ductility could be enhanced by impeding the penetration of shear bands with reinforced particles.     

Human papillomavirus-related squamous cell carcinoma of the anal canal with papillary features

Human papillomavirus(HPV)related squamous cell carcinoma(SCC)involving the anal canal is a well-known carcinoma associated with high-risk types of HPV.HPVrelated SCC with papillary morphology(papillary SCC)has been described in the oropharynx.We describe,for the first time,a case of anal HPV-related squamous carcinoma with papillary morphology.The tumor arose from the anal mucosa.The biopsies revealed a superficially invasive SCCwith prominent papillary features and associated in situ carcinoma.The tumor cells were positive for p16 and were also positive for high-risk types of HPV using chromogenic in situ hybridization.The findings are consistent with a HPV-related SCC of the anal canal with papillary features.This tumor shows histologic features similar to a papillary HPV-related SCC of the oropharynx.Additional studies are needed to characterize these lesions.     

KAI1 gene is differently expressed in papillary and pancreatic cancer: influence on metastasis

AIM:To compare KAI1 in cancer of papilla of Vater and pancreas to evaluate whether there are differences in biologic behavior which might account for prognosis.METHODS:We compared the expression in 24 papillay and 29 pancreatic cancers using Northern blot analysis, immunochemical assay and in situ hybridization, and investigated whether early diagnosis or molecular differences predict the outcome in these tumor entities.RESULTS:By Northern blot analysis there is no statistical difference of KAI1 levels in normal and cancerous papilla. No association between KAI1 mRNA expression and tumor stage or tumor differentiation was found in the tumors. By immunohistochemical assay, KAI1 staining in cytoplasm of papillary cancer cells was similar to that of normal papillary cells. By in situ hybridization, the results of KAI1 mRNA expression in normal and cancerous papilla were similar to those with immunohistochemical assay. The normal and cancerous pancreas tissues were also analyzed by the methods used in papillary samples.CONCLUSION:Although the biologic roles of KAI1 have not been clarified, our results suggest that KAI1 may restrict the progression of malignant papillary cancer, but its expression might not have any effect on the characteristics of papillary tumor, whereas by the analysis of KAI1 gene, its reduced expression is closely related to the progression and metastases of pancreatic cancer.     

Human epidermal growth factor receptor-2 in oesophageal cancers: An observational study

AIM: To determine the incidence of human epidermal growth factor receptor 2 (HER2) over expression in oesophageal cancers.METHODS: A retrospective study, of one hundred consecutive cases of endoscopic histological samples of oesophageal cancers from a single British cancer network were included. Cancer cases were diagnosed between April 2007 and June 2010. HER2 over expression was assessed using immunohistochemistry, those that scored “0” and “+1” were considered “negative” for HER2; those that scored “+3” were considered “Positive”. Cases that were scored “+2” on immunohistochemistry further went on to have HER2 gene analysis using the Ventana HER brightfield dual-colour in situ hybridisations (HER B DISH) assay and either came back to be positive or negative for HER2 over expression. Overall survival was measured from date of histological diagnosis until date of death. 93% of the cases were followed up till five years or death, and all were followed up till two years. Cases of gastro-oesophageal junctional tumours were excluded.RESULTS: The median age of our sample was 66 years (range: 38-91 years). Eighty one were male and 19 female. Ninety-one of the cases were adenocarcinoma of the oesophagus and the rest were cases of squamous cell carcinoma. The anatomical distribution of the tumours was; upper oesophagus 2, middle oesophagus 11, and 87 were in the lower oesophagus. Operative resection was completed in 15 cases; seven cases had attempted surgical resections, i.e., open and close, 33 patients received definitive chemo-radiation and 52 had palliative treatment. Twenty-five of the cancers showed evidence of HER2 over expression, all were adenocarcinomas. Of the 25 cases that showed evidence of HER2 over expression, 21 (84%) were located in the lower third of the oesophagus. On staging, 24 out of the 25 HER2 positive cases were at stage 3 or more (13 at stage 3 and 11 at stage 4), For HER2 negative cases 37 were at stage 3 and 32 were staged as stage 4. Seventeen out of twenty five cases (68%) with HER2 over expression received palliative therapy, in comparison to thirty five out of seventy five (46.7%) in tumours not expressing HER2. No significant difference in overall survival was demonstrated between patients whose cancers showed evidence of HER2 over expression and those who did not; median overall survival for HER2 positive tumours was 15 mo (95%CI, 11-19 mo) compared to 13 mo (95%CI, 9-17 mo) for HER2 negative ones. Two years cumulative survival for cases with HER2 over expression was 33.7% compared to 31.6% in cases without HER2 over expression (P = 0.576). Only cancer’s stage significantly affected overall survival on both univariant and multivariable analysis (P = 0.034 and P = 0.009 respectively). None of the patients included in this study received Trastuzumab.CONCLUSION: Twenty-seven point five percent of oesophageal adenocarcinomas showed evidence of HER2 over expression. Routine testing for human HER2 in oesophageal adenocarcinomas can have significant implication on treatments offered to patients that may potentially affect their prognosis.     

In Vivo Osteogenic Differentiation of Human Embryoid Bodies in an Injectable in Situ-Forming Hydrogel

In this study, we examined the in vivo osteogenic differentiation of human embryoid bodies (hEBs) by using an injectable in situ-forming hydrogel. A solution containing MPEG-b-(polycaprolactone-ran-polylactide) (MCL) and hEBs was easily prepared at room temperature. The MCL solution with hEBs and osteogenic factors was injected into nude mice and developed into in situ-forming hydrogels at the injection sites; these hydrogels maintained their shape even after 12 weeks in vivo, thereby indicating that the in situ-forming MCL hydrogel was a suitable scaffold for hEBs. The in vivo osteogenic differentiation was observed only in the in situ gel-forming MCL hydrogel in the presence of hEBs and osteogenic factors. In conclusion, this preliminary study suggests that hEBs and osteogenic factors embedded in an in situ-forming MCL hydrogel may provide numerous benefits as a noninvasive alternative for allogeneic tissue engineering applications.     

Association of Fusobacterium nucleatum infection with colorectal cancer in Chinese patients

AIM: To investigate Fusobacterium nucleatum(F. nucleatum) abundance in colorectal cancer(CRC) tissues and its association with CRC invasiveness in Chinese patients.METHODS: The resected cancer and adjacent normal tissues(10 cm beyond cancer margins) from 101 consecutive patients with CRC were collected. Fluorescent quantitative polymerase chain reaction(FQ-PCR) was applied to detect F. nucleatum in CRC and normal tissues. The difference of F. nucleatum abundance between cancer and normal tissues and the relationship of F. nucleatum abundance with clinical variables were evaluated. Fluorescence in situ hybridization(FISH) analysis was performed on 22 CRC tissues with the highest F. nucleatum abundance by FQ-PCR testing to confirm FQ-PCR results.RESULTS: The median abundance of F. nucleatum in CRC tissues [0.242(0.178-0.276)] was significantly higher than that in normal controls [0.050(0.023-0.067)](P 0.001). F. nucleatum was over-represented in 88/101(87.1%) CRC samples. The abundance of F. nucleatum determined by 2~(-ΔCT) was significantly greater in tumor samples [0.242(0.178, 0.276)] than in normal controls [0.050(0.023, 0.067)](P 0.001). The frequency of patients with lymph node metastases was higher in the over-abundance group [52/88(59.1%)] than in the under-abundance group [0/13(0%)](P 0.005). No significant association of F. nucleatum with other clinico-pathological variables was observed(P 0.05). FISH analysis also found more F. nucleatum in CRC than in normal tissues(median number 6, 25~(th) 3, 75~(th) 10 vs 2, 25~(th) 1, 75~(th) 5)(P 0.01).CONCLUSION: F. nucleatum was enriched in CRC tissues and associated with CRC development and metastasis.     

Human liver stem/progenitor cells decrease serum bilirubin in hyperbilirubinemic Gunn rat

AIM:To test the ability of adult-derived human liver stem/progenitor cells(ADHLSC)from large scale cultures to conjugate bilirubin in vitro and in bilirubin conjugation deficient rat.METHODS:ADHLSC from large scale cultures were tested for their phenotype and for their capacity to conjugate bilirubin in vitro after hepatogenic differentiation.In vivo,Gunn rats[uridine diphosphate-glucuronosyltransferase 1A1(UGT1A1)deficient animal]were injected with ADHLSC and cryopreserved hepatocytes(positive control).Two,4,13 and 27 wk posttransplantation,transplanted Gunn rat bilirubin serum levels were determined by high performance liquid chromatography.Human cell engraftment of trans-planted cells was assessed 27 wk post-transplantation using immunohistochemistry and RTqPCR.RESULTS:Large scale culture conditions do not modified ADHLSC phenotype,ADHLSC were able to specifically conjugate bilirubin.ADHLSC were intraportally injected into Gunn rats and blood UCB was measured at different times post-transplantation,infused-Gunn rats exhibited a metabolic effect 3 mo post-transplantation and maintained over a 6 mo period.ADHLSC engraftment into Gunn rat’s liver was demonstrated by RTqPCR and immunohistochemistry against albumin and UGT1A1.CONCLUSION:ADHLSC from large scale cultures are efficient in conjugating bilirubin in vitro and in restoring a deficient metabolic function(reducing bilirubin level)in hyperbilirubinemic rats.