A locus on the long arm of chromosome 1 as a possible cause of essential hypertension

None of the genes responsible for essential hypertension has been identified. Recent work in genetically hypertensive rats has shown linkage of blood pressure with alleles of the renin gene. Since the renin gene is a member of a conserved synteny group that in humans spans chromosome 1q21.3-32.3 and includes the gene for antithrombin III (AT3), we used linkage studies to examine the relationship between alleles of AT3 and hypertension in a family having 10 affected members. From the lod score obtained at a recombination fraction of zero the odds for linkage of AT3 and hypertension in this family were calculated as 6:1 in favour of linkage. This result provides grounds for further examination of the possible role of the 1q23 locus in the aetiology of essential hypertension.

Immunosuppressive properties of mesenchymal stromal cell cultures derived from the limbus of human and rabbit corneas

Background aims Mesenchymal stromal cells (MSCs) cultivated from the corneal limbus (L-MSCs) provide a potential source of cells for corneal repair. In the present study, we investigated the immunosuppressive properties of human L-MSCs and putative rabbit L-MSCs to develop an allogeneic therapy and animal model of L-MSC transplantation. Methods MSC-like cultures were established from the limbal stroma of human and rabbit (New Zealand white) corneas using either serum-supplemented medium or a commercial serum-free MSC medium (MesenCult-XF Culture Kit; Stem Cell Technologies, Melbourne, Australia). L-MSC phenotype was examined by flow cytometry. The immunosuppressive properties of L-MSC cultures were assessed using mixed leukocyte reactions. L-MSC cultures were also tested for their ability to support colony formation by primary limbal epithelial (LE) cells. Results Human L-MSC cultures were typically CD34−, CD45− and HLA-DR− and CD73+, CD90+, CD105+ and HLA-ABC+. High levels (>80%) of CD146 expression were observed for L-MSC cultures grown in serum-supplemented medium but not cultures grown in MesenCult-XF (approximately 1%). Rabbit L-MSCs were approximately 95% positive for major histocompatibility complex class I and expressed lower levels of major histocompatibility complex class II (approximately 10%), CD45 (approximately 20%), CD105 (approximately 60%) and CD90 (<10%). Human L-MSCs and rabbit L-MSCs suppressed human T-cell proliferation by up to 75%. Conversely, L-MSCs from either species stimulated a 2-fold to 3-fold increase in LE cell colony formation. Conclusions L-MSCs display immunosuppressive qualities in addition to their established non-immunogenic profile and stimulate LE cell growth in vitro across species boundaries. These results support the potential use of allogeneic L-MSCs in the treatment of corneal disorders and suggest that the rabbit would provide a useful pre-clinical model.

Low temperature pretreatment of sugarcane bagasse at atmospheric pressure using mixtures of ethylene carbonate and ethylene glycol

A low temperature lignocellulose pretreatment process was developed using acid-catalysed mixtures of alkylene carbonate and alkylene glycol. Pretreatment of sugarcane bagasse with mixtures of ethylene carbonate (EC) and ethylene glycol (EG) was more effective than that with mixtures of propylene carbonate (PC) and propylene glycol (PG). These mixtures were more effective than the individual components in making bagasse cellulose more amenable to cellulase digestion. Glucan digestibilities of ≥87% could be achieved with a wide range of EC to EG ratios from 9:1 to 1:1 (w/w). Pretreatment of bagasse by the EC/EG mixture with a ratio of 4:1 in the presence of 1.2% H2SO4 at 90 °C for 30 min led to the highest glucan enzymatic digestibility of 93%. The high glucan digestibilities obtained under these acidic conditions were due to (a) the ability of alkylene carbonate to cause significant biomass size reduction, (b) the ability of alkylene glycol to cause biomass defibrillation, (c) the ability of alkylene carbonate and alkylene glycol to remove xylan and lignin, and (d) the magnified above attributes in the mixtures of alkylene carbonate and alkylene glycol.

IS-RT-PCR assay detection of MT-MMP in a human breast cancer cell line

The in situ-reverse transcription-polymerase chain reaction (IS-RT-PCR) is a method that allows the direct localisation of gene expression. The method utilises the dual buffer mediated activity of the enzyme rTth DNA polymerase enabling both reverse transcription and DNA amplification. Labelled nucleoside triphosphates allow the site of expression to be labelled, rather than the PCR primers themselves, giving a more accurate localisation of transcript expression and decreased background than standard in situ hybridisation (ISH) assays. The MDA-MB-231 human breast carcinoma (HBC) cell line was assayed via the IS-RT-PCR technique, using primers encoding MT-MMP (membrane-type matrix metalloproteinase) and human β-actin. Our results clearly indicate baseline expression of MT-MMP in the relatively invasive MDA-MB-231 cell line at a signal intensity similar to the housekeeping gene β-actin, and results following induction with Concanavalin A (Con A) are consistent with our previous results obtained via Northern blotting.

Frequency in hypertensives of alleles for a RFLP associated with the renin gene

The genetic basis of primary hypertension is not known. Renin is important in blood pressure and volume control and a HindIII restriction fragment length polymorphism (RFLP) is present within the human renin gene locus. To examine whether there is a relationship between this RFLP and primary hypertension, DNA and renin analyses were performed on leukocytes and plasma from hypertensive and normotensive individuals. In hypertensives the frequencies of alleles for the HindIII RFLP were found to be 0.55 and 0.45, compared with 0.60 and 0.40 in the total population of 231 subjects examined, a difference that was not statistically significant. There also appeared to be no significant difference in renin activity in plasma for hypertensive patients of each genotype, nor in their pre- or post-treatment blood pressures. We thus conclude that, within the limits of the present study, the suspected genetic abnormalities associated with primary hypertension in man do not appear to be related to a HindIII RFLP in the renin gene.

Scanning the genome for essential hypertension loci

1. Essential hypertension occurs in people with an underlying genetic predisposition who subject themselves to adverse environmental influences. The number of genes involved is unknown, as is the extent to which each contributes to final blood pressure and the severity of the disease. 2. In the past, studies of potential candidate genes have been performed by association (case-control) analysis of unrelated individuals or linkage (pedigree or sibpair) analysis of families. These studies have resulted in several positive findings but, as one may expect, also an enormous number of negative results. 3. In order to uncover the major genetic loci for essential hypertension, it is proposed that scanning the genome systematically in 100- 200 affected sibships should prove successful. 4. This involves genotyping sets of hypertensive sibships to determine their complement of several hundred microsatellite polymorphisms. Those that are highly informative, by having a high heterozygosity, are most suitable. Also, the markers need to be spaced sufficiently evenly across the genome so as to ensure adequate coverage. 5. Tests are performed to determine increased segregation of alleles of each marker with hypertension. The analytical tools involve specialized statistical programs that can detect such differences. Non- parametric multipoint analysis is an appropriate approach. 6. In this way, loci for essential hypertension are beginning to emerge.

The influence of cellular source on periodontal regeneration using calcium phosphate coated polycaprolactone scaffold supported cell sheets

Cell-based therapy is considered a promising approach to achieving predictable periodontal regeneration. In this study, the regenerative potential of cell sheets derived from different parts of the periodontium (gingival connective tissue, alveolar bone and periodontal ligament) were investigated in an athymic rat periodontal defect model. Periodontal ligament (PDLC), alveolar bone (ABC) and gingival margin-derived cells (GMC) were obtained from human donors. The osteogenic potential of the primary cultures was demonstrated in vitro. Cell sheets supported by a calcium phosphate coated melt electrospun polycaprolactone (CaP-PCL) scaffold were transplanted to denuded root surfaces in surgically created periodontal defects, and allowed to heal for 1 and 4 weeks. The CaP-PCL scaffold alone was able to promote alveolar bone formation within the defect after 4 weeks. The addition of ABC and PDLC sheets resulted in significant periodontal attachment formation. The GMC sheets did not promote periodontal regeneration on the root surface and inhibited bone formation within the CaP-PCL scaffold. In conclusion, the combination of either PDLC or ABC sheets with a CaP-PCL scaffold could promote periodontal regeneration, but ABC sheets were not as effective as PDLC sheets in promoting new attachment formation.

Increasing electrospun scaffold pore size with tailored collectors for improved cell penetration

This study investigates the use of patterned collectors to increase the pore size of electrospun scaffolds for enhanced cell infiltration. The morphology of the patterned scaffolds was investigated by scanning electron microscopy, which showed that the collector pattern was accurately mimicked by the electrospun fibres. We observed an enlargement in the pore size and in the pore size distribution compared with conventional electrospinning. Mechanical testing revealed that the mechanical properties could be tailored, to some extent, according to the patterning and that the patterned scaffolds were softer than standard electrospun scaffolds. When NIH 3T3 fibroblasts were seeded onto patterned collectors improved cell infiltration was observed. Cells were able to penetrate up to 250 μm into the scaffolds, compared with 30 μm for the standard scaffolds. This increase in the depth of infiltration occurred as early as 24 h post-seeding and remained constant over 7 days.

An androgen receptor mutation in the MDA-MB-453 cell line model of molecular apocrine breast cancer compromises receptor activity

Recent evidence indicates that the estrogen receptor-a-negative, androgen receptor (AR)- positive molecular apocrine subtype of breast cancer is driven by AR signaling. The MDA-MB-453 cell line is the prototypical model of this breast cancer subtype; its proliferation is stimulated by androgens such as 5a-dihydrotestosterone (DHT) but inhibited by the progestin medroxyprogesterone acetate (MPA) via AR-mediated mechanisms. We report here that the AR gene in MDAMB- 453 cells contains a G-T transversion in exon 7, resulting in a receptor variant with a glutamine to histidine substitution at amino acid 865 (Q865H) in the ligand binding domain. Compared with wild-type AR, the Q865H variant exhibited reduced sensitivity to DHT and MPA in transactivation assays in MDA-MB-453 and PC-3 cells but did not respond to non-androgenic ligands or receptor antagonists. Ligand binding, molecular modeling, mammalian two-hybrid and immunoblot assays revealed effects of the Q865H mutation on ligand dissociation, AR intramolecular interactions, and receptor stability. Microarray expression profiling demonstrated that DHT and MPA regulate distinct transcriptional programs in MDA-MB-453 cells. Gene Set Enrichment Analysis revealed that DHT- but not MPA-regulated genes were associated with estrogen-responsive transcriptomes from MCF-7 cells and the Wnt signaling pathway. These findings suggest that the divergent proliferative responses of MDA-MB-453 cells to DHT and MPA result from the different genetic programs elicited by these two ligands through the AR-Q865H variant. This work highlights the necessity to characterize additional models of molecular apocrine breast cancer to determine the precise role of AR signaling in this breast cancer subtype. Endocrine-Related Cancer (2012) 19 599–613

A parametric analysis technique for design of fuel cell and hybrid-electric vehicles

This paper presents a new simplified parametric analysis technique for the design of fuel cell and hybrid-electric vehicles. The technique utilizes a comprehensive set of ∼30 parameters to fully characterize the vehicle platform, powertrain components, vehicle performance requirements and driving conditions. It is best applied to the sizing of powertrain components and prediction of energy consumption in a vehicle. This new parametric technique makes a good complement to existing vehicle simulation software packages and therefore represents a potentially valuable tool for the hybrid vehicle designer.

Epstein-Barr virus-positive diffuse large B-cell lymphoma of the elderly expresses EBNA3A with conserved CD8+ T-cell epitopes

Post-transplantation lymphoproliferative disorders (PTLD) arise in the immunosuppressed and are frequently Epstein-Barr virus (EBV) associated. The most common PTLD histological sub-type is diffuse large B-cell lymphoma (EBV+DLBCL-PTLD). Restoration of EBV-specific T-cell immunity can induce EBV+DLBCL-PTLD regression. The most frequent B-cell lymphoma in the immunocompetent is also DLBCL. ‘EBV-positive DLBCL of the elderly’ (EBV+DLBCL) is a rare but well-recognized DLBCL entity that occurs in the overtly immunocompetent, that has an adverse outcome relative to EBV-negative DLBCL. Unlike PTLD (which is classified as viral latency III), literature suggests EBV+DLBCL is typically latency II, i.e. expression is limited to the immuno-subdominant EBNA1, LMP1 and LMP2 EBV-proteins. If correct, this would be a major impediment for T-cell immunotherapeutic strategies. Unexpectedly we observed EBV+DLBCL-PTLD and EBV+DLBCL both shared features consistent with type III EBV-latency, including expression of the immuno-dominant EBNA3A protein. Extensive analysis showed frequent polymorphisms in EBNA1 and LMP1 functionally defined CD8+ T-cell epitope encoding regions, whereas EBNA3A polymorphisms were very rare making this an attractive immunotherapy target. As with EBV+DLBCL-PTLD, the antigen presenting machinery within lymphomatous nodes was intact. EBV+DLBCL express EBNA3A suggesting it is amenable to immunotherapeutic strategies.

Mathematical models of bubble evolution in a Hele-Shaw Cell

This thesis concerns the mathematical model of moving fluid interfaces in a Hele-Shaw cell: an experimental device in which fluid flow is studied by sandwiching the fluid between two closely separated plates. Analytic and numerical methods are developed to gain new insights into interfacial stability and bubble evolution, and the influence of different boundary effects is examined. In particular, the properties of the velocity-dependent kinetic undercooling boundary condition are analysed, with regard to the selection of only discrete possible shapes of travelling fingers of fluid, the formation of corners on the interface, and the interaction of kinetic undercooling with the better known effect of surface tension. Explicit solutions to the problem of an expanding or contracting ring of fluid are also developed.

The role of EBV-encoded microRNAs in B cell differentiation

Epstein Barr virus (EBV) is a common γ-herpes virus, infecting approximately 90% of the world‟s population. It is also one of the first known viruses known to be oncogenic, and is associated with a number of tumour types, primarily lymphomas. MicroRNAs (miRNAs) are post-transcriptional regulators of gene expression and many human miRNAs have been associated with the development of malignancies including cancer. EBV was the first human virus identified to express miRNAs and encodes more than 40 miRNAs within its genome. Yet, an understanding of the targets of EBV-miRNAs, and thereby the function of them in pathogenesis remains sadly limited. This study identifies a potential novel target of EBV-miRNAs, MECP2 and characterises the miRNA:mRNA interactions between two previously identified novel targets; Bim and EBF1. In particular, this study focuses upon the interaction between EBF1 and the EBV-miRNA BART11-5p, demonstrating a 151bp region of the EBF1 3‟UTR that is capable of mediating the silencing of luciferase expression by BART11-5p but is not capable of silencing a full length EBF1-3‟UTR luciferase construct. This study provides evidence that EBF1 may be a target of one or more EBV-miRNAs.