An investigation into the use of human papillomavirus type 16 virus-like particles as a delivery vector system for foreign proteins: N- and C-terminal fusion of GFP to the L1 and L2 capsid proteins

Development of vaccine strategies against human papillomavirus (HPV), which causes cervical cancer, is a priority. We investigated the use of virus-like particles (VLPs) of the most prevalent type, HPV-16, as carriers of foreign proteins. Green fluorescent protein (GFP) was fused to the N or C terminus of both L1 and L2, with L2 chimeras being co-expressed with native L1. Purified chimaeric VLPs were comparable in size (∼55 nm) to native HPV VLPs. Conformation-specific monoclonal antibodies (Mabs) bound to the VLPs, thereby indicating that they possibly retain their antigenicity. In addition, all of the VLPs encapsidated DNA in the range of 6-8 kb. © 2007 Springer-Verlag.

Reasons for substance use among people with mental disorders

Background: Comorbidity of mental disorders and substance use continues to be a major problem. To inform the development of more effective interventions for these co-existing disorders, this paper aimed to determine if there are clear variations in the reasons for tobacco, alcohol or cannabis use across people with different mental disorders. Methods: Data from five randomized controlled trials on co-existing disorders that measured reasons for tobacco, alcohol or cannabis use using the Drug Use Motives Questionnaire, Reasons for Smoking Questionnaire or via free response are reported and combined. Two studies involved participants with depression, two involved participants with a psychotic disorder and one involved participants with a range of mental disorders. A series of logistic regressions were conducted to examine differences in reasons for tobacco, alcohol or cannabis use and to compare these reasons between people with psychotic disorders or depression. Results: Participants had a mean age of 38 (SD=12) and just over half (60%) were male. Forty-six percent of participants had a psychotic disorder and 54% experienced depression. Data from 976 participants across the five studies were included in the analyses. Tobacco and alcohol were primarily used to cope, while cannabis was primarily used for pleasure. People with psychotic disorders were more likely than people with depression to use tobacco for coping, pleasure and illness motives. People with depression, in contrast, were more likely to use alcohol for these reasons and social reasons. Conclusions: It may be important to tailor interventions for co-existing mental disorders and substance use by substance type and type of mental disorder. For example, interventions might be improved by including alternative coping strategies to tobacco and/or alcohol use, by addressing the social role of alcohol and by helping people with mental disorders using cannabis to gain pleasure from their lives in other ways.

Cement-in-cement revision for selected Vancouver Type B1 femoral periprosthetic fractures : A biomechanical analysis

The aim of this study was to perform a biomechanical analysis of the cement-in-cement (c-in-c) technique for fixation of selected Vancouver Type B1 femoral periprosthetic fractures and to assess the degree of cement interposition at the fracture site. Six embalmed cadaveric femora were implanted with a cemented femoral stem. Vancouver Type B1 fractures were created by applying a combined axial and rotational load to failure. The femora were repaired using the c-in-c technique and reloaded to failure. The mean primary fracture torque was 117 Nm (SD 16.6, range 89–133). The mean revision fracture torque was 50 Nm (SD 16.6, range 29–74), which is above the torque previously observed for activities of daily living. Cement interposition at the fracture site was found to be minimal.

Perceived barriers to healthy lifestyle activities in midlife and older Australian women with type 2 diabetes

Purpose: Type 2 diabetes is a leading cause of morbidity and mortality in midlife and older Australian women with known modifiable risk factors for type 2 diabetes including smoking, nutrition, physical activity and obesity. In Australia little research has been done to investigate the perceived barriers to healthy lifestyle activities in midlife and older women with type 2 diabetes. The primary aim of this study was to explore the level and type of perceived barriers to health promotion activities. The secondary aim was to explore the relationship of perceived barriers to smoking behaviour, fruit and vegetable intake, physical activity, and body mass index. Methods: The study was a cross sectional survey of women, aged over 45 with type 2 diabetes, attending metropolitan community health clinics (N = 41). Data was collected from self-report questionnaire and analysed using descriptive and inferential statistics. Results: Women in the study had average total barriers scores similar to those reported in the literature for women with a range of physical disabilities and illnesses. The leading barriers for this group of women were: lack of interest, concern about safety, too tired, lack of money and feeling what they do does not help. There was no association between total barriers scores and body mass index, physical activity, fruit and vegetable intake or socio-demographic variables. Conclusion: This study contributes to understanding the perceptions of midlife and older women with type 2 diabetes about the level and type of barriers to healthy lifestyle activities that they experience. Evidence from this study can be applied to inform health promotion for lifestyle risk factor reduction in women with type 2 diabetes.

The AusD Study: A population-based study of the determinants of serum 25-hydroxyvitamin D concentration across a broad latitude range

Observational studies suggest that people with a high serum 25-hydroxyvitamin D (25(OH)D) concentration may have reduced risk of chronic diseases such as osteoporosis, multiple sclerosis, type 1 diabetes, cardiovascular disease, and some cancers. The AusD Study (A Quantitative Assessment of Solar UV Exposure for Vitamin D Synthesis in Australian Adults) was conducted to clarify the relationships between ultraviolet (UV) radiation exposure, dietary intake of vitamin D, and serum 25(OH)D concentration among Australian adults residing in Townsville (19.3°S), Brisbane (27.5°S), Canberra (35.3°S), and Hobart (42.8°S). Participants aged 18-75 years were recruited from the Australian Electoral Roll between 2009 and 2010. Measurements were made of height, weight, waist:hip ratio, skin, hair, and eye color, blood pressure, and grip strength. Participants completed a questionnaire on sun exposure and vitamin D intake, together with 10 days of personal UV dosimetry and an associated sun-exposure and physical-activity diary that was temporally linked to a blood test for measurement of 25(OH)D concentration. Ambient solar UV radiation was also monitored at all study sites. We collected comprehensive, high-quality data from 1,002 participants (459 males, 543 females) assessed simultaneously across a range of latitudes and through all seasons. Here we describe the scientific and methodological issues considered in designing the AusD Study.

Memory effect in the deposition of C20 fullerenes on a diamond surface

In this paper, the deposition of C-20 fullerenes on a diamond (001)-(2x1) surface and the fabrication of C-20 thin film at 100 K were investigated by a molecular dynamics (MD) simulation using the many-body Brenner bond order potential. First, we found that the collision dynamic of a single C-20 fullerene on a diamond surface was strongly dependent on its impact energy. Within the energy range 10-45 eV, the C-20 fullerene chemisorbed on the surface retained its free cage structure. This is consistent with the experimental observation, where it was called the memory effect in "C-20-type" films [P. Melion , Int. J. Mod. B 9, 339 (1995); P. Milani , Cluster Beam Synthesis of Nanostructured Materials (Springer, Berlin, 1999)]. Next, more than one hundred C-20 (10-25 eV) were deposited one after the other onto the surface. The initial growth stage of C-20 thin film was observed to be in the three-dimensional island mode. The randomly deposited C-20 fullerenes stacked on diamond surface and acted as building blocks forming a polymerlike structure. The assembled film was also highly porous due to cluster-cluster interaction. The bond angle distribution and the neighbor-atom-number distribution of the film presented a well-defined local order, which is of sp(3) hybridization character, the same as that of a free C-20 cage. These simulation results are again in good agreement with the experimental observation. Finally, the deposited C-20 film showed high stability even when the temperature was raised up to 1500 K.

Lattice-free models of cell invasion : discrete simulations and travelling waves

Invasion waves of cells play an important role in development, disease and repair. Standard discrete models of such processes typically involve simulating cell motility, cell proliferation and cell-to-cell crowding effects in a lattice-based framework. The continuum-limit description is often given by a reaction–diffusion equation that is related to the Fisher–Kolmogorov equation. One of the limitations of a standard lattice-based approach is that real cells move and proliferate in continuous space and are not restricted to a predefined lattice structure. We present a lattice-free model of cell motility and proliferation, with cell-to-cell crowding effects, and we use the model to replicate invasion wave-type behaviour. The continuum-limit description of the discrete model is a reaction–diffusion equation with a proliferation term that is different from lattice-based models. Comparing lattice based and lattice-free simulations indicates that both models lead to invasion fronts that are similar at the leading edge, where the cell density is low. Conversely, the two models make different predictions in the high density region of the domain, well behind the leading edge. We analyse the continuum-limit description of the lattice based and lattice-free models to show that both give rise to invasion wave type solutions that move with the same speed but have very different shapes. We explore the significance of these differences by calibrating the parameters in the standard Fisher–Kolmogorov equation using data from the lattice-free model. We conclude that estimating parameters using this kind of standard procedure can produce misleading results.

A retrospective cohort investigation of active range of motion within one week of open reduction and internal fixation of distal radius fractures

Distal radius fractures stabilized by open reduction internal fixation (ORIF) have become increasingly common. There is currently no consensus on the optimal time to commence range of motion (ROM) exercises post-ORIF. A retrospective cohort review was conducted over a five-year period to compare wrist and forearm range of motion outcomes and number of therapy sessions between patients who commenced active ROM exercises within the first seven days and from day eight onward following ORIF of distal radius fractures. One hundred and twenty-one patient cases were identified. Clinical data, active ROM at initial and discharge therapy assessments, fracture type, surgical approaches, and number of therapy sessions attended were recorded. One hundred and seven (88.4%) cases had complete datasets. The early active ROM group (n = 37) commenced ROM a mean (SD) of 4.27 (1.8) days post-ORIF. The comparator group (n = 70) commenced ROM exercises 24.3 (13.6) days post-ORIF. No significant differences were identified between groups in ROM at initial or discharge assessments, or therapy sessions attended. The results from this study indicate that patients who commenced active ROM exercises an average of 24 days after surgery achieved comparable ROM outcomes with similar number of therapy sessions to those who commenced ROM exercises within the first week.

Perceived barriers to healthy lifestyle activities in midlife and older Australian women with type 2 diabetes

Background Type 2 diabetes is a leading cause of morbidity and mortality in midlife and older Australian women with known modifiable risk factors for type 2 diabetes including smoking, nutrition, physical activity and obesity. In Australia little research has been done to investigate the perceived barriers to healthy lifestyle activities in midlife and older women with type 2 diabetes. Aims The primary aim of this study was to explore the level and type of perceived barriers to health promotion activities. The secondary aim was to explore the relationship of perceived barriers to smoking behaviour, fruit and vegetable intake, physical activity, and body mass index. Methods The study was a cross sectional survey of women, aged over 45 with type 2 diabetes, recruited from four metropolitan community health clinics (n = 41). Data were collected from self-report questionnaires and analysed using quantitative methods. Results Women in the study had average total barriers scores similar to those reported in the literature for women with a range of physical disabilities and illnesses. The leading barriers for this group of women were: lack of interest, concern about safety, too tired, lack of money and feeling what they do does not help. There was no association between total barriers scores and body mass index, physical activity, fruit and vegetable intake or socio-demographic variables. Conclusion This study contributes to understanding the perceptions of midlife and older women with type 2 diabetes about the level and type of barriers to healthy lifestyle activities that they experience. The participants reported a high level perceived barriers with a range of personal, social and environmental issues identified and described. This study suggests that health promotion education and interventions for risk factor reduction in women with type 2 diabetes may be enhanced by explicitly addressing perceived barriers to healthy lifestyle activities.

A novel protecting group methodology for syntheses using nitroxides

The methoxyamine group represents an ideal protecting group for the nitroxide moiety. It can be easily and selectively introduced in high yield (typically >90%) to a range of functionalised nitroxides using FeSO4.7H2O and H2O2 in DMSO. Its removal is readily achieved under mild conditions in high yield (70-90%) using mCPBA in a Cope-type elimination process.

Rapid quantification of free cholesterol in tears using direct insertion/electron ionization-Mass spectrometry

Purpose. To establish a simple and rapid analytical method, based on direct insertion/electron ionization-mass spectrometry (DI/EI-MS), for measuring free cholesterol in tears from humans and rabbits. Methods. A stable-isotope dilution protocol employing DI/EI-MS in selected ion monitoring mode was developed and validated. It was used to quantify the free cholesterol content in human and rabbit tear extracts. Tears were collected from adult humans (n = 15) and rabbits (n = 10) and lipids extracted. Results. Screening, full-scan (m/z 40-600) DI/EI-MS analysis of crude tear extracts showed that diagnostic ions located in the mass range m/z 350 to 400 were those derived from free cholesterol, with no contribution from cholesterol esters. DI/EI-MS data acquired using selected ion monitoring (SIM) were analyzed for the abundance ratios of diagnostic ions with their stable isotope-labeled analogues arising from the D6-cholesterol internal standard. Standard curves of good linearity were produced and an on-probe limit of detection of 3 ng (at 3:1 signal to noise) and limit of quantification of 8 ng (at 10:1 signal to noise). The concentration of free cholesterol in human tears was 15 ± 6 μg/g, which was higher than in rabbit tears (10 ± 5 μg/g). Conclusions. A stable-isotope dilution DI/EI-SIM method for free cholesterol quantification without prior chromatographic separation was established. Using this method demonstrated that humans have higher free cholesterol levels in their tears than rabbits. This is in agreement with previous reports. This paper provides a rapid and reliable method to measure free cholesterol in small-volume clinical samples. © 2013 The Association for Research in Vision and Ophthalmology, Inc.

Automatic classification of human epithelial type 2 cell indirect immunofluorescence images using cell pyramid matching

This paper describes a novel system for automatic classification of images obtained from Anti-Nuclear Antibody (ANA) pathology tests on Human Epithelial type 2 (HEp-2) cells using the Indirect Immunofluorescence (IIF) protocol. The IIF protocol on HEp-2 cells has been the hallmark method to identify the presence of ANAs, due to its high sensitivity and the large range of antigens that can be detected. However, it suffers from numerous shortcomings, such as being subjective as well as time and labour intensive. Computer Aided Diagnostic (CAD) systems have been developed to address these problems, which automatically classify a HEp-2 cell image into one of its known patterns (eg. speckled, homogeneous). Most of the existing CAD systems use handpicked features to represent a HEp-2 cell image, which may only work in limited scenarios. We propose a novel automatic cell image classification method termed Cell Pyramid Matching (CPM), which is comprised of regional histograms of visual words coupled with the Multiple Kernel Learning framework. We present a study of several variations of generating histograms and show the efficacy of the system on two publicly available datasets: the ICPR HEp-2 cell classification contest dataset and the SNPHEp-2 dataset.

Activation of matrix metalloproteinase-2 (MMP-2) by membrane type 1 matrix metalloproteinase through an artificial receptor for ProMMP-2 generates active MMP-2

The suggested model for pro-matrix metalloproteinase-2 (proMMP-2) activation by membrane type 1 MMP (MT1-MMP) implicates the complex between MT1-MMP and tissue inhibitor of MMP-2 (TIMP-2) as a receptor for proMMP-2. To dissect this model and assess the pathologic significance of MMP-2 activation, an artificial receptor for proMMP-2 was created by replacing the signal sequence of TIMP-2 with cytoplasmic/transmembrane domain of type II transmembrane mosaic serine protease (MSP-T2). Unlike TIMP-2, MSP-T2 served as a receptor for proMMP-2 without inhibiting MT1-MMP, and generated TIMP-2-free active MMP-2 even at a low level of MT1-MMP. Thus, MSP-T2 did not affect direct cleavage of the substrate testican-1 by MT1-MMP, whereas TIMP-2 inhibited it even at the level that stimulates proMMP-2 processing. Expression of MSP-T2 in HT1080 cells enhanced MMP-2 activation by endogenous MT1-MMP and caused intensive hydrolysis of collagen gel. Expression of MSP-T2 in U87 glioma cells, which express a trace level of endogenous MT1-MMP, induced MMP-2 activation and enhanced cell-associated protease activity, activation of extracellular signal-regulated kinase, and metastatic ability into chick embryonic liver and lung. MT1-MMP can exert both maximum MMP-2 activation and direct cleavage of substrates with MSP-T2, which cannot be achieved with TIMP-2. These results suggest that MMP-2 activation by MT1-MMP potentially amplifies protease activity, and combination with direct cleavage of substrate causes effective tissue degradation and enhances tumor invasion and metastasis, which highlights the complex role of TIMP-2. MSP-T2 is a unique tool to analyze physiologic and pathologic roles of MMP-2 and MT1-MMP in comparison with TIMP-2.

Substrate choice of membrane-type 1 matrix metalloproteinase is dictated by tissue inhibitor of metalloproteinase-2 levels

Although tissue inhibitor of metalloproteinase-2 (TIMP-2) is known to be not only an inhibitor of matrix metalloproteinases (MMP) but also a cofactor for membrane-type 1 MMP (MT1-MMP)-mediated MMP-2 activation, it is still unclear how TIMP-2 regulates MMP-2 activation and cleavage of substrates by MT1-MMP. In the present study we examined the levels of cell-surface MT1-MMP, MMP-2 activation and cleavage of MT1-MMP substrates in 293T cells transfected with the MT1-MMP and TIMP-2 genes. Co-expression of TIMP-2 at an appropriate level increased the level of cell-surface MT1-MMP, both the TIMP-2-bound and free forms, and generated processed MMP-2 with gelatin-degrading activity. In contrast, MT1-MMP substrates testican-1 and syndecan-1 were cleaved by the cells expressing MT1-MMP, which was inhibited by TIMP-2 even at levels that stimulate MMP-2 activation. These results suggest that TIMP-2 environment determines MT1-MMP substrate choice between direct cleavage of its own substrates and MMP-2 activation.

Implication of collagen type I-induced membrane-type 1-matrix metalloproteinase expression and matrix metalloproteinase-2 activation in the metastatic progression of breast carcinoma

We have previously demonstrated that fibroblasts and invasive human breast carcinoma (HBC) cells specifically activate matrix metalloproteinase- 2 (MMP-2) when cultured on 3-dimensional gels of type I collagen but not a range of other substrates. We show here the constitutive expression of membrane-type 1 (MT1)-MMP in both fibroblasts, and invasive HBC cell lines, that have fibroblastic attributes presumably acquired through an epithelial- to-mesenchymal transition (EMT). Treatment with collagen type I increased the steady-state MT1-MMP mRNA levels in these cells but did not induce either MT1-MMP expression or MMP-2 activation in noninvasive breast carcinoma cell lines, which retain epithelial features. Basal MT3-MMP mRNA expression had a pattern similar to that of MT1-MMP but was not up-regulated by collagen. MT4- MMP mRNA was seen in both invasive and noninvasive HBC cell lines and was also not collagen-regulated, and MT2-MMP mRNA was not detected in any of the HBC cell lines tested. These data support a role for MT1-MMP in the collagen- induced MMP-2-activation seen in these cells. In situ hybridization analysis of archival breast cancer specimens revealed a close parallel in expression of both collagen type I and MT1-MMP mRNA in peritumoral fibroblasts, which was correlated with aggressiveness of the lesion. Relatively high levels of expression of both mRNA species were seen in fibroblasts close to invasive tumor nests and, although only focally, in certain areas close to preinvasive tumors. These foci may represent hot spots for local degradation and invasive progression. Collectively, these results implicate MT1-MMP in collagen- stimulated MMP-2 activation and suggest that this mechanism may be employed in vivo by both tumor-associated fibroblasts and EMT-derived carcinoma cells to facilitate increased invasion and/or metastasis.