Serine protease inhibition and mitochondrial dysfunction associated with cisplatin resistance in human tumor cell lines: targets for therapy

Indicators of mitochondrial function were studied in two different cell culture models of cis-diamminedichloroplatinum-II (CDDP) resistance: the intrinsically resistant human ovarian cancer cell line CI-80-13S, and resistant clones (HeLa-S1a and HeLa-S1b) generated by stable expression of the serine protease inhibitor—plasminogen activator inhibitor type-2 (PAI-2), in the human cervical cancer cell line HeLa. In both models, CDDP resistance was associated with sensitivity to killing by adriamycin, etoposide, auranofin, bis[1,2-bis(diphenylphosphino)ethane]gold(I) chloride {[Au(DPPE)2]Cl}, CdCl2 and the mitochondrial inhibitors rhodamine-123 (Rhl23), dequalinium chloride (DeCH), tetraphenylphosphonium (TPP), and ethidium bromide (EtBr) and with lower constitutive levels of ATP. Unlike the HeLa clones, CI-80-13S cells were additionally sensitive to chloramphenicol, 1-methyl-4-phenylpyridinium ion (MPP+), rotenone, thenoyltrifluoroacetone (TTFA), and antimycin A, and showed poor reduction of 1-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT), suggesting a deficiency in NADH dehydrogenase and/or succinate dehydrogenase activities. Total platinum uptake and DNA-bound platinum were slightly lower in CI-80-13S than in sensitive cells. The HeLa-S1a and HeLa-S1b clones, on the other hand, showed poor reduction of triphenyltetrazolium chloride (TTC), indicative of low cytochrome c oxidase activity. Total platinum uptake by HeLa-S1a was similar to HeLa, but DNA-bound platinum was much lower than for the parent cell line. The mitochondria of CI-80-13S and HeLa-S1a showed altered morphology and were fewer in number than those of JAM and HeLa. In both models, CDDP resistance was associated with less platinum accumulation and with mitochondrial and membrane defects, brought about one case with expression of a protease inhibitor which is implicated in tumor progression. Such markers may identify tumors suitable for treatment with gold phosphine complexes or other mitochondrial inhibitors.

Protective effects of sweroside on human MG-63 cells and rat osteoblasts

Herbal Fructus Corni is a folk medicine with a long history of safe use for treating osteoporosis in postmenopausal women or elderly men in Asia. Sweroside is a bioactive herbal ingredient isolated from Fructus Corni, which has been widely used for the treatment of osteoporosis in traditional Chinese medicine (TCM). Unfortunately, the working mechanisms of this compound are difficult to determine and thus remain unclear. The aim of the study was performed to determine the potential molecular mechanism of the anti-osteoporotic effect of sweroside on the human MG-63 cells and rat osteoblasts. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test was used to observe the effect of sweroside on cell proliferation. The activity of alkaline phosphatase (ALP) and the amount of osteocalcin were also assayed the cell differentiation. Sweroside significantly increased the proliferation of human MG-63 cells and rat osteoblasts (P<0.01). It increased the activity of ALP, and osteocalcin was also elevated in response to sweroside (P<0.05). Of note, flowcytometer assay showed that sweroside can attenuate and inhibit apoptosis. Sweroside has a direct osteogenic effect on the proliferation and differentiation of cultured human MG-63 cells and rat osteoblasts in vitro. These data will help in understanding the molecular mechanisms of therapeutic efficacy of sweroside, and highlight insights into drug discovery. In the current study, sweroside has been suggested to be a promising osteoporosis therapeutic natural product.

Characterisation of combustion related acoustic emission sources for diesel engine condition monitoring

This thesis represents a major step forward in understanding the link between the development of combustion related faults in diesel engines and the generation of acoustic emissions. The findings presented throughout the thesis provide a foundation so that future diesel engine monitoring systems are able to more effectively detect and monitor developing faults. In undertaking this research knowledge concerning engine function and relevant failure mechanisms was combined with different modelling methods to generate a framework that was used to effectively identify fault related activity within acoustic emissions recorded from different engines.

Isolation of bioactive compounds that relate to the anti-platelet activity of Cymbopogon ambiguus

Infusions and decoctions of Cymbopogon ambiguus have been used traditionally in Australia for the treatment of headache, chest infections and muscle cramps. The aim of the present study was to screen and identify bioactive compounds from C. ambiguus that could explain this plant's anti-headache activity. A dichloromethane extract of C. ambiguus was identified as having activity in adenosine-diphosphate-induced human platelet aggregation and serotonin-release inhibition bioassays. Subsequent fractionation of this extract led to the isolation of four phenylpropenoids, eugenol, elemicin, Eugenol methylether and trans-isoelemicin. While both Eugenol and elemicin exhibited dose-dependent inhibition of ADP-induced human platelet serotonin release, only eugenol displayed potent inhibitory activity with an IC(50) value of 46.6 microM, in comparison to aspirin, with an IC(50) value of 46.1 microM. These findings provide evidence to support the therapeutic efficacy of C. ambiguus in the non-conventional treatment of Headache and Inflammatory conditions.

The microbial content of vacuum cleaner bag dust and emitted bioaerosols : implications for human exposures indoors

Vacuum cleaners can release large concentrations of particles, both in their exhaust air and from resuspension of settled dust. However, the size, variability and microbial diversity of these emissions are unknown, despite evidence to suggest they may contribute to allergic responses and infection transmission indoors. This study aimed to evaluate bioaerosol emission from various vacuum cleaners. We sampled the air in an experimental flow tunnel where vacuum cleaners were run and their airborne emissions sampled with closed-face cassettes. Dust samples were also 35 collected from the dust bag. Total bacteria, total archaea, Penicillium/Aspergillus and total Clostridium cluster 1 were quantified with specific qPCR protocols and emission rates were calculated. Clostridium botulinum, as well as antibiotic resistance genes were detected in each sample using endpoint PCR. Bacterial diversity was also analyzed using denaturing gel electrophoresis (DGGE), image analysis and band sequencing. We demonstrated that emission of bacteria and moulds (Pen/Asp) can reach values as high as 1E05/min and that those emissions are not related to each other. The bag dust bacterial and mould content was also consistently across the vacuums we assessed, reaching up to 1E07 bacteria or moulds equivalent/g. Antibiotic resistance genes were detected in several samples. No archaea or C. botulinum were detected in any air samples. Diversity analyses showed that most bacteria are from human sources, in keeping with other recent results. These results highlight the potential capability of vacuum cleaners to disseminate appreciable quantities of moulds and human-associated bacteria indoors and their role as a source of exposure to bioaerosols.

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.

Relationship of moderate and low isometric lumbar extension through architectural and muscular activity variables: a cross sectional study

Background No study relating the changes obtained in the architecture of erector spinae (ES) muscle were registered with ultrasound and different intensities of muscle contraction recorded by surface EMG (electromyography) on the ES muscle was found. The aim of this study was analyse the relationship in the response of the ES muscle during isometric moderate and light lumbar isometric extension considering architecture and functional muscle variables. Methods Cross-sectional study. 46 subjects (52% men) with a group mean age of 30.4 (±7.78). The participants developed isometric lumbar extension while performing moderate and low isometric trunk and hip extension in a sitting position with hips flexed 90 degrees and the lumbar spine in neutral position. During these measurements, electromyography recordings and ultrasound images were taken bilaterally. Bilaterally pennation angle, muscle thickness, torque and muscle activation were measured. This study was developed at the human movement analysis laboratory of the Health Science Faculty of the University of Malaga (Spain). Results Strong and moderate correlations were found at moderate and low intensities contraction between the variable of the same intensity, with correlation values ranging from 0.726 (Torque Moderate – EMG Left Moderate) to 0.923 (Angle Left Light – Angle Right Light) (p < 0.001). This correlation is observed between the variables that describe the same intensity of contraction, showing a poor correlation between variables of different intensities. Conclusion There is a strong relationship between architecture and function variables of ES muscle when describe an isometric lumbar extension at light or moderate intensity. Keywords: Ultrasonography; Surface electromyography; Thickness; Pennation angle; Erector spinae

Exploring the role of activity in genre

This paper provides an outline of genre as we currently know it, and examines the changes occurring as games become more complex. Recent research we've undertaken suggests that our perception of which games fall into which genre category is subjective and that genre hybridization continues to blur our understanding of these categories. Consequently, it is becoming increasingly difficult to categorise game play experience based on traditional genre classifications. We examine the use of videogame activities as a useful mechanism for supplementing our understanding of videogame genre. Through considering activity as a means of classifying game experiences we may obtain a much more nuanced understanding of how players engage with games within a particular genre and across genres.

The role of the molecular footprint of EGFR in tailoring treatment decisions in NSCLC

The majority of patients with non-small-cell lung cancer (NSCLC) present with advanced disease, with targeted therapies providing some improvement in clinical outcomes. The epidermal growth factor receptor (EGFR) tyrosine kinase (TK) plays an important role in the pathogenesis of NSCLC. Tyrosine kinase inhibitors (TKIs), which target the EGFR TK domain, have proven to be an effective treatment strategy; however, patient responses to treatment vary considerably. Therefore, the identification of patients most likely to respond to treatment is essential to optimise the benefit of TKIs. Tumour-associated activating mutations in EGFR can identify patients with NSCLC who are likely to have a good response to TKIs. Nonetheless, the majority of patients relapse within a year of starting treatment. Studies of tumours at relapse have demonstrated expression of a T790M mutation in exon 20 of the EGFR TK domain in approximately 50% of cases. Although conferring resistance to reversible TKIs, these patients may remain sensitive to new-generation irreversible/panerb inhibitors. A number of techniques have been employed for genotypic assessment of tumourassociated DNA to identify EGFR mutations, each of which has advantages and disadvantages. This review presents an overview of the current methodologies used to identify such molecular markers. Recent developments in technology may make the monitoring of changes in patients' tumour genotypes easier in clinical practice, which may enable patients' treatment regimens to be tailored during the course of their disease, potentially leading to improved patient outcomes.

bcl-2 and c-erbB-2 proteins are involved in the regulation of VEGF and of thymidine phosphorylase angiogenic activity in non-small-cell lung cancer

Tumour angiogenesis has been recently recognised as one of the most important prognostic factors in lung cancer. Although a variety of angiogenic factors have been identified, the angiogenesis process remains poorly understood. Bcl-2, c-erbB-2 and p53 are well-known oncogenes involved in non- small-cell lung cancer pathogenesis. A direct correlation of thymidine phosphorylase (TP) and of vascular endothelial growth factor (VEGF) with intratumoural angiogenesis has been reported. In the present study we investigated the possible regulatory role if bcl-2, c-erB-2 proteins in angiogenesis and in VEGF and TP expression in non-small-cell lung cancer. Two hundred sixteen specimens from T1,2-NO, 1 staged patients treated with surgery alone were immunohistochemically examined. Bcl-2 and c-erbB-2 were significantly inversely related to each other (P = 0.04) and both were inversely associated with microvessel density (P < 0.02). High TP and VEGF reactivity was statistically related to loss of bcl-2 expression (P < 0.01). A significant co-expression of c-erbB-2 with TP was noted (P = 0.01). However, TP expression was related to high angiogenesis only in cases with absence of c-erB-2 expression (P < 0.0001). c-erbB-2 expression in poorly vascularised tumours was linked with poor outcome (P = 0.03). The present study provides strong evidence that the bcl-2 gene has a suppressive function over genes involved in both angiogenesis (VEGF and TP) and cell migration (c- erbB-2) in NSCLC. TP and c-erbB-2 proteins are significantly, and often simultaneously, expressed in bcl-2 negative cases. However, expression of the c-erbB-2 abolishes the TP-related angiogenic activity. Whether this is a result of a direct activity of the c-erbB-2 protein or a consequence of a c- erbB-2-related immune response remains to be further investigated.

Reduced biceps femoris myoelectrical activity influences eccentric knee flexor weakness after repeat sprint running

The aim of this study was to determine whether declines in knee flexor strength following overground repeat sprints were related to changes in hamstrings myoelectrical activity. Seventeen recreationally active males completed maximal isokinetic concentric and eccentric knee flexor strength assessments at 1800.s-1 before and after repeat sprint running. Myoelectrical activity of the biceps femoris (BF) and medial hamstrings (MH) was measured during all isokinetic contractions. Repeated measures mixed model (Fixed factors = time [pre- and post- repeat sprint] and leg [dominant and non-dominant], random factor = participants) design was fitted with the restricted maximal likelihood method. Repeat sprint running resulted in significant declines in eccentric, and concentric, knee flexor strength (eccentric = 25 ± 34 Nm, 15% p<0.001; concentric 11 Nm± 22 Nm, 10% p = 0.001). Eccentric BF myoelectrical activity was significantly reduced (10%; p= 0.033). Concentric BF and all MH myoelectrical activity were not altered. The declines in maximal eccentric torque were associated with the change in eccentric biceps femoris myoelectrical activity (p = 0.013). Following repeat sprint running there were preferential declines in the myoelectrical activity of the BF, which explained declines in eccentric knee flexor strength.

Time course-dependent changes in the transcriptome of human skeletal muscle during recovery from endurance exercise: From inflammation to adaptive remodeling

Re-programming of gene expression is fundamental for skeletal muscle adaptations in response to endurance exercise. This study investigated the time-course dependent changes in the muscular transcriptome following an endurance exercise trial consisting of 1 h of intense cycling immediately followed by 1 h of intense running. Skeletal muscle samples were taken at baseline, 3 h, 48 h, and 96 h post-exercise from eight healthy, endurance-trained, male individuals. RNA was extracted from muscle. Differential gene expression was evaluated using Illumina microarrays and validated with qPCR. Gene set enrichment analysis identified enriched molecular signatures chosen from the Molecular Signatures Database. Three h post-exercise, 102 gene sets were up-regulated [family wise error rate (FWER), P < 0.05]; including groups of genes related with leukocyte migration, immune and chaperone activation, and cyclic AMP responsive element binding protein (CREB) 1-signaling. Forty-eight h post-exercise, among 19 enriched gene sets (FWER, P < 0.05), two gene sets related to actin cytoskeleton remodeling were up-regulated. Ninety-six h post-exercise, 83 gene sets were enriched (FWER, P < 0.05), 80 of which were up-regulated; including gene groups related to chemokine signaling, cell stress management, and extracellular matrix remodeling. These data provide comprehensive insights into the molecular pathways involved in acute stress, recovery, and adaptive muscular responses to endurance exercise. The novel 96 h post-exercise transcriptome indicates substantial transcriptional activity, potentially associated with the prolonged presence of leukocytes in the muscles. This suggests that muscular recovery, from a transcriptional perspective, is incomplete 96 h after endurance exercise involving muscle damage.

Outsider board activity, ownership structure and firm value : evidence from Korea

Using a sample of publicly listed firm in Korea from 2002 to 2006, this article examines the impact of board monitoring on firm value and productivity. We use outsider's attendance of board meetings as a proxy for board monitoring. Consistent with the commitment hypothesis, we find that outsider's attendance rate increases firm value, suggesting that attending board meeting itself is a strong signal that reflects outsider's intention to monitor insiders. While ownership of controlling shareholders negatively affects firm value, this relationship is not moderated by increased monitoring by outsiders. Our findings provide further evidence that the outside director system is less effective in chaebol-affiliated firms. Results also indicate that the effect of outsider's board monitoring activity on investor's valuation of the firm is greater than on productivity improvement of the firm. Our conclusions are robust for possible endogeneity in the relationship between firm value and board attendance by outside directors.

Monitoring policy and actions on food environments : rationale and outline of the INFORMAS policy engagement and communication strategies

The International Network for Food and Obesity/non-communicable diseases Research, Monitoring and Action Support (INFORMAS) proposes to collect performance indicators on food policies, actions and environments related to obesity and non-communicable diseases. This paper reviews existing communications strategies used for performance indicators and proposes the approach to be taken for INFORMAS. Twenty-seven scoring and rating tools were identified in various fields of public health including alcohol, tobacco, physical activity, infant feeding and food environments. These were compared based on the types of indicators used and how they were quantified, scoring methods, presentation and the communication and reporting strategies used. There are several implications of these analyses for INFORMAS: the ratings/benchmarking approach is very commonly used, presumably because it is an effective way to communicate progress and stimulate action, although this has not been formally evaluated; the tools used must be trustworthy, pragmatic and policy-relevant; multiple channels of communication will be needed; communications need to be tailored and targeted to decision-makers; data and methods should be freely accessible. The proposed communications strategy for INFORMAS has been built around these lessons to ensure that INFORMAS's outputs have the greatest chance of being used to improve food environments.