Effectiveness of porous silicon nanoparticle treatment at inhibiting the migration of a heterogeneous glioma cell population

Background: Approximately 80% of brain tumours are gliomas. Despite treatment, patient mortality remains high due to local metastasis and relapse. It has been shown that transferrin-functionalised porous silicon nanoparticles (Tf@pSiNPs) can inhibit the migration of U87 glioma cells. However, the underlying mechanisms and the effect of glioma cell heterogeneity, which is a hallmark of the disease, on the efficacy of Tf@pSiNPs remains to be addressed. Results: Here, we observed that Tf@pSiNPs inhibited heterogeneous patient-derived glioma cells’ (WK1) migration across small perforations (3 μm) by approximately 30%. A phenotypical characterisation of the migrated subpopulations revealed that the majority of them were nestin and fibroblast growth factor receptor 1 positive, an indication of their cancer stem cell origin. The treatment did not inhibit cell migration across large perforations (8 μm), nor cytoskeleton formation. This is in agreement with our previous observations that cellular-volume regulation is a mediator of Tf@pSiNPs’ cell migration inhibition. Since aquaporin 9 (AQP9) is closely linked to cellular-volume regulation, and is highly expressed in glioma, the effect of AQP9 expression on WK1 migration was investigated. We showed that WK1 migration is correlated to the differential expression patterns of AQP9. However, AQP9-silencing did not affect WK1 cell migration across perforations, nor the efficacy of cell migration inhibition mediated by Tf@pSiNPs, suggesting that AQP9 is not a mediator of the inhibition. Conclusion: This in vitro investigation highlights the unique therapeutic potentials of Tf@pSiNPs against glioma cell migration and indicates further optimisations that are required to maximise its therapeutic efficacies. Graphic Abstract: [Figure not available: see fulltext.

Isolation of Circulating Tumour Cells in Patients With Glioblastoma Using Spiral Microfluidic Technology – A Pilot Study

Glioblastoma (GBM) is the most common and aggressive type of tumour arising from the central nervous system. GBM remains an incurable disease despite advancement in therapies, with overall survival of approximately 15 months. Recent literature has highlighted that GBM releases tumoural content which crosses the blood-brain barrier (BBB) and is detected in patients' blood, such as circulating tumour cells (CTCs). CTCs carry tumour information and have shown promise as prognostic and predictive biomarkers in different cancer types. Currently, there is limited data for the clinical utility of CTCs in GBM. Here, we report the use of spiral microfluidic technology to isolate CTCs from whole blood of newly diagnosed GBM patients before and after surgery, followed by characterization for GFAP, cell-surface vimentin protein expression and EGFR amplification. CTCs were found in 13 out of 20 patients (9/20 before surgery and 11/19 after surgery). Patients with CTC counts equal to 0 after surgery had a significantly longer recurrence-free survival (p=0.0370). This is the first investigation using the spiral microfluidics technology for the enrichment of CTCs from GBM patients and these results support the use of this technology to better understand the clinical value of CTCs in the management of GBM in future studies

Spot the Difference-Development of a Syndrome Based Protein Microarray for Specific Serological Detection of Multiple Flavivirus Infections in Travelers

Background The family Flaviviridae, genus Flavivirus, holds many of the world’s most prevalent arboviral diseases that are also considered the most important travel related arboviral infections. In most cases, flavivirus diagnosis in travelers is primarily based on serology as viremia is often low and typically has already been reduced to undetectable levels when symptoms set in and patients seek medical attention. Serological differentiation between flaviviruses and the false-positive results caused by vaccination and cross-reactivity among the different species, are problematic for surveillance and diagnostics of flaviviruses. Their partially overlapping geographic distribution and symptoms, combined with increase in travel, and preexisting antibodies due to flavivirus vaccinations, expand the need for rapid and reliable multiplex diagnostic tests to supplement currently used methods. Goal We describe the development of a multiplex serological protein microarray using recombinant NS1 proteins for detection of medically important viruses within the genus Flavivirus. Sera from clinical flavivirus patients were used for primary development of the protein microarray. Results Results show a high IgG and IgM sensitivity and specificity for individual NS1 antigens, and limited cross reactivity, even within serocomplexes. In addition, the serology based on this array allows for discrimination between infection and vaccination response for JEV vaccine, and no cross-reactivity with TBEV and YFV vaccine induced antibodies when testing for antibodies to other flaviviruses. Conclusion Based on these data, multiplex NS1-based protein microarray is a promising tool for surveillance and diagnosis of flaviviruses.

Cross-sectional associations between sleep duration, sedentary time, physical activity, and adiposity indicators among Canadian preschool-aged children using compositional analyses

Abstract Background Sleep duration, sedentary behaviour, and physical activity are three co-dependent behaviours that fall on the movement/non-movement intensity continuum. Compositional data analyses provide an appropriate method for analyzing the association between co-dependent movement behaviour data and health indicators. The objectives of this study were to examine: (1) the combined associations of the composition of time spent in sleep, sedentary behaviour, light-intensity physical activity (LPA), and moderate- to vigorous-intensity physical activity (MVPA) with adiposity indicators; and (2) the association of the time spent in sleep, sedentary behaviour, LPA, or MVPA with adiposity indicators relative to the time spent in the other behaviours in a representative sample of Canadian preschool-aged children. Methods Participants were 552 children aged 3 to 4 years from cycles 2 and 3 of the Canadian Health Measures Survey. Sedentary time, LPA, and MVPA were measured with Actical accelerometers (Philips Respironics, Bend, OR USA), and sleep duration was parental reported. Adiposity indicators included waist circumference (WC) and body mass index (BMI) z-scores based on World Health Organization growth standards. Compositional data analyses were used to examine the cross-sectional associations. Results The composition of movement behaviours was significantly associated with BMI z-scores (p = 0.006) but not with WC (p = 0.718). Further, the time spent in sleep (BMI z-score: γ sleep  = −0.72; p = 0.138; WC: γ sleep  = −1.95; p = 0.285), sedentary behaviour (BMI z-score: γ SB  = 0.19; p = 0.624; WC: γ SB  = 0.87; p = 0.614), LPA (BMI z-score: γ LPA  = 0.62; p = 0.213, WC: γ LPA  = 0.23; p = 0.902), or MVPA (BMI z-score: γ MVPA  = −0.09; p = 0.733, WC: γ MVPA  = 0.08; p = 0.288) relative to the other behaviours was not significantly associated with the adiposity indicators. Conclusions This study is the first to use compositional analyses when examining associations of co-dependent sleep duration, sedentary time, and physical activity behaviours with adiposity indicators in preschool-aged children. The overall composition of movement behaviours appears important for healthy BMI z-scores in preschool-aged children. Future research is needed to determine the optimal movement behaviour composition that should be promoted in this age group

Shaping bursting by electrical coupling and noise

Gap-junctional coupling is an important way of communication between neurons and other excitable cells. Strong electrical coupling synchronizes activity across cell ensembles. Surprisingly, in the presence of noise synchronous oscillations generated by an electrically coupled network may differ qualitatively from the oscillations produced by uncoupled individual cells forming the network. A prominent example of such behavior is the synchronized bursting in islets of Langerhans formed by pancreatic \beta-cells, which in isolation are known to exhibit irregular spiking. At the heart of this intriguing phenomenon lies denoising, a remarkable ability of electrical coupling to diminish the effects of noise acting on individual cells. In this paper, we derive quantitative estimates characterizing denoising in electrically coupled networks of conductance-based models of square wave bursting cells. Our analysis reveals the interplay of the intrinsic properties of the individual cells and network topology and their respective contributions to this important effect. In particular, we show that networks on graphs with large algebraic connectivity or small total effective resistance are better equipped for implementing denoising. As a by-product of the analysis of denoising, we analytically estimate the rate with which trajectories converge to the synchronization subspace and the stability of the latter to random perturbations. These estimates reveal the role of the network topology in synchronization. The analysis is complemented by numerical simulations of electrically coupled conductance-based networks. Taken together, these results explain the mechanisms underlying synchronization and denoising in an important class of biological models

Discovery of a diaminoquinoxaline benzenesulfonamide antagonist of HIV-1 Nef function using a yeast-based phenotypic screen

Background: HIV-1 Nef is a viral accessory protein critical for AIDS progression. Nef lacks intrinsic catalytic activity and binds multiple host cell signaling proteins, including Hck and other Src-family tyrosine kinases. Nef binding induces constitutive Hck activation that may contribute to HIV pathogenesis by promoting viral infectivity, replication and downregulation of cell-surface MHC-I molecules. In this study, we developed a yeast-based phenotypic screen to identify small molecules that inhibit the Nef-Hck complex. Results: Nef-Hck interaction was faithfully reconstituted in yeast cells, resulting in kinase activation and growth arrest. Yeast cells expressing the Nef-Hck complex were used to screen a library of small heterocyclic compounds for their ability to rescue growth inhibition. The screen identified a dihydrobenzo-1,4-dioxin-substituted analog of 2-quinoxalinyl-3-aminobenzene-sulfonamide (DQBS) as a potent inhibitor of Nef-dependent HIV-1 replication and MHC-I downregulation in T-cells. Docking studies predicted direct binding of DQBS to Nef which was confirmed in differential scanning fluorimetry assays with recombinant purified Nef protein. DQBS also potently inhibited the replication of HIV-1 NL4-3 chimeras expressing Nef alleles representative of all M-group HIV-1 clades.Conclusions: Our findings demonstrate the utility of a yeast-based growth reversion assay for the identification of small molecule Nef antagonists. Inhibitors of Nef function discovered with this assay, such as DQBS, may complement the activity of current antiretroviral therapies by enabling immune recognition of HIV-infected cells through the rescue of cell surface MHC-I. © 2013 Trible et al.; licensee BioMed Central Ltd

Intermittent Hypoxia-Induced Cognitive Deficits Are Mediated by NADPH Oxidase Activity in a Murine Model of Sleep Apnea

Background: In rodents, exposure to intermittent hypoxia (IH), a hallmark of obstructive sleep apnea (OSA), is associated with neurobehavioral impairments, increased apoptosis in the hippocampus and cortex, as well as increased oxidant stress and inflammation. Excessive NADPH oxidase activity may play a role in IH-induced CNS dysfunction. Methods and Findings: The effect of IH during light period on two forms of spatial learning in the water maze and well as markers of oxidative stress was assessed in mice lacking NADPH oxidase activity (gp91phox _/Y) and wild-type littermates. On a standard place training task, gp91phox _/Y displayed normal learning, and were protected from the spatial learning deficits observed in wild-type littermates exposed to IH. Moreover, anxiety levels were increased in wild-type mice exposed to IH as compared to room air (RA) controls, while no changes emerged in gp91phox _/Y mice. Additionally, wild-type mice, but not gp91phox _/Y mice had significantly elevated levels of NADPH oxidase expression and activity, as well as MDA and 8-OHDG in cortical and hippocampal lysates following IH exposures. Conclusions: The oxidative stress responses and neurobehavioral impairments induced by IH during sleep are mediated, at least in part, by excessive NADPH oxidase activity, and thus pharmacological agents targeting NADPH oxidase may provid

Metagenomic analysis of the turkey gut RNA virus community

Viral enteric disease is an ongoing economic burden to poultry producers worldwide, and despite considerable research, no single virus has emerged as a likely causative agent and target for prevention and control efforts. Historically, electron microscopy has been used to identify suspect viruses, with many small, round viruses eluding classification based solely on morphology. National and regional surveys using molecular diagnostics have revealed that suspect viruses continuously circulate in United States poultry, with many viruses appearing concomitantly and in healthy birds. High-throughput nucleic acid pyrosequencing is a powerful diagnostic technology capable of determining the full genomic repertoire present in a complex environmental sample. We utilized the Roche/454 Life Sciences GS-FLX platform to compile an RNA virus metagenome from turkey flocks experiencing enteric disease. This approach yielded numerous sequences homologous to viruses in the BLAST nr protein database, many of which have not been described in turkeys. Our analysis of this turkey gut RNA metagenome focuses in particular on the turkey-origin members of the Picornavirales, the Caliciviridae, and the turkey Picobirnaviruses

PACE - The first placebo controlled trial of paracetamol for acute low back pain: design of a randomised controlled trial

<p>Abstract</p> <p>Background</p> <p>Clinical practice guidelines recommend that the initial treatment of acute low back pain (LBP) should consist of advice to stay active and regular simple analgesics such as paracetamol 4 g daily. Despite this recommendation in all international LBP guidelines there are no placebo controlled trials assessing the efficacy of paracetamol for LBP at any dose or dose regimen. This study aims to determine whether 4 g of paracetamol daily (in divided doses) results in a more rapid recovery from acute LBP than placebo. A secondary aim is to determine if ingesting paracetamol in a time-contingent manner is more effective than paracetamol taken when required (PRN) for recovery from acute LBP.</p> <p>Methods/Design</p> <p>The study is a randomised double dummy placebo controlled trial. 1650 care seeking people with significant acute LBP will be recruited. All participants will receive advice to stay active and will be randomised to 1 of 3 treatment groups: time-contingent paracetamol dose regimen (plus placebo PRN paracetamol), PRN paracetamol (plus placebo time-contingent paracetamol) or a double placebo study arm. The primary outcome will be time (days) to recovery from pain recorded in a daily pain diary. Other outcomes will be pain intensity, disability, function, global perceived effect and sleep quality, captured at baseline and at weeks 1, 2, 4 and 12 by an assessor blind to treatment allocation. An economic analysis will be conducted to determine the cost-effectiveness of treatment from the health sector and societal perspectives.</p> <p>Discussion</p> <p>The successful completion of the trial will provide the first high quality evidence on the effectiveness of the use of paracetamol, a guideline endorsed treatment for acute LBP.</p> <p>Trail registration</p> <p>ACTRN12609000966291.</p

Patient-derived glioblastoma cells show significant heterogeneity in treatment responses to the inhibitor-of-apoptosis-protein antagonist birinapant.

BACKGROUND: Resistance to temozolomide (TMZ) greatly limits chemotherapeutic effectiveness in glioblastoma (GBM). Here we analysed the ability of the Inhibitor-of-apoptosis-protein (IAP) antagonist birinapant to enhance treatment responses to TMZ in both commercially available and patient-derived GBM cells. METHODS: Responses to TMZ and birinapant were analysed in a panel of commercial and patient-derived GBM cell lines using colorimetric viability assays, flow cytometry, morphological analysis and protein expression profiling of pro- and antiapoptotic proteins. Responses in vivo were analysed in an orthotopic xenograft GBM model. RESULTS: Single-agent treatment experiments categorised GBM cells into TMZ-sensitive cells, birinapant-sensitive cells, and cells that were insensitive to either treatment. Combination treatment allowed sensitisation to therapy in only a subset of resistant GBM cells. Cell death analysis identified three principal response patterns: Type A cells that readily activated caspase-8 and cell death in response to TMZ while addition of birinapant further sensitised the cells to TMZ-induced cell death; Type B cells that readily activated caspase-8 and cell death in response to birinapant but did not show further sensitisation with TMZ; and Type C cells that showed no significant cell death or moderately enhanced cell death in the combined treatment paradigm. Furthermore, in vivo, a Type C patient-derived cell line that was TMZ-insensitive in vitro and showed a strong sensitivity to TMZ and TMZ plus birinapant treatments. CONCLUSIONS: Our results demonstrate remarkable differences in responses of patient-derived GBM cells to birinapant single and combination treatments, and suggest that therapeutic responses in vivo may be greatly affected by the tumour microenvironment