A GLP1 receptor agonist diabetes drug ameliorates neurodegeneration in a mouse model of infantile neurometabolic disease.

Infantile neuroaxonal dystrophy (INAD) is a rare paediatric neurodegenerative condition caused by mutations in the PLA2G6 gene, which is also the causative gene for PARK14-linked young adult-onset dystonia parkinsonism. INAD patients usually die within their first decade of life, and there are currently no effective treatments available. GLP1 receptor (GLP-1R) agonists are licensed for treating type 2 diabetes mellitus but have also demonstrated neuroprotective properties in a clinical trial for Parkinson's disease. Therefore, we evaluated the therapeutic efficacy of a new recently licensed GLP-1R agonist diabetes drug in a mouse model of INAD. Systemically administered high-dose semaglutide delivered weekly to juvenile INAD mice improved locomotor function and extended the lifespan. An investigation into the mechanisms underlying these therapeutic effects revealed that semaglutide significantly increased levels of key neuroprotective molecules while decreasing those involved in pro-neurodegenerative pathways. The expression of mediators in both the apoptotic and necroptotic pathways were also significantly reduced in semaglutide treated mice. A reduction of neuronal loss and neuroinflammation was observed. Finally, there was no obvious inflammatory response in wild-type mice associated with the repeated high doses of semaglutide used in this study.

Effects of anterior compartment fasciotomy on intramuscular compartment pressure in patients with chronic exertional compartment syndrome.

BACKGROUND: Patients with chronic exertional compartment syndrome (CECS) have pain during exercise that usually subsides at rest. Diagnosis is usually confirmed by measurement of intramuscular compartment pressure (IMCP) following exclusion of other possible causes. Management usually requires fasciotomy but reported outcomes vary widely. There is little evidence of the effectiveness of fasciotomy on IMCP. Testing is rarely repeated postoperatively and reported follow-up is poor. Improved diagnostic criteria based on preselection and IMCP levels during dynamic exercise testing have recently been reported. OBJECTIVES: (1) To compare IMCP in three groups, one with classical symptoms and no treatment and the other with symptoms of CECS who have been treated with fasciotomy and an asymptomatic control group. (2) Establish if differences in IMCP in these groups as a result of fasciotomy relate to functional and symptomatic improvement. METHODS: Twenty subjects with symptoms of CECS of the anterior compartment, 20 asymptomatic controls and 20 patients who had undergone fasciotomy for CECS were compared. All other possible diagnoses were excluded using rigorous inclusion criteria and MRI. Dynamic IMCP was measured using an electronic catheter wire before, during and after participants exercised on a treadmill during a standardised 15 min exercise challenge. Statistical analysis included t-tests and analysis of variance. RESULTS: Fasciotomy results in reduced IMCP at all time points during a standardised exercise protocol compared with preoperative cases. In subjects responding to fasciotomy, there is a significant reduction in IMCP below that of preoperative groups (P<0.001). Postoperative responders to fasciotomy have no significant differences in IMCP from asymptomatic controls (P=0.182). CONCLUSION: Fasciotomy reduces IMCP in all patients. Larger studies are required to confirm that the reduction in IMCP accounts for differences in functional outcomes and pain reductions seen in postoperative patients with CECS.

Barefoot plantar pressure measurement in Chronic Exertional Compartment Syndrome.

BACKGROUND: Patients with Chronic Exertional Compartment Syndrome (CECS) have exercise-limiting pain that subsides at rest. Diagnosis is confirmed by intramuscular compartment pressure (IMCP) measurement. Accompanying CECS, subjective changes to gait (foot slap) are frequently reported by patients. This has not previously been investigated. The aim of this study was to investigate differences in barefoot plantar pressure (BFPP) between CECS cases and asymptomatic controls prior to the onset of painful symptoms. METHODS: 40 male military volunteers, 20 with symptoms of CECS and 20 asymptomatic controls were studied. Alternative diagnoses were excluded with rigorous inclusion criteria, magnetic resonance imaging and dynamic IMCP measurement. BFPP was measured during walking and marching. Data were analysed for: Stance Time (ST); foot progression angle (FPA); centre of force; plantarflexion rate after heel strike (IFFC-time); the distribution of pressure under the heel; and, the ratio between inner and outer metatarsal loading. Correlation coefficients of each variable with speed and leg length were calculated followed by ANCOVA or t-test. Receiver operating characteristic (ROC) curves were constructed for IFFC-time. RESULTS: Caseshad shorter ST and IFFC-times than controls. FPA was inversely related to walking speed (WS) in controls only. The area under the ROC curve for IFFC-time ranged from 0.746 (95%CI: 0.636-0.87) to 0.773 (95%CI: 0.671-0.875) representing 'fair predictive validity'. CONCLUSION: Patients with CECS have an increased speed of ankle plantarflexion after heel strike that precedes the onset of painful symptoms likely resulting from a mechanical disadvantage of Tibialis Anterior. These findings provide further insight into the pathophysiology of CECS and support further investigation of this non-invasive diagnostic. The predictive value of IFFC-time in the diagnosis of CECS is comparable to post-exercise IMCP but falls short of dynamic IMCP measured during painful symptoms.

Apoptosis progression studied using parallel dielectrophoresis electrophysiological analysis and flow cytometry.

Apoptosis is characterised by many cellular events, but the standard Annexin-V assay identifies two; the transfer of the phospholipid phosphatidylserine (PS) from inner to outer leaflets of the plasma membrane, acting as an "eat me" signal to macrophages, and the permeabilisation of the plasma membrane. In this paper we compare the results from the Annexin-V assay with electrophysiology data obtained in parallel using dielectrophoresis, which highlights two changes in cell electrophysiology; a change in cytoplasmic conductivity which correlates with PS expression, and a membrane conductance spike that correlates with permeabilisation. Combining results from both methods shows a strong inverse relationship between conductivity and PS externalisation. One mechanism which may explain this correlation is related to intracellular Ca(2+), which is known to increase early in apoptosis. PS expression occurs when enzymes called scramblases swap external and internal phospholipids, and which are usually activated by Ca(2+), whilst the change in cytoplasmic conductivity may be due to K(+) efflux from intermediate conductance (IK) ion channels that are also activated by Ca(2+).

A dielectrophoretic method of discrimination between normal oral epithelium, and oral and oropharyngeal cancer in a clinical setting.

Despite the accessibility of the oral cavity to clinical examination, delays in diagnosis of oral and oropharyngeal carcinoma (OOPC) are observed in a large majority of patients, with negative impact on prognosis. Diagnostic aids might help detection and improve early diagnosis, but there remains little robust evidence supporting the use of any particular diagnostic technology at the moment. The aim of the present feasibility first-in-human study was to evaluate the preliminary diagnostic validity of a novel technology platform based on dielectrophoresis (DEP). DEP does not require labeling with antibodies or stains and it is an ideal tool for rapid analysis of cell properties. Cells from OOPC/dysplasia tissue and healthy oral mucosa were collected from 57 study participants via minimally-invasive brush biopsies and tested with a prototype DEP platform using median membrane midpoint frequency as main analysis parameter. Results indicate that the current DEP platform can discriminate between brush biopsy samples from cancerous and healthy oral tissue with a diagnostic sensitivity of 81.6% and a specificity of 81.0%. The present ex vivo results support the potential application of DEP testing for identification of OOPC. This result indicates that DEP has the potential to be developed into a low-cost, rapid platform as an assistive tool for the early identification of oral cancer in primary care; given the rapid, minimally-invasive and non-expensive nature of the test, dielectric characterization represents a promising platform for cost-effective early cancer detection.

Characterization of human skeletal stem and bone cell populations using dielectrophoresis.

Dielectrophoresis (DEP) is a non-invasive cell analysis method that uses differences in electrical properties between particles and surrounding medium to determine a unique set of cellular properties that can be used as a basis for cell separation. Cell-based therapies using skeletal stem cells are currently one of the most promising areas for treating a variety of skeletal and muscular disorders. However, identifying and sorting these cells remains a challenge in the absence of unique skeletal stem cell markers. DEP provides an ideal method for identifying subsets of cells without the need for markers by using their dielectric properties. This study used a 3D dielectrophoretic well chip device to determine the dielectric characteristics of two osteosarcoma cell lines (MG-63 and SAOS-2) and an immunoselected enriched skeletal stem cell fraction (STRO-1 positive cell) of human bone marrow. Skeletal cells were exposed to a series of different frequencies to induce dielectrophoretic cell movement, and a model was developed to generate the membrane and cytoplasmic properties of the cell populations. Differences were observed in the dielectric properties of MG-63, SAOS-2 and STRO-1 enriched skeletal populations, which could potentially be used to sort cells in mixed populations. This study provide evidence of the ability to characterize different human skeletal stem and mature cell populations, and acts as a proof-of-concept that dielectrophoresis can be exploited to detect, isolate and separate skeletal cell populations from heterogeneous bone marrow cell populations.

Dielectrophoretic response of DNA shows different conduction mechanisms for poly(dG)-poly(dC) and poly(dA)-Poly(dT) in solution.

Although the subject of some scrutiny over the years, the mechanism of conduction in DNA has not yet been resolved, with competing theories suggesting either electronic and ionic conduction mechanisms. In this paper we use dielectrophoresis to determine the electrical properties of poly(dG)-poly(dC) (GC) and poly(dA)-poly(dT) (AT) DNA in solution. The molecules show different conduction mechanisms; GC DNA exhibits conduction primarily through the molecule, whereas in AT DNA conduction through the counterion cloud surrounding the molecule in solution is more significant.

Solution processable multi-channel ZnO nanowire field-effect transistors with organic gate dielectric.

The present work focuses on nanowire (NW) applications as semiconducting elements in solution processable field-effect transistors (FETs) targeting large-area low-cost electronics. We address one of the main challenges related to NW deposition and alignment by using dielectrophoresis (DEP) to select multiple ZnO nanowires with the correct length, and to attract, orientate and position them in predefined substrate locations. High-performance top-gate ZnO NW FETs are demonstrated on glass substrates with organic gate dielectric layers and surround source-drain contacts. Such devices are hybrids, in which inorganic multiple single-crystal ZnO NWs and organic gate dielectric are synergic in a single system. Current-voltage (I-V) measurements of a representative hybrid device demonstrate excellent device performance with high on/off ratio of ~10(7), steep subthreshold swing (s-s) of ~400 mV/dec and high electron mobility of ~35 cm(2) V(-1) s(-1) in N2 ambient. Stable device operation is demonstrated after 3 months of air exposure, where similar device parameters are extracted including on/off ratio of ~4 × 10(6), s-s ~500 mV/dec and field-effect mobility of ~28 cm(2) V(-1) s(-1). These results demonstrate that DEP can be used to assemble multiples of NWs from solvent formulations to enable low-temperature hybrid transistor fabrication for large-area inexpensive electronics.

Cancer, pre-cancer and normal oral cells distinguished by dielectrophoresis.

Most oral cancers are oral squamous cell carcinomas (OSCC) that arise from the epithelial lining of the oral mucosa. Given that the oral cavity is easily accessible, the disease lends itself to early detection; however, most oral cancers are diagnosed at a late stage, and approximately half of oral cancer sufferers do not survive beyond five years, post-diagnosis. The low survival rate has been attributed to late detection, but there is no accepted, reliable and convenient method for the detection of oral cancer and oral pre-cancer. Dielectrophoresis (DEP) is a label-free technique which can be used to obtain multi-parametric measurements of cell electrical properties. Parameters such as cytoplasmic conductivity and effective membrane capacitance (C(Eff)) can be non-invasively determined by the technique. In this study, a novel lab-on-a-chip device was used to determine the cytoplasmic conductivity and C(Eff) of primary normal oral keratinocytes, and pre-cancerous and cancerous oral keratinocyte cell lines. Our results show that the electrical properties of normal, pre-cancerous and cancerous oral keratinocytes are distinct. Furthermore, increasing C (Eff) and decreasing cytoplasmic conductivity correlate with disease progression which could prove significant for diagnostic and prognostic applications. DEP has the potential to be used as a non-invasive technique to detect oral cancer and oral pre-cancer. Clinical investigation is needed to establish the reliability and temporal relationship of the correlation between oncologic disease progression and the electrical parameters identified in this study. To use this technique as an OSCC detection tool in a clinical setting, further characterisation and refinement is warranted.

Dielectrophoretic analysis of changes in cytoplasmic ion levels due to ion channel blocker action reveals underlying differences between drug-sensitive and multidrug-resistant leukaemic cells.

Dielectrophoresis (DEP)--the motion of particles in non-uniform AC fields-has been used in the investigation of cell electrophysiology. The technique offers the advantages of rapid determination of the conductance and capacitance of membrane and cytoplasm. However, it is unable to directly determine the ionic strengths of individual cytoplasmic ions, which has potentially limited its application in assessing cell composition. In this paper, we demonstrate how dielectrophoresis can be used to investigate the cytoplasmic ion composition by using ion channel blocking agents. By blocking key ion transporters individually, it is possible to determine their overall contribution to the free ions in the cytoplasm. We use this technique to evaluate the relative contributions of chloride, potassium and calcium ions to the cytoplasmic conductivities of drug sensitive and resistant myelogenous leukaemic (K562) cells in order to determine the contributions of individual ion channel activity in mediating multi-drug resistance in cancer. Results indicate that whilst K(+) and Ca(2+) levels were extremely similar between sensitive and resistant lines, levels of Cl(-) were elevated by three times to that in the resistant line, implying increased chloride channel activity. This result is in line with current theories of MDR, and validates the use of ion channel blockers with DEP to investigate ion channel function.

Parallel measurements of drug actions on Erythrocytes by dielectrophoresis, using a three-dimensional electrode design.

A type of well-based assay that uses a laminated three-dimensional electrode design to characterise the effects of different drugs on red blood cells using dielectrophoresis is presented. The capability of the system to determine the effects of chemical agents on the electrophysiology of red blood cells is demonstrated using saponin and valinomycin as two examples of drugs that can penetrate the cell membrane and therefore change the dielectric properties of the cell. Light intensity changes are measured in the well over a period of time at various frequencies and the dielectric properties of the cells determined using an ellipsoidal multi-shell model. It is shown that the laminated electrode permits a high degree of automation and thus a high number of parallel experiments, which reduces both the time and effort needed to examine differences between populations of red blood cells. The technique is directly compatible with the industry-standard 1536 well-plate analysis technique.

Finite element modelling of a 3 dimensional dielectrophoretic flow separator device for optimal bioprocessing conditions.

Planar 2-dimensional dielectrophoresis electrode geometries are limited in only being capable of handling fluid volumes ranging from picolitres to hundreds of microliters per hour. A 3-dimensional electrode system has been developed capable of handling significantly larger volumes of fluid. Using finite element modeling the electric field distribution within various bore sizes was realized. From these simulations it is possible to optimize bioprocessing factors influencing the performance of a dielectrophoretic separator. Process calculations have shown that flow-rates of 25ml hr/sup -1/ or more can be attained for the separation of heterogeneous populations of bio-particles based on their dielectric properties.

Surface-enhanced resonance Raman scattering of black inkjet dyes in solution and in situ printed onto paper.

Understanding the changes that occur when dyes are absorbed onto paper is crucial for the design of new inkjet dyes. This problem is particularly difficult for black dyes that have complex chromophores, and as a result, spectroscopic information on electronic and structural changes can be of importance. Surface-enhanced resonance Raman scattering (SERRS) and electronic structure calculations were used to probe in situ changes in the chromophore in black di-azo dyes printed onto paper. The data indicate that the low-energy chromophore is due mainly to the hydrazone group and the high-energy chromophore to both the azo and hydrazone groups. A comparison of SERRS from the dyes adsorbed onto silver particles in suspension and from the dyes on paper demonstrated a broadening of the chromophore into the red for both dyes and evidence of a structural change in one dye.

Predictors of serum selenium in cigarette smokers and the lack of association with lung and prostate cancer risk.

Epidemiological studies have suggested that low levels of selenium are associated with a higher incidence of both lung and prostate cancer. We analyzed the selenium serum concentration in 356 Carotene and Retinol Efficacy Trial (CARET) participants who later developed lung cancer and 356 matched controls and in 235 prostate cancer cases and 456 matched controls. Serum samples were obtained a mean of 4.7 years before diagnosis for both tumor types. Controls were matched to cases by year of randomization, age, smoking status, treatment arm, exposure population (asbestos workers or cigarette smokers), and year of blood draw. In the control population (n = 820), significant predictors of low serum selenium concentration were current smoking status and East Coast locations of the study center. Overall, there was no significant difference in mean serum selenium in lung cancer cases versus controls (11.91 microg/dl versus 11.77 microg/dl) or prostate cancer cases versus controls (11.48 microg/dl versus 11.43 microg/dl). No statistically significant trend in odds ratio was seen across quartiles of serum selenium for lung cancer (P = 0.49) or prostate cancer (P = 0.69). In a subpopulation of 174 prostate cancer patients who had clinical and pathological staging material reviewed, there was no association between serum selenium and Gleason score or clinical or pathological stage. In the CARET population of current and former smokers consuming an ad libitum diet, the serum concentration of selenium was not a risk factor for either lung cancer or prostate cancer.

Dielectrophoretic manipulation and characterization of herpes simplex virus-1 capsids.

The dielectrophoretic behaviour of the capsids of herpes simplex virus type-1 has been measured over a range of conductivities of KCl solutions, with and without the addition of mannitol. The dielectrophoretic response of the capsids was recorded by measuring the frequency corresponding to zero dielectrophoretic force. The data were analysed using a multi-shelled model, and the permittivity and conductivity of the particles estimated. The capsid was modelled as a porous protein shell through which suspending medium passes, an inner chamber containing suspending medium in equilibrium with the outside, and a central core of protein (the scaffold). Capsids suspended in KCl without mannitol exhibited a different behaviour to those suspended in KCl with mannitol.

Selenium deficiency is associated with shedding of HIV-1--infected cells in the female genital tract.

OBJECTIVE: To assess the relation between selenium deficiency and vaginal or cervical shedding of HIV-1-infected cells. DESIGN: Cross-sectional study of 318 HIV-1 seropositive women in Mombasa, Kenya. METHODS: Vaginal and cervical swab specimens were tested for the presence of HIV-1 DNA by polymerase chain reaction. Multivariate logistic regression models, adjusting for CD4 count and vitamin A deficiency, were used. RESULTS: Selenium deficiency (defined as levels <85 microg/L) was observed in 11% of the study population. In unstratified multivariate analyses, there was no significant association between selenium deficiency and vaginal or cervical shedding. In stratified analyses, however, significant associations became apparent after excluding women with predictors of shedding with strong local effects on the genital tract mucosa. Among women who did not use oral contraceptives and who did not have vaginal candidiasis, selenium deficiency was significantly associated with vaginal shedding (adjusted odds ratio [AOR] 2.9, 95% confidence interval [CI] 1.0--8.8, p =.05). Effect modification was also observed in the relation between selenium deficiency and cervical shedding, with a significant association seen among those women who were not using oral contraceptive pills or depot medroxyprogesterone acetate and who did not have Neisseria gonorrhoeae infection (AOR 2.8, 95% CI 1.1--7.0, p =.02). CONCLUSIONS: We found selenium deficiency to be associated with a nearly threefold higher likelihood of genital mucosal shedding of HIV-1--infected cells, suggesting that deficiency may increase the infectiousness of women with HIV-1. Nutritional interventions to prevent HIV-1 transmission warrant investigation.

Separation of submicron bioparticles by dielectrophoresis.

Submicron particles such as latex spheres and viruses can be manipulated and characterized using dielectrophoresis. By the use of appropriate microelectrode arrays, particles can be trapped or moved between regions of high or low electric fields. The magnitude and direction of the dielectrophoretic force on the particle depends on its dielectric properties, so that a heterogeneous mixture of particles can be separated to produce a more homogeneous population. In this paper the controlled separation of submicron bioparticles is demonstrated. With electrode arrays fabricated using direct write electron beam lithography, it is shown that different types of submicron latex spheres can be spatially separated. The separation occurs as a result of differences in magnitude and/or direction of the dielectrophoretic force on different populations of particles. These differences arise mainly because the surface properties of submicron particles dominate their dielectrophoretic behavior. It is also demonstrated that tobacco mosaic virus and herpes simplex virus can be manipulated and spatially separated in a microelectrode array.

Measurement of bacterial flagellar thrust by negative dielectrophoresis.

The force produced by the flagella of the bacterium Salmonella typhimurium has been measured using negative dielectrophoretic methods. The bacteria are held in a force funnel, produced using a nonuniform electric field. When the motor force is balanced against an opposing negative dielectrophoretic force the bacteria become motionless. Numerical simulations have been used to estimate the electric field gradient in the electrodes. Together with experimental observations of bacterial motion the data gives a value of the force produced by the bacterial motor to be 0.37 pN.

Detection of domoic acid in northern anchovies and California sea lions associated with an unusual mortality event.

The occurrence of an unusual mortality event involving California sea lions (Zalophus californianus) along the central California coast in May 1998 was recently reported. The potent neurotoxin domoic acid (DA), produced naturally by the diatom Pseudo-nitzschia australis and transmitted to the sea lions via planktivorous northern anchovies (Engraulis mordax), was identified as the probable causative agent. Details of DA analyses for anchovy tissues and sea lion feces are described. Domoic acid levels were estimated in anchovy samples by HPLC-UV, and in sea lion feces using the same method as well as a microplate receptor binding assay, with absolute confirmation by tandem mass spectrometry. The highest DA concentrations in anchovies occurred in the viscera (223 +/- 5 microg DA g(-1)), exceeding values in the body tissues by seven-fold and suggesting minimal bioaccumulation of DA in anchovy tissue. HPLC values for DA in sea lion fecal material (ranging from 152 to 136.5 microg DA g(-1)) required correction for interference from an unidentified compound. Inter-laboratory comparisons of HPLC data showed close quantitative agreement. Fecal DA activity determined using the receptor binding assay corresponded with HPLC values to within a factor of two. Finally, our detection of P. australis frustules, via scanning electron microscopy, in both anchovy viscera and fecal material from sea lions exhibiting seizures provides corroborating evidence that this toxic algal species was involved in this unusual sea lion mortality event.