Abstract
High-throughput gene expression analysis is widely used. However, analysis is not straightforward. Multiple approaches should be applied and methods to combine their results implemented and ...investigated. We present methodology for the comprehensive analysis of expression data, including co-expression module detection and result integration via data-fusion, threshold based methods, and a Naïve Bayes classifier trained on simulated data. Application to rare-disease model datasets confirms existing knowledge related to immune cell infiltration and suggest novel hypotheses including the role of calcium channels. Application to simulated and spike-in experiments shows that combining multiple methods using consensus and classifiers leads to optimal results. ExpHunter Suite is implemented as an R/Bioconductor package available from
https://bioconductor.org/packages/ExpHunterSuite
. It can be applied to model and non-model organisms and can be run modularly in R; it can also be run from the command line, allowing scalability with large datasets. Code and reports for the studies are available from
https://github.com/fmjabato/ExpHunterSuiteExamples
.
Oxidative stress contributes to dysfunction of glial cells in the optic nerve head (ONH). However, the biological basis of the precise functional role of mitochondria in this dysfunction is not fully ...understood. Coenzyme Q10 (CoQ10), an essential cofactor of the electron transport chain and a potent antioxidant, acts by scavenging reactive oxygen species (ROS) for protecting neuronal cells against oxidative stress in many neurodegenerative diseases. Here, we tested whether hydrogen peroxide (100 μM H2O2)-induced oxidative stress alters the mitochondrial network, oxidative phosphorylation (OXPHOS) complex (Cx) expression and bioenergetics, as well as whether CoQ10 can ameliorate oxidative stress-mediated alterations in mitochondria of the ONH astrocytes in vitro. Oxidative stress triggered the activation of ONH astrocytes and the upregulation of superoxide dismutase 2 (SOD2) and heme oxygenase-1 (HO-1) protein expression in the ONH astrocytes. In contrast, CoQ10 not only prevented activation of ONH astrocytes but also significantly decreased SOD2 and HO-1 protein expression in the ONH astrocytes against oxidative stress. Further, CoQ10 prevented a significant loss of mitochondrial mass by increasing mitochondrial number and volume density and by preserving mitochondrial cristae structure, as well as promoted mitofilin and peroxisome-proliferator-activated receptor-γ coactivator-1 protein expression in the ONH astrocyte, suggesting an induction of mitochondrial biogenesis. Finally, oxidative stress triggered the upregulation of OXPHOS Cx protein expression, as well as reduction of cellular adeonsine triphosphate (ATP) production and increase of ROS generation in the ONH astocytes. However, CoQ10 preserved OXPHOS protein expression and cellular ATP production, as well as decreased ROS generation in the ONH astrocytes. On the basis of these observations, we suggest that oxidative stress-mediated mitochondrial dysfunction or alteration may be an important pathophysiological mechanism in the dysfunction of ONH astrocytes. CoQ10 may provide new therapeutic potentials and strategies for protecting ONH astrocytes against oxidative stress-mediated mitochondrial dysfunction or alteration in glaucoma and other optic neuropathies.
PMM2-CDG (MIM # 212065), the most common congenital disorder of glycosylation, is caused by the deficiency of phosphomannomutase 2 (PMM2). It is a multisystemic disease of variable severity that ...particularly affects the nervous system; however, its molecular pathophysiology remains poorly understood. Currently, there is no effective treatment. We performed an RNA-seq based transcriptomic study using patient-derived fibroblasts to gain insight into the mechanisms underlying the clinical symptomatology and to identify druggable targets. Systems biology methods were used to identify cellular pathways potentially affected by PMM2 deficiency, including Senescence, Bone regulation, Cell adhesion and Extracellular Matrix (ECM) and Response to cytokines. Functional validation assays using patients' fibroblasts revealed defects related to cell proliferation, cell cycle, the composition of the ECM and cell migration, and showed a potential role of the inflammatory response in the pathophysiology of the disease. Furthermore, treatment with a previously described pharmacological chaperone reverted the differential expression of some of the dysregulated genes. The results presented from transcriptomic data might serve as a platform for identifying therapeutic targets for PMM2-CDG, as well as for monitoring the effectiveness of therapeutic strategies, including pharmacological candidates and mannose-1-P, drug repurposing.
•Patient-derived fibroblasts from PMM2-CDG are models for studies of pathomechanisms underlying the clinical symptomatology.•Identification of novel druggable targets in PMM2-CDG by transcriptomic analysis.•Senescence, bone regulation, cell adhesion, ECM, and cytokine response are affected pathways in PMM2-CDG.
Elevated rates of burnout and post-traumatic stress have been found in staff working in critical care settings, but the aspect of moral distress has been harder to quantify until a recent revision of ...a scale previously designed for nurses, was adapted for use with a range of health professionals, including physicians. In this cross-sectional survey, n = 171 nurses and physicians working in intensive care in the United Kingdom completed the Moral Distress Scale-Revised in relation to their experiences at work. Mean (SD) Moral Distress Scale-Revised score was 70.2 (39.6). Significant associations were found with female gender (female 74.1 (40.2) vs. male 55.5 (33.8), p = 0.010); depression (r = 0.165, p = 0.035) and with intention to leave job (considering leaving 85.5 (42.4) vs. not considering leaving 67.2 (38.6), p = 0.040). These results highlight the importance of considering the moral impact of work-related issues when addressing staff wellbeing in critical care settings.
Glutamate excitotoxicity leads to fragmented mitochondria in neurodegenerative diseases, mediated by nitric oxide and S-nitrosylation of dynamin-related protein 1, a mitochondrial outer membrane ...fission protein. Optic atrophy gene 1 (OPA1) is an inner membrane protein important for mitochondrial fusion. Autosomal dominant optic atrophy (ADOA), caused by mutations in OPA1, is a neurodegenerative disease affecting mainly retinal ganglion cells (RGCs). Here, we showed that OPA1 deficiency in an ADOA model influences N-methyl-D-aspartate (NMDA) receptor expression, which is involved in glutamate excitotoxicity and oxidative stress. Opa1(enu/+) mice show a slow progressive loss of RGCs, activation of astroglia and microglia, and pronounced mitochondrial fission in optic nerve heads as found by electron tomography. Expression of NMDA receptors (NR1, 2A, and 2B) in the retina of Opa1(enu/+) mice was significantly increased as determined by western blot and immunohistochemistry. Superoxide dismutase 2 (SOD2) expression was significantly decreased, the apoptotic pathway was activated as Bax was increased, and phosphorylated Bad and BcL-xL were decreased. Our results conclusively demonstrate that not only glutamate excitotoxicity and/or oxidative stress alters mitochondrial fission/fusion, but that an imbalance in mitochondrial fission/fusion in turn leads to NMDA receptor upregulation and oxidative stress. Therefore, we propose a new vicious cycle involved in neurodegeneration that includes glutamate excitotoxicity, oxidative stress, and mitochondrial dynamics.
An optical waveguide array biosensor suitable for rapid detection of multiple bio-hazardous agents is presented. SpectroSens™ optical microchip sensors contain multiple spatially-separated waveguide ...channels with integral high-precision Bragg gratings sensitive to changes in refractive-index; selective surface-functionalisation of discrete sensing channels with different antibodies as bio-recognition elements enables selective multi-analyte biological detection. Interactions between target antigens in the test sample and respective surface-immobilised antibodies result in localised changes in refractive-index; the biosensor response manifests as increases in wavelength of light reflected from specific sensing channels. Multiplexed, label-free detection of 8 different biological agents, encompassing bacterial spores, vegetative cells, viruses and proteinaceous toxins has been demonstrated in real-time. Selective detection of
Bacillus atrophaeus (BG) spores,
Escherichia coli cells, MS2 viruses and ovalbumin (OVA) protein (simulant bio-hazardous agents) was first demonstrated as proof-of-concept; subsequently, detection of
Bacillus anthracis (BA) spores (UM23CL2 strain),
Franciscella tularensis (FT) cells (live vaccine strain), Vaccinia viruses (heat-killed) and ricin toxin (bio-hazardous agents) was proven. Two optical microchip sensors, each comprising 8 sensing channels were packaged into a single disposable cartridge allowing simultaneous 16-channel data acquisition. The specific antibody deposition sequence used in this study enabled detection of either 4 simulants or 4 bio-hazardous agents using a single consumable. The final device, a culmination of the multidisciplinary convergence of the fields of biology, chemistry, optoelectronics and microfluidics, is man-portable and inherently robust. The performance characteristics of the SpectroSens™ technology platform highlight its potential for exploitation as a ‘detect to warn/treat’ biodetector in security and defence operations.
Aim
To assess the association between allopurinol and mortality and cardiovascular outcomes in an allopurinol‐treated diabetes cohort.
Materials and Methods
We conducted a population‐based ...retrospective cohort study in Ontario, Canada. Eligible subjects were ≥ 66 years old with diabetes and a first prescription for allopurinol between 1 April, 2002 and 31 March, 2012 and were followed until 31 March, 2016. The primary outcome was a composite: all‐cause mortality, non‐fatal cardiovascular event (myocardial infarction, revascularization procedure, or stroke) or congestive heart failure (CHF). Secondary outcomes were components of the primary outcome and pneumonia as a negative tracer. Allopurinol was modelled as time‐varying exposed versus unexposed, daily dose category and cumulative dose using sex‐specific multivariable Cox proportional hazards models.
Results
Over a median follow‐up of 4.65 years (interquartile range 1.79–7.81), 16 266/23 103 males and 10 571/15 313 females experienced the primary outcome. Allopurinol was associated with a reduction in the primary outcome adjusted hazard ratios (aHR) 0.77 (95% confidence interval 0.75–0.80) and 0.81 (0.78–0.84) for males and females, respectively, driven by marked reductions in all‐cause mortality and modest reductions in cardiovascular events/CHF. There was no effect of cumulative allopurinol dose on any outcome, and allopurinol was also associated with reduced risk of pneumonia in males aHR 0.88 (0.83, 0.93).
Conclusions
Allopurinol was associated with reduced mortality and cardiovascular outcomes. However, lack of cumulative dose effect and a positive tracer outcome in males suggests residual bias. Future research assessing whether allopurinol prevents vascular complications in diabetes requires a clinical trial.
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