MiRNAs are small non-coding RNAs that interact with their target mRNAs for posttranscriptional gene regulation. Finely controlled miRNA biogenesis, target recognition and degradation indicate that ...maintaining miRNA homeostasis is essential for regulating cell proliferation, growth, differentiation and apoptosis. Increasingly, miRNAs have been recognized as a potential biomarker for disease diagnosis. MiRNAs can be found in blood, plasma, and tissues, and miRNA expression and activity differ in developmental stages, tissues and in response to external stimuli. MiRNA transcripts are matured from pri-miRNA over pre-miRNA to mature miRNA, a process that includes multiple steps and enzymes. Many tools are available to identify and quantify specific miRNAs, ranging from measuring total miRNA, specific miRNA activity, miRNA arrays and miRNA localization. The various miRNA assays differ in accuracy, cost, efficiency and convenience of monitoring miRNA dynamics. To acknowledge the significance and increasing research interest in miRNAs, we summarize the traditional as well as novel methods of miRNA quantification with strengths and limitations of various techniques in biochemical and medical research.
•DNA oxidation plays the prominent role in the early stages of Parkinson’s disease.•RNA oxidation CSF markers are supportive indicators of the neurodegeneration.•Disease duration and age influence ...the RNA/DNA oxidation in Parkinson’s disease.
8-hydroxy-2 deoxyguanosine (8-OHdG) and the 8-hydroxyguanosine (8-OHG) are the most widely used biomarkers of nucleoside oxidation affecting DNA and RNA and are considered reliable markers of oxidative stress. Increased levels of these markers are found in the various biological fluids of patients with neurodegenerative disorders.
The primary aim of our study was to assess the differences of investigated markers between patient groups and subsequently study the influence of clinical factors that might modify the levels of investigated markers during the disease progression.
In this study, we analysed the 8-OHdG and 8-OHG levels in the cerebrospinal fluid (CSF) and serum from 44 patients with Parkinson’s disease (PD) and 32 controls using an ELISA.
There were significantly higher CSF levels of both investigated markers in Parkinson’s disease patients as compared to controls (p=0.02 and p=0.04). Significantly higher CSF values of 8-OHdG were found in PD patients without dementia (p=0.05), whereas patients with dementia recorded lower 8-OHG CSF levels compared to controls (p=0.04). The disease duration and age influenced the levels of both markers within investigated groups.
Oxidative DNA damage plays an important role in the early stages of PD, whereas during the progression of the disease the process is more complex, and other mechanisms are in the foreground. The measurement of 8-OHdG might be used as an “early-stage marker”, whereas the decrease of 8-OHG in CSF might reflect the degree of neurodegeneration during the disease progression, suggesting its utility as a prognostic marker of advanced PD stages.
Abstract Recent studies showed that spreading depolarizations (SDs) occurs abundantly in patients following ischemic stroke and experimental evidence suggests that SDs recruit tissue at risk into ...necrosis. We hypothesized that BBB opening with consequent alterations of the extracellular electrolyte composition and extravasation of albumin facilitates generation of SDs since albumin mediates an astrocyte transcriptional response with consequent disturbance of potassium and glutamate homeostasis. Here we show extravasation of Evans blue–albumin complex into the hippocampus following cortical photothrombotic stroke in the neighboring neocortex. Using extracellular field potential recordings and exposure to serum electrolytes we observed spontaneous SDs in 80% of hippocampal slices obtained from rats 24 h after cortical photothrombosis. Hippocampal exposure to albumin for 24 h through intraventricular application together with serum electrolytes lowered the threshold for the induction of SDs in most slices irrespective of the pathway of stimulation. Exposing acute slices from naive animals to albumin led also to a reduced SD threshold. In albumin-exposed slices the onset of SDs was usually associated with larger stimulus-induced accumulation of extracellular potassium, and preceded by epileptiform activity, which was also observed during the recovery phase of SDs. Application of ifenprodil (3 μM), an NMDA-receptor type 2 B antagonist, blocked stimulus dependent epileptiform discharges and generation of SDs in slices from animals treated with albumin in-vivo . We suggest that BBB opening facilitates the induction of peri-infarct SDs through impaired homeostasis of K+.
Currently available antiepileptic drugs (AEDs) were developed to suppress seizure activity but less for prevention of epileptogenesis or for treatment of epileptogenic encephalopathies. Despite ...considerable efforts towards pharmacological control of seizures, about 30 % of epileptic patients do not achieve complete seizure control, and these numbers are even higher in patients suffering from partial seizures - a common form of epilepsy in adults. The mechanisms behind drug-resistance are far from being understood. Likely several unrelated mechanisms might lead in concert to reduced efficacy of the AEDs. Consequently, there is a need for predictive biomarkers of susceptibility to pharmacoresistant seizures and for new therapies interfering with epileptogenesis and preventing development of drug-resistance instead of merely suppressing seizures. This also necessitates the design of novel in vitro and in vivo epilepsy models that would better mimic the progressive nature of epilepsy and resemble the state of a chronic epileptic tissue. In this review we discuss current theories of drug-resistance and give a short summary of the epilepsy models that are frequently used for testing AEDs. We will also highlight caveats of the different models and consider novel approaches to overcome these difficulties. Finally we give a short outlook on unconventional therapies interfering with epileptogenesis as well as with drug delivery and retention.
Several molecular subtypes of sporadic Creutzfeldt–Jakob disease have been identified and electroencephalogram and cerebrospinal fluid biomarkers have been reported to support clinical diagnosis but ...with variable utility according to subtype. In recent years, a series of publications have demonstrated a potentially important role for magnetic resonance imaging in the pre-mortem diagnosis of sporadic Creutzfeldt–Jakob disease. Magnetic resonance imaging signal alterations correlate with distinct sporadic Creutzfeldt–Jakob disease molecular subtypes and thus might contribute to the earlier identification of the whole spectrum of sporadic Creutzfeldt–Jakob disease cases. This multi-centre international study aimed to provide a rationale for the amendment of the clinical diagnostic criteria for sporadic Creutzfeldt–Jakob disease. Patients with sporadic Creutzfeldt–Jakob disease and fluid attenuated inversion recovery or diffusion-weight imaging were recruited from 12 countries. Patients referred as ‘suspected sporadic Creutzfeldt–Jakob disease’ but with an alternative diagnosis after thorough follow up, were analysed as controls. All magnetic resonance imaging scans were assessed for signal changes according to a standard protocol encompassing seven cortical regions, basal ganglia, thalamus and cerebellum. Magnetic resonance imaging scans were evaluated in 436 sporadic Creutzfeldt–Jakob disease patients and 141 controls. The pattern of high signal intensity with the best sensitivity and specificity in the differential diagnosis of sporadic Creutzfeldt–Jakob disease was identified. The optimum diagnostic accuracy in the differential diagnosis of rapid progressive dementia was obtained when either at least two cortical regions (temporal, parietal or occipital) or both caudate nucleus and putamen displayed a high signal in fluid attenuated inversion recovery or diffusion-weight imaging magnetic resonance imaging. Based on our analyses, magnetic resonance imaging was positive in 83% of cases. In all definite cases, the amended criteria would cover the vast majority of suspected cases, being positive in 98%. Cerebral cortical signal increase and high signal in caudate nucleus and putamen on fluid attenuated inversion recovery or diffusion-weight imaging magnetic resonance imaging are useful in the diagnosis of sporadic Creutzfeldt–Jakob disease. We propose an amendment to the clinical diagnostic criteria for sporadic Creutzfeldt–Jakob disease to include findings from magnetic resonance imaging scans.
High-fidelity protein synthesis requires properly aminoacylated transfer RNAs (tRNAs), yet diverse cell types, from bacteria to humans, show a surprising ability to tolerate errors in translation ...resulting from mutations in tRNAs, aminoacyl-tRNA synthetases, and other components of protein synthesis. Recently, we characterized a tRNA
G35A mutant (tRNA
) that occurs in 2% of the human population. The mutant tRNA decodes phenylalanine codons with serine, inhibits protein synthesis, and is defective in protein and aggregate degradation. Here, we used cell culture models to test our hypothesis that tRNA-dependent mistranslation will exacerbate toxicity caused by amyotrophic lateral sclerosis (ALS)-associated protein aggregation. Relative to wild-type tRNA, we found cells expressing tRNA
showed slower but effective aggregation of the fused in sarcoma (FUS) protein. Despite reduced levels in mistranslating cells, wild-type FUS aggregates showed similar toxicity in mistranslating cells and normal cells. The aggregation kinetics of the ALS-causative FUS R521C variant were distinct and more toxic in mistranslating cells, where rapid FUS aggregation caused cells to rupture. We observed synthetic toxicity in neuroblastoma cells co-expressing the mistranslating tRNA mutant and the ALS-causative FUS R521C variant. Our data demonstrate that a naturally occurring human tRNA variant enhances cellular toxicity associated with a known causative allele for neurodegenerative disease.
Delivery of Active AKT1 to Human Cells Siddika, Tarana; Balasuriya, Nileeka; Frederick, Mallory I ...
Cells,
11/2022, Letnik:
11, Številka:
23
Journal Article
Recenzirano
Odprti dostop
Protein kinase B (AKT1) is a serine/threonine kinase and central transducer of cell survival pathways. Typical approaches to study AKT1 biology in cells rely on growth factor or insulin stimulation ...that activates AKT1 via phosphorylation at two key regulatory sites (Thr308, Ser473), yet cell stimulation also activates many other kinases. To produce cells with specific AKT1 activity, we developed a novel system to deliver active AKT1 to human cells. We recently established a method to produce AKT1 phospho-variants from
with programmed phosphorylation. Here, we fused AKT1 with an N-terminal cell penetrating peptide tag derived from the human immunodeficiency virus trans-activator of transcription (TAT) protein. The TAT-tag did not alter AKT1 kinase activity and was necessary and sufficient to rapidly deliver AKT1 protein variants that persisted in human cells for 24 h without the need to use transfection reagents. TAT-pAKT1
induced selective phosphorylation of the known AKT1 substrate GSK-3α, but not GSK-3β, and downstream stimulation of the AKT1 pathway as evidenced by phosphorylation of ribosomal protein S6 at Ser240/244. The data demonstrate efficient delivery of AKT1 with programmed phosphorylation to human cells, thus establishing a cell-based model system to investigate signaling that is dependent on AKT1 activity.
In porous materials, such as foams or pressed powder boards, the thermal conductivity via the gas phase represents at ambient temperatures a significant or even dominant contribution to the total ...thermal conductivity. The variation of the thermal conductivity of an open porous material with gas pressure is a function of the pore size. In the pressure range below 1
bar, measurement of thermal conductivity offers a non-destructive probe of pore sizes larger than 100
nm. If the thermal conductivity setup also allows for measurements at higher pressures, the pore size range can be extended to even smaller values.
Experimental data for a variety of different porous materials such as open porous foams, granular porous and nonporous materials and aerogels, porous solids with an adjustable mean pore size in the meso- or macro-pore range, are presented. The pore characteristics determined from thermal conductivity measurements are compared to morphological data derived from nitrogen sorption and small angle X-ray scattering data.
The potential and limits of this method in terms of the extraction of an average pore size and a pore size distribution are discussed.
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