Validation and optimization of the loop-mediated isothermal amplification (LAMP) technique for rapid detection of wheat stripe mosaic virus, a wheat-infecting pathogen.

BACKGROUND: Wheat stripe mosaic virus (WhSMV) is a significant wheat pathogen that causes substantial yield losses in Brazil and other countries. Although several detection methods are available, reliable and efficient tools for on-site WhSMV detection are currently lacking. In this study, a Loop-Mediated Isothermal Amplification (LAMP) method was developed for rapid and reliable field detection of WhSMV. We designed WhSMV-specific primers for the LAMP assay and optimized reaction conditions for increased sensitivity and specificity using infected plant samples. RESULTS: We have developed a diagnostic method utilizing the Loop-Mediated Isothermal Amplification (LAMP) technique capable of rapidly and reliably detecting WhSMV. The LAMP assay has been optimized to enhance sensitivity, specificity, and cost-effectiveness. CONCLUSION: The LAMP assay described here represents a valuable tool for early WhSMV detection, serving to mitigate the adverse economic and social impacts of this viral pathogen. By enabling swift and accurate identification, this assay can significantly improve the sustainability of cereal production systems, safeguarding crop yields against the detrimental effects of WhSMV.

Biofeedback interventions for short term upper limb function following stroke: A systematic review with meta-analysis.

BACKGROUND: Biofeedback has been used by rehabilitation professionals in the treatment of poststroke function impairments. PURPOSE: Investigate the efficacy of any type of biofeedback intervention for the treatment of upper limb function in individuals following stroke. STUDY DESIGN: Systematic review of literature with meta-analysis. METHODS: Literature searches were conducted using MESH terms and text words in PubMed, Lilacs, Scielo, Scopus, PEDro, and Web of Science databases. The main outcome was improvement in upper limb's motor function and motor function in activities of daily living. We calculated the Mean Difference and Standardized Mean Difference for the assessment scales reported as primary outcome. The methodological quality of included studies was assessed using PEDro scale. The overall quality of the evidence was assessed using GRADE system. RESULTS: From 1360 articles identified, 16 were included in the review (09 in the meta-analysis). Three forest plots of hemiparesis and one of hemiplegia showed that biofeedback therapy associated with conventional therapy has a greater improvement in participants upper limb motor function when compared to isolated conventional therapy. Two forest plots of hemiparesis and one of hemiplegia showed no superiority in participants improvement for biofeedback associated with conventional therapy when compared to isolated conventional therapy. CONCLUSION: Biofeedback therapy associated with conventional therapy showed a small clinical effect when associated to conventional therapy and very low quality of evidence. Although further research with higher quality evidence is needed.

Effectiveness of Circuit-Based Exercises on Gait Speed, Balance, and Functional Mobility in People Affected by Stroke: A Meta-Analysis.

BACKGROUND: Several interventions have been proposed to rehabilitate patients with neurologic dysfunctions due to stroke. However, the effectiveness of circuit-based exercises according to its actual definition, ie, an overall program to improve strength, stamina, balance or functioning, was not provided. OBJECTIVE: To examine the effectiveness of circuit-based exercise in the treatment of people affected by stroke. METHODS: A search through PubMed, Embase, Cochrane Library, and Physiotherapy Evidence Database databases was performed to identify controlled clinical trials without language or date restriction. The overall mean difference with 95% confidence interval was calculated for all outcomes. Two independent reviewers assessed the risk of bias. RESULTS: Eleven studies met the inclusion criteria, and 8 presented suitable data to perform a meta-analysis. Quantitative analysis showed that circuit-based exercise was more effective than conventional intervention on gait speed (mean difference of 0.11 m/s) and circuit-based exercise was not significantly more effective than conventional intervention on balance and functional mobility. CONCLUSION: Our results demonstrated that circuit-based exercise presents better effects on gait when compared with conventional intervention and that its effects on balance and functional mobility were not better than conventional interventions. LEVEL OF EVIDENCE: I.

PERK activation mitigates tau pathology in vitro and in vivo.

The RNA-like endoplasmic reticulum kinase (PERK) is genetically associated with the tauopathy progressive supranuclear palsy (PSP). To elucidate the functional mechanisms underlying this association, we explored PERK activity in brains of PSP patients and its function in three tauopathy models (cultured human neurons overexpressing 4-repeat wild-type tau or treated with the environmental neurotoxin annonacin, and P301S tau transgenic mice). In vitro, treatment with a pharmacological PERK activator CCT020312 or PERK overexpression reduced tau phosphorylation, tau conformational change and 4-repeat tau isoforms, and increased cell viability. In vivo, the PERK activator significantly improved memory and locomotor function, reduced tau pathology, and prevented dendritic spine and motoneuron loss in P301S tau mice. Importantly, the PERK substrate EIF2A, mediating some detrimental effects of PERK signaling, was downregulated in PSP brains and tauopathy models, suggesting that the alternative PERK-NRF2 pathway accounts for these beneficial effects in the context of tauopathies. In summary, PERK activation may be a novel strategy to treat PSP and eventually other tauopathies.

Systemically administered neuregulin-1ß1 rescues nigral dopaminergic neurons via the ErbB4 receptor tyrosine kinase in MPTP mouse models of Parkinson's disease.

Previously, we demonstrated that systemically injected extracellular domain of neuregulin-1ß1 (Nrg1ß1), a nerve growth and differentiation factor, passes the blood-brain barrier and rescues dopaminergic neurons of substantia nigra in the 6-hydroxydopamine-mouse model of Parkinson's disease (PD). Here, we studied the effects of peripherally administered Nrg1ß1 in another toxin-based mouse model of PD. For this purpose, (i) nigrostriatal pathway injury was induced by treatment of adult wild-type mice with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in acute and subchronic paradigms; and (ii) Nrg1ß1 or saline (control) were administered 1 h before each MPTP injection. We found that Nrg1ß1 significantly reduced the loss of nigral dopaminergic neurons in both intoxication paradigms (7 days post-injection). However, Nrg1ß1 did not reverse MPTP-induced decrease in dopamine levels and dopaminergic fibers in the striatum. We also show that MPTP conversion to its toxic metabolite 1-methyl-4-phenylpyridinium as well as levels of dopamine transporter, mediating intracellular uptake of 1-methyl-4-phenylpyridinium, are unaffected by Nrg1ß1. Finally, neuroprotective properties of Nrg1ß1 on nigral dopaminergic neurons are specifically mediated by ErbB4 as revealed through the study of ErbB4 knockout mice. In conclusion, systemically administered Nrg1ß1 protects midbrain dopaminergic neurons against this PD-related toxic insult. Thus, Nrg1ß1 may have a benefit in the treatment of PD patients. Previously, we demonstrated that systemically administered neuregulin-1ß1 (Nrg1ß1) passes the blood-brain barrier, phosphorylates ErbB4 receptors and elevates dopamine (DA) levels in the nigrostriatal system of healthy mice. Nrg1ß1 protects nigral DAergic neurons in the 6-hydroxydopamine (6-OHDA) mouse model of Parkinson's disease (PD). Here, we show that Nrg1ß1 rescues nigral DAergic neurons also against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced cell death. ErbB4 expression is essential for the neuroprotective effect of Nrg1ß1 on midbrain DAergic neurons. Nrg1ß1 might be beneficial in PD treatment.

Piericidin A aggravates Tau pathology in P301S transgenic mice.

OBJECTIVE: The P301S mutation in exon 10 of the tau gene causes a hereditary tauopathy. While mitochondrial complex I inhibition has been linked to sporadic tauopathies. Piericidin A is a prototypical member of the group of the piericidins, a class of biologically active natural complex I inhibitors, isolated from streptomyces spp. with global distribution in marine and agricultural habitats. The aim of this study was to determine whether there is a pathogenic interaction of the environmental toxin piericidin A and the P301S mutation. METHODS: Transgenic mice expressing human tau with the P301S-mutation (P301S+/+) and wild-type mice at 12 weeks of age were treated subcutaneously with vehicle (N = 10 P301S+/+, N = 7 wild-type) or piericidin A (N = 9 P301S+/+, N = 9 wild-type mice) at a dose of 0.5 mg/kg/d for a period of 28 days via osmotic minipumps. Tau pathology was measured by stereological counts of cells immunoreative with antibodies against phosphorylated tau (AD2, AT8, AT180, and AT100) and corresponding Western blot analysis. RESULTS: Piericidin A significantly increased the number of phospho-tau immunoreactive cells in the cerebral cortex in P301S+/+ mice, but only to a variable and mild extent in wild-type mice. Furthermore, piericidin A led to increased levels of pathologically phosphorylated tau only in P301S+/+ mice. While we observed no apparent cell loss in the frontal cortex, the synaptic density was reduced by piericidin A treatment in P301S+/+ mice. DISCUSSION: This study shows that exposure to piericidin A aggravates the course of genetically determined tau pathology, providing experimental support for the concept of gene-environment interaction in the etiology of tauopathies.

Mitochondrial complex 1 inhibition increases 4-repeat isoform tau by SRSF2 upregulation.

Progressive Supranuclear Palsy (PSP) is a neurodegenerative disorder characterised by intracellular aggregation of the microtubule-associated protein tau. The tau protein exists in 6 predominant isoforms. Depending on alternative splicing of exon 10, three of these isoforms have four microtubule-binding repeat domains (4R), whilst the others only have three (3R). In PSP there is an excess of the 4R tau isoforms, which are thought to contribute significantly to the pathological process. The cause of this 4R increase is so far unknown. Several lines of evidence link mitochondrial complex I inhibition to the pathogenesis of PSP. We demonstrate here for the first time that annonacin and MPP(+), two prototypical mitochondrial complex I inhibitors, increase the 4R isoforms of tau in human neurons. We show that the splicing factor SRSF2 is necessary to increase 4R tau with complex I inhibition. We also found SRSF2, as well as another tau splicing factor, TRA2B, to be increased in brains of PSP patients. Thereby, we provide new evidence that mitochondrial complex I inhibition may contribute as an upstream event to the pathogenesis of PSP and suggest that splicing factors may represent an attractive therapeutic target to intervene in the disease process.

Memory deficits correlate with tau and spine pathology in P301S MAPT transgenic mice.

AIM: P301S MAPT transgenic mice (P301S mice) are a widely used model of frontotemporal dementia and parkinsonism linked to chromosome 17 with tau pathology (FTDP-17-tau). However, a systematic correlation between cognitive deficits and cellular tau pathology at different ages is still missing. Therefore, our study investigated memory deficits of P301S mice in relation to pathological tau species and dendritic spine pathology throughout adulthood. METHODS: We analysed P301S mice behaviourally with the novel open field, rotarod, and Morris water maze tests to measure deficits in locomotion, balance and cognition, respectively; immunohistochemically with different tau antibodies for specific tau species; and with Golgi staining for dendritic spine pathology. RESULTS: We confirmed the occurrence of locomotor deficits at an age of 5 months and newly report memory deficits from 2.5 months of age onwards. At this early age, MC1 and CP13, but not AT180 immunoreactivity, was prominent in the hippocampus of P301S mice. Neuronal cell loss in the hippocampus of P301S mice was not observed to occur till 6 months of age. However, there was a significant reduction in the density of dendritic spines from young adulthood onwards in hippocampal pyramidal neurones. CONCLUSION: In P301S mice, memory deficits precede the onset of locomotor dysfunction and coincide with the appearance of conformationally changed, S202-phosphorylated tau and reduced spine density in the absence of neuronal cell loss in the hippocampus. Our finding provides insights into the toxic effects of different tau species in vivo and may facilitate the development of new therapies against neurodegenerative tauopathies.

Trifluoperazine rescues human dopaminergic cells from wild-type α-synuclein-induced toxicity.

Parkinson's disease (PD) is the most frequent neurodegenerative movement disorder. Presently, there is no causal therapy available to slow down or halt disease progression. The presynaptic protein alpha-synuclein aggregates to form intraneuronal Lewy bodies in PD. It is generally believed that intermediates on the way from monomers to the large aggregates would mediate neurotoxicity, but the precise species and mechanism responsible for neuronal death are controversially debated. To study alpha-synuclein-mediated toxicity, we developed a new model in which moderate overexpression of wild-type alpha-synuclein led to gradual death of human postmitotic dopaminergic neurons. In accordance with findings in postmortem PD brains, small oligomeric species occurred and the autophagic flux was impaired in our model. The phenothiazine neuroleptic trifluoperazine, an activator of macroautophagy, selectively reduced one particular alpha-synuclein species and rescued cells. Inversely, blocking of autophagy led to an accumulation of this oligomeric species and increased cell death. These data show that activation of autophagy is a promising approach to protect against alpha-synuclein pathology and likely acts by targeting one specific alpha-synuclein species.

Annonacin, a natural lipophilic mitochondrial complex I inhibitor, increases phosphorylation of tau in the brain of FTDP-17 transgenic mice.

Both genetic and environmental factors likely contribute to the neuropathology of tauopathies, but it remains unclear how specific genetic backgrounds affect the susceptibility towards environmental toxins. Mutations in the tau gene have been associated with familial tauopathies, while annonacin, a plant-derived mitochondrial inhibitor, has been implicated in an environmental form of tauopathy. We therefore determined whether there was a pathogenic synergy between annonacin exposure and the expression of the R406W-tau mutation in transgenic mice. We found that annonacin exposure caused an increase in the number of neurons with phosphorylated tau in the somatodendritic compartment in several brain areas in R406W(+/+) mice as opposed to mice that had only the endogenous mouse tau (R406W(-/-)). Western blot analysis demonstrated a concomitant increase in total tau protein without increase in tau mRNA, but reduced proteasomal proteolytic activity in R406W(+/+), but not R406W(-/-) mice, upon annonacin-treatment. Phosphorylated tau levels exceeded the increase in total tau protein, along with increased levels of different tau kinases, foremost a striking increase in the p25/p35 ratio, known to activate the tau kinase Cdk5. In summary, we observed a synergistic interaction between annonacin exposure and the presence of the R406W-tau mutation, which resulted in reduced degradation, increased phosphorylation and redistribution of neuronal tau.

Tau silencing by siRNA in the P301S mouse model of tauopathy.

Suppression of tau protein expression has been shown to improve behavioral deficits in mouse models of tauopathies, offering an attractive therapeutic approach. Experimentally this had been achieved by switching off the promoters controlling the transgenic human tau gene (MAPT), which is not possible in human patients. The aim of the present study was therefore to evaluate the effectiveness of small interfering RNAs (siRNAs) and their cerebral delivery to suppress human tau expression in vivo, which might be a therapeutic option for human tauopathies. We used primary cortical neurons of transgenic mice expressing P301S-mutated human tau and Lund human mesencephalic (LUHMES) cells to validate the suppressive effect of siRNA in vitro. For measuring the effect in vivo, we stereotactically injected siRNA into the brains of P301S mice to reveal the suppression of tau by immunochemistry (AT180, MC1, and CP13 antibodies). We found that the Accell™ SMART pool siRNA against MAPT can effectively suppress tau expression in vitro and in vivo without a specific delivery agent. The siRNA showed a moderate distribution in the hippocampus of mice after single injection. NeuN, GFAP, Iba-1, MHC II immunoreactivities and the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay showed neither signs of neurotoxicity or neuroinflammation nor apoptosis when MAPT siRNA is present in the hippocampus. Our data suggest that siRNA against MAPT can serve as a potential tool for gene therapy in tauopathies.