MicroRNA profiling reveals marker of motor neuron disease in ALS models

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder marked by the loss of motor neurons (MNs) in the brain and spinal cord, leading to fatally debilitating weakness. Because this disease predominantly affects MNs, we aimed to characterize the distinct expression profile of that cell type to elucidate underlying disease mechanisms and to identify novel targets that inform on MN health during ALS disease time course. microRNAs (miRNAs) are short, noncoding RNAs that can shape the expression profile of a cell and thus often exhibit cell-type-enriched expression. To determine MN-enriched miRNA expression, we used Cre recombinase-dependent miRNA tagging and affinity purification in mice. By defining thein vivomiRNA expression of MNs, all neurons, astrocytes, and microglia, we then focused on MN-enriched miRNAs via a comparative analysis and found that they may functionally distinguish MNs postnatally from other spinal neurons. Characterizing the levels of the MN-enriched miRNAs in CSF harvested from ALS models of MN disease demonstrated that one miRNA (miR-218) tracked with MN loss and was responsive to an ALS therapy in rodent models. Therefore, we have used cellular expression profiling tools to define the distinct miRNA expression of MNs, which is likely to enrich future studies of MN disease. This approach enabled the development of a novel, drug-responsive marker of MN disease in ALS rodents.SIGNIFICANCE STATEMENTAmyotrophic lateral sclerosis (ALS) is a neurodegenerative disease in which motor neurons (MNs) in the brain and spinal cord are selectively lost. To develop tools to aid in our understanding of the distinct expression profiles of MNs and, ultimately, to monitor MN disease progression, we identified small regulatory microRNAs (miRNAs) that were highly enriched or exclusive in MNs. The signal for one of these MN-enriched miRNAs is detectable in spinal tap biofluid from an ALS rat model, where its levels change as disease progresses, suggesting that it may be a clinically useful marker of disease status. Furthermore, rats treated with ALS therapy have restored expression of this MN RNA marker, making it an MN-specific and drug-responsive marker for ALS rodents.</jats:p

1000 Norms Project: Protocol of a cross-sectional study cataloging human variation

Background Clinical decision-making regarding diagnosis and management largely depends on comparison with healthy or ‘normal’ values. Physiotherapists and researchers therefore need access to robust patient-centred outcome measures and appropriate reference values. However there is a lack of high-quality reference data for many clinical measures. The aim of the 1000 Norms Project is to generate a freely accessible database of musculoskeletal and neurological reference values representative of the healthy population across the lifespan. Methods/design In 2012 the 1000 Norms Project Consortium defined the concept of ‘normal’, established a sampling strategy and selected measures based on clinical significance, psychometric properties and the need for reference data. Musculoskeletal and neurological items tapping the constructs of dexterity, balance, ambulation, joint range of motion, strength and power, endurance and motor planning will be collected in this cross-sectional study. Standardised questionnaires will evaluate quality of life, physical activity, and musculoskeletal health. Saliva DNA will be analysed for the ACTN3 genotype (‘gene for speed’). A volunteer cohort of 1000 participants aged 3 to 100 years will be recruited according to a set of self-reported health criteria. Descriptive statistics will be generated, creating tables of mean values and standard deviations stratified for age and gender. Quantile regression equations will be used to generate age charts and age-specific centile values. Discussion This project will be a powerful resource to assist physiotherapists and clinicians across all areas of healthcare to diagnose pathology, track disease progression and evaluate treatment response. This reference dataset will also contribute to the development of robust patient-centred clinical trial outcome measures

25th annual computational neuroscience meeting: CNS-2016

The same neuron may play different functional roles in the neural circuits to which it belongs. For example, neurons in the Tritonia pedal ganglia may participate in variable phases of the swim motor rhythms [1]. While such neuronal functional variability is likely to play a major role the delivery of the functionality of neural systems, it is difficult to study it in most nervous systems. We work on the pyloric rhythm network of the crustacean stomatogastric ganglion (STG) [2]. Typically network models of the STG treat neurons of the same functional type as a single model neuron (e.g. PD neurons), assuming the same conductance parameters for these neurons and implying their synchronous firing [3, 4]. However, simultaneous recording of PD neurons shows differences between the timings of spikes of these neurons. This may indicate functional variability of these neurons. Here we modelled separately the two PD neurons of the STG in a multi-neuron model of the pyloric network. Our neuron models comply with known correlations between conductance parameters of ionic currents. Our results reproduce the experimental finding of increasing spike time distance between spikes originating from the two model PD neurons during their synchronised burst phase. The PD neuron with the larger calcium conductance generates its spikes before the other PD neuron. Larger potassium conductance values in the follower neuron imply longer delays between spikes, see Fig. 17.Neuromodulators change the conductance parameters of neurons and maintain the ratios of these parameters [5]. Our results show that such changes may shift the individual contribution of two PD neurons to the PD-phase of the pyloric rhythm altering their functionality within this rhythm. Our work paves the way towards an accessible experimental and computational framework for the analysis of the mechanisms and impact of functional variability of neurons within the neural circuits to which they belong

Rational Design of a 310-Helical PIP-Box Mimetic Targeting PCNA, the Human Sliding Clamp

The human sliding clamp (PCNA) controls access to DNA for many proteins involved in DNA replication and repair. Proteins are recruited to the PCNA surface by means of a short, conserved peptide motif known as the PCNA‐interacting protein box (PIP‐box). Inhibitors of these essential protein‐protein interactions may be useful as cancer therapeutics by disrupting DNA replication and repair in these highly proliferative cells. PIP‐box peptide mimetics have been identified as a potentially rapid route to potent PCNA inhibitors. Here we describe the rational design and synthesis of the first PCNA peptidomimetic ligands, based on the high affinity PIP‐box sequence from the natural PCNA inhibitor p21. These mimetics incorporate covalent i,i+4 side‐chain/side‐chain lactam linkages of different lengths, designed to constrain the peptides into the 310‐helical structure required for PCNA binding. NMR studies confirmed that while the unmodified p21 peptide had little defined structure in solution, mimetic ACR2 pre‐organized into 310‐helical structure prior to interaction with PCNA. ACR2 displayed higher affinity binding than most known PIP‐box peptides, and retains the native PCNA binding mode, as observed in the co‐crystal structure of ACR2 bound to PCNA. This study offers a promising new strategy for PCNA inhibitor design for use as anti‐cancer therapeutics

Rational Design of a 310-Helical PIP-Box Mimetic Targeting PCNA, the Human Sliding Clamp

The human sliding clamp (PCNA) controls access to DNA for many proteins involved in DNA replication and repair. Proteins are recruited to the PCNA surface by means of a short, conserved peptide motif known as the PCNA‐interacting protein box (PIP‐box). Inhibitors of these essential protein‐protein interactions may be useful as cancer therapeutics by disrupting DNA replication and repair in these highly proliferative cells. PIP‐box peptide mimetics have been identified as a potentially rapid route to potent PCNA inhibitors. Here we describe the rational design and synthesis of the first PCNA peptidomimetic ligands, based on the high affinity PIP‐box sequence from the natural PCNA inhibitor p21. These mimetics incorporate covalent i,i+4 side‐chain/side‐chain lactam linkages of different lengths, designed to constrain the peptides into the 310‐helical structure required for PCNA binding. NMR studies confirmed that while the unmodified p21 peptide had little defined structure in solution, mimetic ACR2 pre‐organized into 310‐helical structure prior to interaction with PCNA. ACR2 displayed higher affinity binding than most known PIP‐box peptides, and retains the native PCNA binding mode, as observed in the co‐crystal structure of ACR2 bound to PCNA. This study offers a promising new strategy for PCNA inhibitor design for use as anti‐cancer therapeutics