Modelling the Delta1/Notch1 pathway: in search of the mediator(s) of neural stem cell differentiation

The Notch1 signalling pathway has been shown to control neural stem cell fate through lateral inhibition of mash1, a key promoter of neuronal differentiation. Interaction between the Delta1 ligand of a differentiating cell and the Notch1 protein of a neighbouring cell results in cleavage of the trans-membrane protein, releasing the intracellular domain (NICD) leading to the up regulation of hes1. Hes1 homodimerisation leads to down regulation of mash1. Most mathematical models currently represent this pathway up to the formation of the HES1 dimer. Herein, we present a detailed model ranging from the cleavage of the NICD and how this signal propagates through the Delta1/Notch1 pathway to repress the expression of the proneural genes. Consistent with the current literature, we assume that cells at the self renewal state are represented by a stable limit cycle and through in silico experimentation we conclude that a drastic change in the main pathway is required in order for the transition from self-renewal to differentiation to take place. Specifically, a model analysis based approach is utilised in order to generate hypotheses regarding potential mediators of this change. Through this process of model based hypotheses generation and testing, the degradation rates of Hes1 and Mash1 mRNA and the dissociation constant of Mash1-E47 heterodimers are identified as the most potent mediators of the transition towards neural differentiation.     

Mitotic catastrophe occurs in the absence of apoptosis in p53-null cells with a defective G1 checkpoint

Cell death occurring during mitosis, or mitotic catastrophe, often takes place in conjunction with apoptosis, but the conditions in which mitotic catastrophe may exhibit features of programmed cell death are still unclear. In the work presented here, we studied mitotic cell death by making use of a UV-inactivated parvovirus (adeno-associated virus; AAV) that has been shown to induce a DNA damage response and subsequent death of p53-defective cells in mitosis, without affecting the integrity of the host genome. Osteosarcoma cells (U2OSp53DD) that are deficient in p53 and lack the G1 cell cycle checkpoint respond to AAV infection through a transient G2 arrest. We found that the infected U2OSp53DD cells died through mitotic catastrophe with no signs of chromosome condensation or DNA fragmentation. Moreover, cell death was independent of caspases, apoptosis-inducing factor (AIF), autophagy and necroptosis. These findings were confirmed by time-lapse microscopy of cellular morphology following AAV infection. The assays used readily revealed apoptosis in other cell types when it was indeed occurring. Taken together the results indicate that in the absence of the G1 checkpoint, mitotic catastrophe occurs in these p53-null cells predominantly as a result of mechanical disruption induced by centrosome overduplication, and not as a consequence of a suicide signal.     

Isokinetic knee joint evaluation in track and field events

The purpose of this study was to evaluate maximal torque of the knee flexors and extensors, flexor/extensor ratios, and maximal torque differences between the 2 lower extremities in young track and field athletes. Forty male track and field athletes 13-17 years old and 20 male nonathletes of the same age participated in the study. Athletes were divided into 4 groups according to their age and event (12 runners and 10 jumpers 13-15 years old, 12 runners and 6 jumpers 16-17 years old) and nonathletes into 2 groups of the same age. Maximal torque evaluation of knee flexors and extensors was performed on an isokinetic dynamometer at 60°·s(-1). At the age of 16-17 years, jumpers exhibited higher strength values at extension than did runners and nonathletes, whereas at the age of 13-15 years, no significant differences were found between events. Younger athletes were weaker than older athletes at flexion. Runners and jumpers were stronger than nonathletes in all relative peak torque parameters. Nonathletes exhibited a higher flexor/extensor ratio compared with runners and jumpers. Strength imbalance in athletes was found between the 2 lower extremities in knee flexors and extensors and also at flexor/extensor ratio of the same extremity. Young track and field athletes exhibit strength imbalances that could reduce their athletic performance, and specific strength training for the weak extremity may be needed.     

Incidence and Prevalence of Major Central Nervous System Involvement in Systemic Lupus Erythematosus: A 3-Year Prospective Study of 370 Patients

Background

The incidence and prevalence of CNS involvement in SLE remains unclear owing to conflicting results in the published studies. The aim of the study was to evaluate the incidence and prevalence of major definite CNS events in SLE patients.

Methods

370 SLE patients with no previous history of CNS involvement were prospectively evaluated in a tertiary hospital referral center for 3 years. Major CNS manifestations were codified according to ACR definitions, including chorea, aseptic meningitis, psychosis, seizures, myelopathy, demyelinating syndrome, acute confusional state and strokes. Minor CNS events were excluded. ECLAM and SLEDAI-SELENA Modification scores were used to evaluate disease activity and SLICC/ACR Damage Index was used to assess accumulated damage.

Results

16/370 (4.3%) patients presented with a total of 23 major CNS events. These included seizures (35%), strokes (26%), myelopathy (22%), optic neuritis (8.7%), aseptic meningitis (4.3%) and acute psychosis (4.3%). Incidence was 7.8/100 person years. Among hospitalizations for SLE, 13% were due to CNS manifestations. Epileptic seizures were associated with high disease activity, while myelopathy correlated with lower disease activity and NMO-IgG antibodies (P≤0.05). Stroke incidence correlated with APS coexistence (P = 0.06). Overall, CNS involvement correlated with high ECLAM and SLEDAI scores (P<0.001).

Conclusions

Clinically severe CNS involvement is rare in SLE patients, accounting for 7.8/100 person years. CNS involvement correlates with high disease activity and coexistence of specific features that define the respective CNS syndromes.     

Passive smoking reduces and vitamin C increases exercise-induced oxidative stress: Does this make passive smoking an anti-oxidant and vitamin C a pro-oxidant stimulus?

The current interpretative framework states that, for a certain experimental treatment (usually a chemical substance) to be classified as “anti-oxidant”, it must possess the property of reducing (or even nullifying) exercise-induced oxidative stress. The aim of the study was to compare side by side, in the same experimental setup, redox biomarkers responses to an identical acute eccentric exercise session, before and after chronic passive smoking (considered a pro-oxidant stimulus) or vitamin C supplementation (considered an anti-oxidant stimulus). Twenty men were randomly assigned into either passive smoking or vitamin C group. All participants performed two acute eccentric exercise sessions, one before and one after either exposure to passive smoking or vitamin C supplementation for 12 days. Vitamin C, oxidant biomarkers (F₂-isoprostanes and protein carbonyls) and the non-enzymatic antioxidant (glutathione) were measured, before and after passive smoking, vitamin C supplementation or exercise. It was found that chronic exposure to passive smoking increased the level of F₂-isoprostanes and decreased the level of glutathione at rest, resulting in minimal increase or absence of oxidative stress after exercise. Conversely, chronic supplementation with vitamin C decreased the level of F₂-isoprostanes and increased the level of glutathione at rest, resulting in marked exercise-induced oxidative stress. Contrary to the current scientific consensus, our results show that, when a pro-oxidant stimulus is chronically delivered, it is more likely that oxidative stress induced by subsequent exercise is decreased and not increased. Reversely, it is more likely to find greater exercise-induced oxidative stress after previous exposure to an anti-oxidant stimulus. We believe that the proposed framework will be a useful tool to reach more pragmatic explanations of redox biology phenomena.     

Towards analyzing the potential of exosomes to deliver microRNA therapeutics

Exosome selectivity mechanisms underlying exosome–target cell interactions and the specific traits affecting their capability to communicate still remain unclear. Moreover, the capacity of exosomes to efficiently deliver their molecular cargos intracellularly needs precise investigation towards establishing functional exosome‐based delivery platforms exploitable in the clinical practice. The current study focuses on: (a) exosome production from normal MRC‐5 and Vero cells growing in culture, (b) physicochemical characterization by dynamic light scattering (DLS) and cryo‐transmission electron microscopy; (c) cellular uptake studies of rhodamine‐labeled exosomes in normal and cancer cells, providing to exosomes either “autologous” or “heterologous” cellular delivery environments; and (d) loading exogenous Alexa Fluor 488‐labeled siRNA into exosomes for the assessment of their delivering capacity by immunofluorescence in a panel of recipient cells. The data obtained thus far indicate that MRC‐5 and Vero exosomes, indeed exhibit an interesting delivering profile, as promising “bio‐shuttles,” being pharmacologically exploitable in the context of theranostic applications.