Regulation of DNA replication within the immunoglobulin heavy-chain locus during B cell commitment.

The temporal order of replication of mammalian chromosomes appears to be linked to their functional organization, but the process that establishes and modifies this order during cell differentiation remains largely unknown. Here, we studied how the replication of the Igh locus initiates, progresses, and terminates in bone marrow pro-B cells undergoing B cell commitment. We show that many aspects of DNA replication can be quantitatively explained by a mechanism involving the stochastic firing of origins (across the S phase and the Igh locus) and extensive variations in their firing rate (along the locus). The firing rate of origins shows a high degree of coordination across Igh domains that span tens to hundreds of kilobases, a phenomenon not observed in simple eukaryotes. Differences in domain sizes and firing rates determine the temporal order of replication. During B cell commitment, the expression of the B-cell-specific factor Pax5 sharply alters the temporal order of replication by modifying the rate of origin firing within various Igh domains (particularly those containing Pax5 binding sites). We propose that, within the Igh C(H)-3'RR domain, Pax5 is responsible for both establishing and maintaining high rates of origin firing, mostly by controlling events downstream of the assembly of pre-replication complexes.

Comparison of Three Antagonists of Hedgehog Pathway to Promote Skeletal Muscle Regeneration after High Dose Irradiation.

The current geopolitical context has brought the radiological nuclear risk to the forefront of concerns. High-dose localized radiation exposure leads to the development of a musculocutaneous radiation syndrome affecting the skin and subcutaneous muscles. Despite the implementation of a gold standard treatment based on an invasive surgical procedure coupled with autologous cell therapy, a muscular defect frequently persists. Targeting the modulation of the Hedgehog (Hh) signaling pathway appears to be a promising therapeutic approach. Activation of this pathway enhances cell survival and promotes proliferation after irradiation, while inhibition by Cyclopamine facilitates differentiation. In this study, we compared the effects of three antagonists of Hh, Cyclopamine (CA), Vismodegib (VDG) and Sonidegib (SDG) on differentiation. A stable cell line of murine myoblasts, C2C12, was exposed to X-ray radiation (5 Gy) and treated with CA, VDG or SDG. Analysis of proliferation, survival (apoptosis), morphology, myogenesis genes expression and proteins production were performed. According to the results, VDG does not have a significant impact on C2C12 cells. SDG increases the expression/production of differentiation markers to a similar extent as CA, while morphologically, SDG proves to be more effective than CA. To conclude, SDG can be used in the same way as CA but already has a marketing authorization with an indication against basal cell cancers, facilitating their use in vivo. This proof of concept demonstrates that SDG represents a promising alternative to CA to promotes differentiation of murine myoblasts. Future studies on isolated and cultured satellite cells and in vivo will test this proof of concept.

Assessment of remote ischemic conditioning delivery with optical sensor in acute ischemic stroke: Randomised clinical trial protocol.

BACKGROUND: Remote ischemic conditioning (RIC) is delivered by a blood pressure cuff over the limb, raising pressure 50 mmHg above the systolic blood pressure, to a maximum of 200 mmHg. The cuff is inflated for five minutes and then deflated for five minutes in a sequential ischemia-reperfusion cycle 4-5 times per session. Elevated pressure in the limb may be associated with discomfort and consequently reduced compliance. Continuous assessment of relative blood concentration and oxygenation with a tissue reflectance spectroscopy (a type of optical sensor device) placed over the forearm during the RIC sessions of the arm will allow us to observe the effect of inflation and deflation of the pressure cuff. We hypothesize, in patients with acute ischemic stroke (AIS) and small vessel disease, RIC delivered together with a tissue reflectance sensor will be feasible. METHODS: The study is a prospective, single-center, randomized control trial testing the feasibility of the device. Patients with AIS within 7 days from symptoms onset; who also have small vessel disease will be randomized 2:1 to intervention or sham control arms. All patients randomized to the intervention arm will receive 5 cycles of ischemia/reperfusion in the non-paralyzed upper limb with a tissue reflectance sensor and patients in the sham control arm will receive pressure by keeping the cuff pressure at 30 mmHg for 5 minutes. A total of 51 patients will be randomized, 17 in the sham control arm and 34 in the intervention arm. The primary outcome measure will be the feasibility of RIC delivered for 7 days or at the time of discharge. The secondary device-related outcome measures are fidelity of RIC delivery and the completion rate of intervention. The secondary clinical outcome includes a modified Rankin scale, recurrent stroke and cognitive assessment at 90 days. DISCUSSION: RIC delivery together with a tissue reflectance sensor will allow insight into the blood concentration and blood oxygenation changes in the skin. This will allow individualized delivery of the RIC and improve compliance. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT05408130, June 7, 2022.

Acoustic monitoring of laser-induced phase transitions in minerals: implication for Mars exploration with SuperCam.

The SuperCam instrument suite onboard the Mars 2020 Perseverance rover uses the laser-induced breakdown spectroscopy (LIBS) technique to determine the elemental composition of rocks and soils of the Mars surface. It is associated with a microphone to retrieve the physical properties of the ablated targets when listening to the laser-induced acoustic signal. In this study, we report the monitoring of laser-induced mineral phase transitions in acoustic data. Sound data recorded during the laser ablation of hematite, goethite and diamond showed a sharp increase of the acoustic signal amplitude over the first laser shots. Analyses of the laser-induced craters with Raman spectroscopy and scanning electron microscopy indicate that both hematite and goethite have been transformed into magnetite and that diamond has been transformed into amorphous-like carbon over the first laser shots. It is shown that these transitions are the root cause of the increase in acoustic signal, likely due to a change in target's physical properties as the material is transformed. These results give insights into the influence of the target's optical and thermal properties over the acoustic signal. But most importantly, in the context of the Mars surface exploration with SuperCam, as this behavior occurs only for specific phases, it demonstrates that the microphone data may help discriminating mineral phases whereas LIBS data only have limited capabilities.

Defects and DNA replication.

We introduce a rate-equation formalism to study DNA replication kinetics in the presence of defects resulting from DNA damage and find a crossover between two regimes: a normal regime, where the influence of defects is local, and an initiation-limited regime. In the latter, defects have a global impact on replication, whose progress is set by the rate at which origins of replication are activated, or initiated. Normal, healthy cells have defect densities in the normal regime. Our model can explain an observed correlation between interorigin separation and rate of DNA replication.

Computational methods to study kinetics of DNA replication.

New technologies such as DNA combing have led to the availability of large quantities of data that describe the state of DNA while undergoing replication in S phase. In this chapter, we describe methods used to extract various parameters of replication--fork velocity, origin initiation rate, fork density, numbers of potential and utilized origins--from such data. We first present a version of the technique that applies to "ideal" data. We then show how to deal with, a number of real-world complications, such as the asynchrony of starting times of a population of cells, the finite length of fragments used in the analysis, and the finite amount of DNA in a chromosome.

Nondriven polymer translocation through a nanopore: computational evidence that the escape and relaxation processes are coupled.

Most of the theoretical models describing the translocation of a polymer chain through a nanopore use the hypothesis that the polymer is always relaxed during the complete process. In other words, models generally assume that the characteristic relaxation time of the chain is small enough compared to the translocation time that nonequilibrium molecular conformations can be ignored. In this paper, we use molecular dynamics simulations to directly test this hypothesis by looking at the escape time of unbiased polymer chains starting with different initial conditions. We find that the translocation process is not quite in equilibrium for the systems studied, even though the translocation time tau is about 10 times larger than the relaxation time tau{r}. Our most striking result is the observation that the last half of the chain escapes in less than approximately 12% of the total escape time, which implies that there is a large acceleration of the chain at the end of its escape from the channel.

A Monte Carlo algorithm to study polymer translocation through nanopores. II. Scaling laws.

In the first paper of this series, we developed a new one-dimensional Monte Carlo approach for the study of flexible chains that are translocating through a small channel. We also presented a numerical scheme that can be used to obtain exact values for both the escape times and the escape probabilities given an initial pore-polymer configuration. We now present and discuss the fundamental scaling behaviors predicted by this Monte Carlo method. Our most important result is the fact that, in the presence of an external bias E, we observe a change in the scaling law for the translocation time tau as function of the polymer length N: In the general expression tau approximately N(beta)E, the exponent changes from beta=1 for moderately long chains to beta=1+nu or beta=2nu for very large values of N (for Rouse and Zimm dynamics, respectively). We also observe an increase in the effective diffusion coefficient due to the presence of entropic pulling on unbiased polymer chains.

Biased random walks on a lattice: exact numerical method to study the effect of alternating fields in disordered and asymmetric systems of obstacles.

The migration of a particle in a system of obstacles under the action of an external field is often modeled using lattice Monte Carlo algorithms. For example, such simulation methods have been used to study the electrophoresis of charged molecules in sieving gels and the separation of particles using ratchet systems. In the case of constant fields or low-frequency alternating fields, the Monte Carlo simulation method can be mapped onto a numerical or algebraic matrix problem that can be solved exactly. In this Rapid Communication, we generalize this matrix approach to treat periodic time-dependent fields. The evolution of the spatial distribution function during a period is computed using a sequence of transfer matrices, and a steady-state closure relation allows us to calculate the exact mean velocity of the particle during a complete cycle. As an example, we examine the properties of a simple spatially asymmetric ratchet system in the presence of periodic alternating fields (symmetric and asymmetric) as well as random telegraph signals.

Assessment of Transition Element Speciation in Glasses Using a Portable Transmission Ultraviolet-Visible-Near-Infrared (UV-Vis-NIR) Spectrometer.

A new low-cost experimental setup based on two compact dispersive optical spectrometers has been developed to measure optical absorption transmission spectra over the 350-2500 nm energy range. We demonstrate how near-infrared (NIR) data are essential to identify the coloring species in addition to ultraviolet visible data. After calibration with reference glasses, the use of an original sample stage that maintains the window panel in the vertical position enables the comparison of ancient and modern glasses embedded in a panel from the Sainte-Chapelle of Paris, without any sampling. The spectral resolution enables to observe fine resonances arising in the absorption bands of Cr(3+), and the complementary information obtained in the NIR enables to determine the contribution of Fe(2+), a key indicator of glassmaking conditions.

The theory of DNA separation by capillary electrophoresis.

The Human Genome has been sequenced in large part owing to the invention of capillary electrophoresis. Although this technology has matured enough to allow such amazing achievements, the physical mechanisms at play during separation have yet to be completely understood and optimized. Recently, new separation regimes and new physical mechanisms have been investigated. The use of free-flow electrophoresis and new modes of pulsed-field electrophoresis have been suggested, while we have observed a shift towards single nucleotide polymorphism analysis and microchip technologies. A strong theoretical basis remains essential for the efficient development of new methods.

Intraspinal micro stimulation generates locomotor-like and feedback-controlled movements.

Intraspinal microstimulation (ISMS) may provide a means for improving motor function in people suffering from spinal cord injuries, head trauma, or stroke. The goal of this study was to determine whether microstimulation of the mammalian spinal cord could generate locomotor-like stepping and feedback-controlled movements of the hindlimbs. Under pentobarbital anesthesia, 24 insulated microwires were implanted in the lumbosacral cord of three adult cats. The cats were placed in a sling leaving all limbs pendent. Bilateral alternating stepping of the hindlimbs was achieved by stimulating through as few as two electrodes in each side of the spinal cord. Typical stride lengths were 23.5 cm, and ample foot clearance was achieved during swing. Mean ground reaction force during stance was 36.4 N, sufficient for load-bearing. Feedback-controlled movements of the cat's foot were achieved by reciprocally modulating the amplitude of stimuli delivered through two intraspinal electrodes generating ankle flexion and extension such that the distance between a sensor on the cat's foot and a free sensor moved back and forth by the investigators was minimized. The foot tracked the displacements of the target sensor through its normal range of motion. Stimulation through electrodes with tips in or near lamina IX elicited movements most suitable for locomotion. In chronically implanted awake cats, stimulation through dorsally located electrodes generated paw shakes and flexion-withdrawals consistent with sensory perception but no weight-bearing extensor movements. These locations would not be suitable for ISMS in incomplete spinal cord injuries. Despite the complexity of the spinal neuronal networks, our results demonstrate that by stimulating through a few intraspinal microwires, near-normal bipedal locomotor-like stepping and feedback-controlled movements could be achieved.

Building reliable lattice Monte Carlo models for real drift and diffusion problems.

We revisit the well-known issue of representing an overdamped drift-and-diffusion system by an equivalent lattice random-walk model. We demonstrate that commonly used Monte Carlo algorithms do not conserve the diffusion coefficient when a driving field of arbitrary amplitude is present, and that such algorithms would actually require fluctuating jumping times and one clock per Cartesian direction to work properly. Although it is in principle possible to construct valid algorithms with fixed time steps, we show that no such algorithm can be used in more than two dimensions if the jumps are made along only one axis at each time step.

Size-dependent surface phase change of lithium iron phosphate during carbon coating.

Carbon coating is a simple, effective and common technique for improving the conductivity of active materials in lithium ion batteries. However, carbon coating provides a strong reducing atmosphere and many factors remain unclear concerning the interface nature and underlying interaction mechanism that occurs between carbon and the active materials. Here, we present a size-dependent surface phase change occurring in lithium iron phosphate during the carbon coating process. Intriguingly, nanoscale particles exhibit an extremely high stability during the carbon coating process, whereas microscale particles display a direct visualization of surface phase changes occurring at the interface at elevated temperatures. Our findings provide a comprehensive understanding of the effect of particle size during carbon coating and the interface interaction that occurs on carbon-coated battery material--allowing for further improvement in materials synthesis and manufacturing processes for advanced battery materials.

Modeling inhomogeneous DNA replication kinetics.

In eukaryotic organisms, DNA replication is initiated at a series of chromosomal locations called origins, where replication forks are assembled proceeding bidirectionally to replicate the genome. The distribution and firing rate of these origins, in conjunction with the velocity at which forks progress, dictate the program of the replication process. Previous attempts at modeling DNA replication in eukaryotes have focused on cases where the firing rate and the velocity of replication forks are homogeneous, or uniform, across the genome. However, it is now known that there are large variations in origin activity along the genome and variations in fork velocities can also take place. Here, we generalize previous approaches to modeling replication, to allow for arbitrary spatial variation of initiation rates and fork velocities. We derive rate equations for left- and right-moving forks and for replication probability over time that can be solved numerically to obtain the mean-field replication program. This method accurately reproduces the results of DNA replication simulation. We also successfully adapted our approach to the inverse problem of fitting measurements of DNA replication performed on single DNA molecules. Since such measurements are performed on specified portion of the genome, the examined DNA molecules may be replicated by forks that originate either within the studied molecule or outside of it. This problem was solved by using an effective flux of incoming replication forks at the model boundaries to represent the origin activity outside the studied region. Using this approach, we show that reliable inferences can be made about the replication of specific portions of the genome even if the amount of data that can be obtained from single-molecule experiments is generally limited.

First permanent human implant of the Stimulus Router System, a novel neuroprosthesis: preliminary testing of a polarity reversing stimulation technique.

Neuroprostheses (NPs) are electrical stimulators that help to restore sensory or motor functions lost as a result of neural damage. The Stimulus Router System (SRS) is a new type of NP developed in our laboratory. The system uses fully implanted, passive leads to "capture" and "route" some of the current flowing between pairs of surface electrodes to the vicinity of the target nerves, hence eliminating the need for an implanted stimulator. In June 2008, 3 SRS leads were implanted in a tetraplegic man for restoration of grasp and release. To reduce the size of the external wristlet and thereby optimize usability, we recently implemented a polarity reversing stimulation technique that allowed us to eliminate a reference electrode. Selective activation of three target muscles was achieved by switching the polarities of the stimulus current delivered between pairs of surface electrodes located over the pick-up terminals of the implanted leads and reducing the amplitude of the secondary phases of the stimulus pulses.

Evaluation of tooth-click triggering and speech recognition in assistive technology for computer access.

BACKGROUND: Computer access can play an important role in employment and leisure activities following spinal cord injury. The authors' prior work has shown that a tooth-click detecting device, when paired with an optical head mouse, may be used by people with tetraplegia for controlling cursor movement and mouse button clicks. OBJECTIVE: To compare the efficacy of tooth clicks to speech recognition and that of an optical head mouse to a gyrometer head mouse for cursor and mouse button control of a computer. METHODS: Six able-bodied and 3 tetraplegic subjects used the devices listed above to produce cursor movements and mouse clicks in response to a series of prompts displayed on a computer. The time taken to move to and click on each target was recorded. RESULTS: The use of tooth clicks in combination with either an optical head mouse or a gyrometer head mouse can provide hands-free cursor movement and mouse button control at a speed of up to 22% of that of a standard mouse. Tooth clicks were significantly faster at generating mouse button clicks than speech recognition when paired with either type of head mouse device. CONCLUSIONS: Tooth-click detection performed better than speech recognition when paired with both the optical head mouse and the gyrometer head mouse. Such a system may improve computer access for people with tetraplegia.

Control of DNA replication by anomalous reaction-diffusion kinetics.

We propose a simple model for the control of DNA replication in which the rate of initiation of replication origins is controlled by protein-DNA interactions. Analyzing recent data from Xenopus frog embryos, we find that the initiation rate is reaction limited until nearly the end of replication, when it becomes diffusion limited. Initiation of origins is suppressed when the diffusion-limited search time dominates. To fit the experimental data, we find that the interaction between DNA and the rate-limiting protein must be subdiffusive.

Sequence effects on the forced translocation of heteropolymers through a small channel.

By using a recently developed Monte Carlo algorithm and an exact numerical method, we calculate the translocation probability and the average translocation time for charged heterogeneous polymers driven through a nanopore by an external electric field. The heteropolymer chains are composed of two types of monomers (A and B) which differ only in terms of their electric charge. We present an exhaustive study of chains composed of eight monomers by calculating the average translocation time associated with the 256 possible arrangements for various ratios of the monomer charges (lambda(A)lambda(B)) and electric field intensities E. We find that each sequence leads to a unique value of the translocation probability and time. We also show that the distribution of translocation times is strongly dependent on the two forces felt by the monomers ( approximately lambda(A)E and approximately lambda(B)E). Finally, we present results that highlight the effect of having repetitive patterns by studying the translocation times of various block copolymer structures for a very long chain composed of N=2(18) monomers (all with the same number of A and B monomers).