Tagless LysoIP for immunoaffinity enrichment of native lysosomes from clinical samples.

Lysosomes are implicated in a wide spectrum of human diseases including monogenic lysosomal storage disorders (LSDs), age-associated neurodegeneration and cancer. Profiling lysosomal content using tag-based lysosomal immunoprecipitation (LysoTagIP) in cell and animal models has substantially moved the field forward, but studying lysosomal dysfunction in human patients remains challenging. Here, we report the development of the 'tagless LysoIP' method, designed to enable the rapid enrichment of lysosomes, via immunoprecipitation, using the endogenous integral lysosomal membrane protein TMEM192, directly from clinical samples and human cell lines (e.

In-depth mass-spectrometry reveals phospho-RAB12 as a blood biomarker of G2019S LRRK2-driven Parkinson's disease.

Leucine-rich repeat kinase 2 (LRRK2) inhibition is a promising disease-modifying therapy for LRRK2-associated Parkinson's disease (L2PD) and idiopathic PD (iPD). However, pharmaco-dynamic readouts and progression biomarkers for clinical trials aiming for disease modification are insufficient since no endogenous marker reflecting enhanced kinase activity of the most common LRRK2 G2019S mutation has been reported yet in L2PD patients. Employing phospho-/proteomic analyses we assessed the impact that LRRK2 activating mutations had in peripheral blood mononuclear cells (PBMCs) from a LRRK2 clinical cohort from Spain (n=174).

Silencing Multiple Crustacean Hyperglycaemic Hormone-Encoding Genes in the Redclaw Crayfish Induces Faster Molt Rates with Anomalies.

RNA interference (RNAi)-based biotechnology has been previously implemented in decapod crustaceans. Unlike traditional RNAi methodologies that investigate single gene silencing, we employed a multigene silencing approach in decapods based on chimeric double-stranded RNA (dsRNA) molecules coined 'gene blocks'. Two dsRNA constructs, each targeting three genes of the crustacean hyperglycaemic hormone (CHH) superfamily of neuropeptides, were produced: Type II construct targeting Molt-inhibiting hormone 1 (MIH1), MIH-like 1 (MIHL1), and MIHL2 isoforms and Type I construct targeting ion transport peptide (ITP; a putative hybrid of CHH and MIH) and CHH and CHH-like (CHHL) isoforms.

Transcriptomic Analysis across Crayfish () Claw Regeneration Reveals Potential Stem Cell Sources for Cultivated Crustacean Meat.

In the face of rising global demand and unsustainable production methods, cultivated crustacean meat (CCM) is proposed as an alternative means to produce delicious lobster, shrimp, and crab products. Cultivated meat requires starting stem cells that may vary in terms of potency and the propensity to proliferate or differentiate into myogenic (muscle-related) tissues. Recognizing that regenerating limbs are a non-lethal source of tissue and may harbor relevant stem cells, we selected those of the crayfish as our model.

Transcriptomic Changes Following Induced De-Masculinisation of Australian Red Claw Crayfish .

The Australian red claw crayfish , an emerging species within the freshwater aquaculture trade, is not only an ideal species for commercial production due to its high fecundity, fast growth, and physiological robustness but also notoriously invasive. Investigating the reproductive axis of this species has been of great interest to farmers, geneticists, and conservationists alike for many decades; however, aside from the characterisation of the key masculinising insulin-like androgenic gland hormone (IAG) produced by the male-specific androgenic gland (AG), little remains known about this system and the downstream signalling cascade involved. This investigation used RNA interference to silence in adult intersex (), known to be functionally male but genotypically female, successfully inducing sexual redifferentiation in all individuals.

Unravelling the neuropeptidome of the ornate spiny lobster Panulirus ornatus: A focus on peptide hormones and their processing enzymes expressed in the reproductive tissues.

Neuropeptides are commonly produced in the neural tissues yet can have effects on far-reaching targets, with varied biological responses. We describe here the neuropeptidome of the ornate spiny lobster, Panulirus ornatus, a species of emerging importance to closed-system aquaculture, with a focus on peptide hormones produced by the reproductive tissues. Transcripts for a precursor to one neuropeptide, adipokinetic hormone/corazonin-related peptide (ACP) were identified in high numbers in the sperm duct of adult spiny lobsters suggesting a role for ACP in the reproduction of this species.

Pneumomediastinum due to spontaneous tracheal breach in COVID-19.

Patients who have contracted coronavirus disease 2019 (COVID-19) have a wide variety of complications, many of them involving the respiratory system. One noted complication has been pneumomediastinum. The 63-year-old gentleman, in this case, had contracted COVID-19 and was admitted to the hospital for hypoxemia.

A dual high-energy radiography platform with 15 μm resolution at the National Ignition Facility.

To study matter at extreme densities and pressures, we need mega laser facilities such as the National Ignition Facility as well as creative methods to make observations during timescales of a billionth of a second. To facilitate this, we developed a platform and diagnostic to characterize a new point-projection radiography configuration using two micro-wires irradiated by a short pulse laser system that provides a large field of view with up to 3.6 ns separation between images.

Ion regulation in a freshwater crab, Potamonautes warreni: The effects of trace metal exposure.

Crustaceans inhabiting metal-contaminated freshwaters are susceptible to toxic insult to their osmoregulatory systems. The main osmoregulatory organs of decapod crustaceans, the gills, are continually bathed in freshwater and are therefore at risk from trace metal impacts. The effects of chronic (21 d) exposure to raised dissolved concentrations of Zn, Cd, Cu and Pb on aspects of hydromineral balance were investigated in Potamonautes warreni, a freshwater crab endemic to rivers in South Africa at potential risk from trace metal contamination from mining operations.

Single line-of-sight dual energy backlighter for mix width experiments.

We present a diagnostic technique used to spatially multiplex two x-ray radiographs of an object onto a detector along a single line-of-sight. This technique uses a thin, <2 μm, cosputtered backlighter target to simultaneously produce both Ni and Zn Heα emission. A Ni picket fence filter, 500 μm wide bars and troughs, is then placed in front of the detector to pass only the Ni Heα emission in the bar region and both energies in the trough region thereby spatially multiplexing the two radiographs on a single image.

Glutamate-gated chloride channels of Haemonchus contortus restore drug sensitivity to ivermectin resistant Caenorhabditis elegans.

Anthelmintic resistance is a major problem in livestock farming, especially of small ruminants, but our understanding of it has been limited by the difficulty in carrying out functional genetic studies on parasitic nematodes. An important nematode infecting sheep and goats is Haemonchus contortus; in many parts of the world this species is resistant to almost all the currently available drugs, including ivermectin. It is extremely polymorphic and to date it has proved impossible to relate any sequence polymorphisms to its ivermectin resistance status.

Long duration backlighter experiments at Omega.

We have successfully demonstrated a 7.5 ns duration pinhole-apertured backlighter at the Omega laser facility. Pinhole-apertured point-projection backlighting for 8 ns will be useful for imaging evolving features in experiments at the National Ignition Facility.

Demonstration of the density dependence of x-ray flux in a laser-driven hohlraum.

Experiments have been conducted using laser-driven cylindrical hohlraums whose walls are machined from Ta2O5 foams of 100 mg/cc and 4 g/cc densities. Measurements of the radiation temperature demonstrate that the lower density walls produce higher radiation temperatures than the high density walls. This is the first experimental demonstration of the prediction that this would occur [M.

High-Energy X-ray Microscopy Techniques for Laser-Fusion Plasma Research at the National Ignition Facility.

Multi-kilo-electron-volt x-ray microscopy will be an important laser-produced plasma diagnostic at future megajoule facilities such as the National Ignition Facility (NIF). However, laser energies and plasma characteristics imply that x-ray microscopy will be more challenging at NIF than at existing facilities. We use analytical estimates and numerical ray tracing to investigate several instrumentation options in detail, and we conclude that near-normal-incidence single spherical or toroidal crystals may offer the best general solution for high-energy x-ray microscopy at NIF and similar large facilities.

Dynamic Hohlraums as x-ray sources in high-energy density science.

The first demonstration of laser driven dynamic Hohlraums (LDDH) as a spectrally smooth backlighter source for opacity and temperature measurements through absorption spectrometry of materials in local thermodynamic equilibrium at temperatures >150 eV has been made. This is a crucial temperature regime for future astrophysics and ignition fusion experiments at the nearly completed National Ignition Facility (NIF) [E. I.

Efficient, 1-100-keV x-ray radiography with high spatial and temporal resolution.

Transient x-ray radiography using laser-plasma emission is a powerful tool for diagnosing a large variety of high-energy-density phenomena. Traditional area- and point-backlighting techniques used at inertial confinement fusion facilities such as Nova and Omega cannot be extended efficiently to the future 100-times-larger National Ignition Facility. We have developed an x-ray backlighting technique that uses a backlit pinhole as a source for point-projection radiography.

Treatment of sewage sludge using electrokinetic geosynthetics.

The treatment and disposal of sewage sludge is one of the most problematical issues affecting wastewater treatment in the developed world. The traditional outlets for sewage sludge are to spread it on agricultural land, or to form a cake for deposit to landfill or incineration. In order to create a sludge cake, water must be removed.

Experimental investigation of high-mach-number 3D hydrodynamic jets at the national ignition facility.

The first hydrodynamic experiments were performed on the National Ignition Facility. A supersonic jet was formed via the interaction of a laser driven shock ( approximately 40 Mbar) with 2D and 3D density perturbations. The temporal evolution of the jet's spatial scales and ejected mass were measured with point-projection x-ray radiography.

Chained time trade-off and standard gamble methods. Applications in oesophageal cancer.

It may be difficult to value palliative health states using health state valuation methods such as the time trade-off (TTO) and standard gamble (SG) where health states are traditionally valued relative to perfect/good health and death. Chained methods have been developed to help in this context. However, few studies have compared the values produced by chained TTO and SG methods.