A family with partially penetrant multicentric carpotarsal osteolysis due to gonadal mosaicism: First reported case

Multicentric carpotarsal osteolysis (MCTO) is an autosomal dominant condition characterized by carpal-tarsal abnormalities; over half of affected individuals also develop renal disease. MCTO is caused by mutations of MAFB; however, there is no clear phenotype-genotype correlation. We describe the first reported family of variable MCTO phenotype due to mosaicism: the proband had classical skeletal features and renal involvement due to focal segmental glomerulosclerosis (FSGS), and the father had profound renal impairment due to FSGS, necessitating kidney transplantation. Mosaicism was first suspected in this family due to unequal allele ratios in the sequencing chromatograph of the initial blood sample of proband's father and confirmed by sequencing DNA extracted from the father's hair, collected from different bodily parts. This case highlights the need for a high index of clinical suspicion to detect low-level parental mosaicism, as well as a potential role for MAFB mutation screening in individuals with isolated FSGS.Peer reviewe

TiO2 Coating and UV Photofunctionalization Enhance Blood Coagulation on Zirconia Surfaces

This in vitro study was designed to evaluate the effect of sol-gel derived TiO2 coating on blood coagulation, blood protein adsorption, and platelet response on zirconia surfaces. Square-shaped zirconia (n=96) (10x10x2 mm) was cut, ground, sintered, and finally cleansed ultrasonically in each of acetone and ethanol for 5 minutes. Three experimental groups (n=32) were fabricated: (a) zirconia coated with sol-gel derived TiO2, (b) zirconia coated with sol-gel derived TiO2 and treated with ultraviolet (UV) irradiation for 1 hour, and (c) non-coated zirconia as control. The coatings were prepared from tetraisopropyl orthotitanate solution by dip-coating. The thrombogenicity of the specimens was evaluated using a whole blood kinetic clotting time method where the extent of blood clotting was evaluated at 10, 20, 30, 40, 50, and 60 minutes (n=4/time point, total n=24/group). Scanning electron microscope images were taken to observe platelet morphologies after 1-hour incubation with platelet-rich plasma (PRP) (n=5/group). Surface characteristics were visualized using atomic force microscopy (n=1/group). Adsorption of plasma proteins and fibronectin on each surface was studied by gel electrophoresis (n=2/group). Significant differences were observed in blood coagulation between the test groups at 20-, 30-, 40-, and 50-minute time points (p<0.005). UV treated TiO2 coated specimens showed fastest blood coagulation followed by TiO2 coated and non-coated specimens. Furthermore, platelets appeared at a higher activation state on coated specimens. Gel electrophoresis revealed no difference in protein adsorption among the experimental groups. In summary, TiO2 coatings promoted blood coagulation, and it was further enhanced by UV treatment, which has the potential to hasten the wound healing process in vivo

The Effect of Exposed Glass Fibers and Particles of Bioactive Glass on the Surface Wettability of Composite Implants

Measurement of the wettability of a material is a predictive index of cytocompatibility. This study was designed to evaluate the effect of exposed E-glass fibers and bioactive glass (BAG) particles on the surface wettability behavior of composite implants. Two different groups were investigated: (a) fiber reinforced composites (FRCs) with different fiber orientations and (b) polymer composites with different wt. % of BAG particles. Photopolymerized and heat postpolymerized composite substrates were made for both groups. The surface wettability, topography, and roughness were analyzed. Equilibrium contact angles were measured using the sessile drop method. Three liquids were used as a probe for surface free energy (SFE) calculations. SFE values were calculated from contact angles obtained on smooth surfaces. The surface with transverse distribution of fibers showed higher () polar () and total SFE () components (16.9 and 51.04 mJ/m2, resp.) than the surface with in-plane distribution of fibers (13.77 and 48.27 mJ/m2, resp.). The increase in BAG particle wt. % increased the polar () value, while the dispersive () value decreased. Postpolymerization by heat treatment improved the SFE components on all the surfaces investigated (). Composites containing E-glass fibers and BAG particles are hydrophilic materials that show good wettability characteristics.</p

Site-specific perturbations of alpha-synuclein fibril structure by the Parkinson's disease associated mutations A53T and E46K.

PMCID: PMC3591419This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Parkinson's disease (PD) is pathologically characterized by the presence of Lewy bodies (LBs) in dopaminergic neurons of the substantia nigra. These intracellular inclusions are largely composed of misfolded α-synuclein (AS), a neuronal protein that is abundant in the vertebrate brain. Point mutations in AS are associated with rare, early-onset forms of PD, although aggregation of the wild-type (WT) protein is observed in the more common sporadic forms of the disease. Here, we employed multidimensional solid-state NMR experiments to assess A53T and E46K mutant fibrils, in comparison to our recent description of WT AS fibrils. We made de novo chemical shift assignments for the mutants, and used these chemical shifts to empirically determine secondary structures. We observe significant perturbations in secondary structure throughout the fibril core for the E46K fibril, while the A53T fibril exhibits more localized perturbations near the mutation site. Overall, these results demonstrate that the secondary structure of A53T has some small differences from the WT and the secondary structure of E46K has significant differences, which may alter the overall structural arrangement of the fibrils

Physical characterixation and in vitro biological impact of highly aggregated antibodies separated into size-enriched populations by fluorescence-activated cell sorting

An IgG2 monoclonal antibody (mAb) solution was subjected to stirring, generating high concentrations of nanometer and subvisible particles, which were then successfully size enriched into different size bins by low speed centrifugation or a combination of gravitational sedimentation and Fluorescence-Activated Cell Sorting (FACS). The size-fractionated mAb particles were assessed for their ability to elicit the release of cytokines from a population of donor-derived human peripheral blood mononuclear cells (PBMC) at two phases of the immune response. Fractions enriched in nanometer-sized particles showed a lower response than those enriched in micron-sized particles in this assay. Particles of 5–10 μm in size displayed elevated cytokine release profiles compared to other size ranges. Stir-stressed mAb particles had amorphous morphology, contained protein with partially altered secondary structure, elevated surface hydrophobicity (compared to controls), and trace levels of elemental fluorine. FACS size-enriched the mAb particle samples, yet did not notably alter the overall morphology or composition of particles as measured by Microflow imaging, Transmission Electron Microscopy, and Scanning Electron Microscopy-Energy Dispersive X-ray Spectroscopy. The utility and limitations of FACS for size separation of mAb particles and potential of in-vitro PBMC studies to rank order the immunogenic potential of various types of mAb particles is discussed

Characterization of Particles in Protein Solutions: Reaching the Limits of Current Technologies

Recent publications have emphasized the lack of characterization methods available for protein particles in a size range comprised between 0.1 and 10 μm and the potential risk of immunogenicity associated with such particles. In the present paper, we have investigated the performance of light obscuration, flow microscopy, and Coulter counter instruments for particle counting and sizing in protein formulations. We focused on particles 2–10 μm in diameter and studied the effect of silicon oil droplets originating from the barrel of pre-filled syringes, as well as the effect of high protein concentrations (up to 150 mg/ml) on the accuracy of particle characterization. Silicon oil was demonstrated to contribute significantly to the particle counts observed in pre-filled syringes. Inconsistent results were observed between different protein concentrations in the range 7.5–150 mg/ml for particles <10 μm studied by optical techniques (light obscuration and flow microscopy). However, the Coulter counter measurements were consistent across the same studied concentration range but required sufficient solution conductivity from the formulation buffer or excipients. Our results show that currently available technologies, while allowing comparisons between samples of a given protein at a fixed concentration, may be unable to measure particle numbers accurately in a variety of protein formulations, e.g., at high concentration in sugar-based formulations

Signs of oral dryness in relation to salivary flow rate, pH, buffering capacity and dry mouth complaints

<p>Abstract</p> <p>Background</p> <p>This study aimed to investigate the signs of oral dryness in relation to different salivary variables and to correlate subjective complaints of oral dryness with salivary flow rate.</p> <p>Methods</p> <p>312 unmedicated healthy individuals belonging to three age groups, (6–11, 12–17, and 18–40 years) were examined clinically for signs of oral dryness. Resting and stimulated saliva were collected to determine flow rate, pH and buffering capacity. A questionnaire was used to obtain information on subjective sensation of dry mouth.</p> <p>Results</p> <p>Dry lip and dry mucosa were present in 37.5% and 3.2% of the sample respectively. The proportion of subjects who complained of oral dryness (19%) showed a stimulated salivary flow rate significantly lower than non complainers. Dry lip was significantly related to low resting flow rate but pH and buffering capacity did not show any significant relation to dry lip. Dry mucosa was not related to any of the above mentioned parameters.</p> <p>Conclusion</p> <p>The finding that the stimulated salivary flow rate was reduced in subjects complaining of dry mouth is of great clinical relevance, since the reduction is expected to be reflected in compromising various salivary functions.</p

The Role of Alpha-Synuclein Oligomerization and Aggregation in Cellular and Animal Models of Parkinson’s Disease

α-synuclein (α-syn) is a synaptic protein in which four mutations (A53T, A30P, E46K and gene triplication) have been found to cause an autosomal dominant form of Parkinson’s disease (PD). It is also the major component of intraneuronal protein aggregates, designated as Lewy bodies (LBs), a prominent pathological hallmark of PD. How α-syn contributes to LB formation and PD is still not well-understood. It has been proposed that aggregation of α-syn contributes to the formation of LBs, which then leads to neurodegeneration in PD. However, studies have also suggested that aggregates formation is a protective mechanism against more toxic α-syn oligomers. In this study, we have generated α-syn mutants that have increased propensity to form aggregates by attaching a CL1 peptide to the C-terminal of α-syn. Data from our cellular study suggest an inverse correlation between cell viability and the amount of α-syn aggregates formed in the cells. In addition, our animal model of PD indicates that attachment of CL1 to α-syn enhanced its toxicity to dopaminergic neurons in an age-dependent manner and induced the formation of Lewy body-like α-syn aggregates in the substantia nigra. These results provide new insights into how α-syn-induced toxicity is related to its aggregation

Distinct clinical and neuropathological features of G51D SNCA mutation cases compared with SNCA duplication and H50Q mutation

Background: We and others have described the neurodegenerative disorder caused by G51D SNCA mutation which shares characteristics of Parkinson’s disease (PD) and multiple system atrophy (MSA). The objective of this investigation was to extend the description of the clinical and neuropathological hallmarks of G51D mutant SNCA-associated disease by the study of two additional cases from a further G51D SNCA kindred and to compare the features of this group with a SNCA duplication case and a H50Q SNCA mutation case. Results: All three G51D patients were clinically characterised by parkinsonism, dementia, visual hallucinations, autonomic dysfunction and pyramidal signs with variable age at disease onset and levodopa response. The H50Q SNCA mutation case had a clinical picture that mimicked late-onset idiopathic PD with a good and sustained levodopa response. The SNCA duplication case presented with a clinical phenotype of frontotemporal dementia with marked behavioural changes, pyramidal signs, postural hypotension and transiently levodopa responsive parkinsonism. Detailed post-mortem neuropathological analysis was performed in all cases. All three G51D cases had abundant α-synuclein pathology with characteristics of both PD and MSA. These included widespread cortical and subcortical neuronal α-synuclein inclusions together with small numbers of inclusions resembling glial cytoplasmic inclusions (GCIs) in oligodendrocytes. In contrast the H50Q and SNCA duplication cases, had α-synuclein pathology resembling idiopathic PD without GCIs. Phosphorylated α-synuclein was present in all inclusions types in G51D cases but was more restricted in SNCA duplication and H50Q mutation. Inclusions were also immunoreactive for the 5G4 antibody indicating their highly aggregated and likely fibrillar state. Conclusions: Our characterisation of the clinical and neuropathological features of the present small series of G51D SNCA mutation cases should aid the recognition of this clinico-pathological entity. The neuropathological features of these cases consistently share characteristics of PD and MSA and are distinct from PD patients carrying the H50Q or SNCA duplication