Nusinersen treatment and cerebrospinal fluid neurofilaments: An explorative study on Spinal Muscular Atrophy type 3 patients

The antisense oligonucleotide Nusinersen has been recently licensed to treat spinal muscular atrophy (SMA). Since SMA type 3 is characterized by variable phenotype and milder progression, biomarkers of early treatment response are urgently needed. We investigated the cerebrospinal fluid (CSF) concentration of neurofilaments in SMA type 3 patients treated with Nusinersen as a potential biomarker of treatment efficacy. The concentration of phosphorylated neurofilaments heavy chain (pNfH) and light chain (NfL) in the CSF of SMA type 3 patients was evaluated before and after six months since the first Nusinersen administration, performed with commercially available enzyme-linked immunosorbent assay (ELISA) kits. Clinical evaluation of SMA patients was performed with standardized motor function scales. Baseline neurofilament levels in patients were comparable to controls, but significantly decreased after six months of treatment, while motor functions were only marginally ameliorated. No significant correlation was observed between the change in motor functions and that of neurofilaments over time. The reduction of neurofilament levels suggests a possible early biochemical effect of treatment on axonal degeneration, which may precede changes in motor performance. Our study mandates further investigations to assess neurofilaments as a marker of treatment response.     

MicroRNAs as regulators of cell death mechanisms in amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder affecting upper and lower motor neurons (MNs), resulting in paralysis and precocious death from respiratory failure. Although the causes of ALS are incompletely understood, the role of alterations in RNA metabolism seems central. MicroRNAs (miRNAs) are noncoding RNAs implicated in the regulation of gene expression of many relevant physiological processes, including cell death. The recent model of programmed cell death (PCD) encompasses different mechanisms, from apoptosis to regulated necrosis (RN), in particular necroptosis. Both apoptosis and necroptosis play a significant role in the progressive death of MNs in ALS. In this review, we present key research related to miRNAs that modulate apoptosis and RN pathways in ALS. We also discuss whether these miRNAs represent potential targets for therapeutic development in patients.     

Synthesis of a green polyurethane foam from a biopolyol obtained by enzymatic glycerolysis and its use for immobilization of lipase NS-40116

The use of green sources for materials synthesis has gained popularity in recent years. This work investigated the immobilization of lipase NS-40116 (Thermomyces lanuginosus lipase) in polyurethane foam (PUF) using a biopolyol obtained through the enzymatic glycerolysis between castor oil and glycerol, catalyzed by the commercial lipase Novozym 435 for the PUF formation. The reaction was performed to obtain biopolyol resulting in the conversion of 64% in mono- and diacylglycerol, promoting the efficient use of the reaction product as biopolyol to obtain polyurethane foam. The enzymatic derivative with immobilized lipase NS-40116 presented apparent density of 0.19 ± 0.03 g/cm³ and an immobilization yield was 94 ± 4%. Free and immobilized lipase NS-40116 were characterized in different solvents (methanol, ethanol, and propanol), temperatures (20, 40, 60 and 80 °C), pH (3, 5, 7, 9 and 11) and presence of ions Na⁺, Mg⁺⁺, and Ca⁺⁺. The support provided higher stability to the enzyme, mainly when subjected to acid pH (free lipase lost 80% of relative activity after 360 h of contact, when the enzymatic derivative lost around 22%) and high-temperature free lipase lost 50% of relative activity, while the immobilized remained 95%. The enzymatic derivative was also used for esterification reactions and conversions around 66% in fatty acid methyl esters, using abdominal chicken fat as feedstock, were obtained in the first use, maintaining this high conversion until the fourth reuse, proving that the support obtained using environmentally friendly techniques is applicable.

     

Restoration of human dystrophin following transplantation of exon-skipping-engineered DMD patient stem cells into dystrophic mice

Duchenne muscular dystrophy (DMD) is a hereditary disease caused by mutations that disrupt the dystrophin mRNA reading frame. In some cases, forced exclusion (skipping) of a single exon can restore the reading frame, giving rise to a shorter, but still functional, protein. In this study, we constructed lentiviral vectors expressing antisense oligonucleotides in order to induce an efficient exon skipping and to correct the initial frameshift caused by the DMD deletion of CD133+ stem cells. The intramuscular and intra-arterial delivery of genetically corrected CD133 expressing myogenic progenitors isolated from the blood and muscle of DMD patients results in a significant recovery of muscle morphology, function, and dystrophin expression in scid/mdx mice. These data demonstrate that autologous engrafting of blood or muscle-derived CD133+ cells, previously genetically modified to reexpress a functional dystrophin, represents a promising approach for DMD.     

Coumarin content and physicochemical profile of Mikania laevigata extracts

The 'guaco' lianous herb Mikania laevigata, which is widespread in Southern Brazil, is traditionally used to treat bronchitis, asthma and cough. This work investigates the influence of the extraction method, solvent:drug ratio, ethanol proportion, harvest season (summer or winter) and solvent heating on the physicochemical profile of the extracts (dry weight, density, pH) and the coumarin (1,2-benzopyrone) content determined by LC. Among the results obtained, it is observed that higher ethanol content increases the amount of coumarin in the extract. Leaves harvested in summer also produce an extract with a high coumarin yield. The most efficient method of extraction is percolation, independent of the solvent used.     

Comparative analysis of the human dystrophin and utrophin gene structures

We present analysis of intronic sequences in the human DMD and UTRN genes. In both genes accumulation of repeated elements could account for intron expansion. Out-of-frame rod-domain exons have stronger splice sites and are separated by significantly longer introns as compared to in-frame exons. These features are unique for the two homologs and not shared by other spectrin superfamily genes.     

Physicochemical and mechanical characterization of galactomannan from Mimosa scabrella: Effect of drying method

The galactomannan from Mimosa scabrella Bentham was extracted on a pilot plant scale and dried either by vacuum oven (GVO) or by spray-drier (GSD) to evaluate the effect of the drying technique on the powder quality and its applicability as excipient in solid dosage forms. The analysis by high performance size exclusion chromatography (HPSEC) coupled to multiangle laser light scattering (MALLS) suggests that both products behave as semi-flexible polymers, although the GSD showed more aggregation at molecular level (～10%) and higher chain stiffness (Lp 9.1 nm). TG and DSC analysis showed weight loss event with peak at 299.7 and 311.9 °C to GVO and GSD, respectively, as well and higher ash content for GSD sample, in both inert and oxidant atmosphere. The X-ray diffraction confirmed the amorphous nature of both galactomannans although GVO showed high crystallinity. The GSD showed lower density (1.009 g/cm³), higher cohesiveness (repose angle 35.5°, compressibility 32.2% and absence of flow), smaller and more spherical particles than GVO sample, both with high polydispersion. As vacuum oven-drying resulted in a like fibrous material, spray-drying appears as an alternative method easy to extrapolate in industry, requiring a glidant incorporation to improve the powder flowability.