Disease Mechanisms and Therapeutic Approaches in SMARD1-Insights from Animal Models and Cell Models.

Spinal muscular atrophy with respiratory distress type 1 (SMARD1) is a fatal childhood motoneuron disease caused by mutations in the IGHMBP2 gene. It is characterized by muscle weakness, initially affecting the distal extremities due to the degeneration of spinal α-motoneurons, and respiratory distress, due to the paralysis of the diaphragm. Infantile forms with a severe course of the disease can be distinguished from juvenile forms with a milder course. Mutations in the IGHMBP2 gene have also been found in patients with peripheral neuropathy Charcot-Marie-Tooth type 2S (CMT2S). IGHMBP2 is an ATP-dependent 5'→3' RNA helicase thought to be involved in translational mechanisms. In recent years, several animal models representing both SMARD1 forms and CMT2S have been generated to initially study disease mechanisms. Later, the models showed very well that both stem cell therapies and the delivery of the human IGHMBP2 cDNA by AAV9 approaches (AAV9-IGHMBP2) can lead to significant improvements in disease symptoms. Therefore, the SMARD1 animal models, in addition to the cellular models, provide an inexhaustible source for obtaining knowledge of disease mechanisms, disease progression at the cellular level, and deeper insights into the development of therapies against SMARD1.

Sub-millisecond microfluidic mixers coupled to time-resolved in situ photonics to study ultra-fast reaction kinetics: the case of ultra-small gold nanoparticle synthesis.

We report a continuous microreactor platform achieving sub-millisecond homogeneous reagent mixing (∼300 µs) for a time-resolved study on the synthesis of ultra-small gold nanoparticles (NPs). The microreactor (coupled with small angle X-ray scattering, UV-vis, and X-ray absorption spectroscopy for in situ and in operando characterizations), operates within mixing time frames below system characteristic times, providing a unique opportunity to deepen the comprehension of reaction and phase transition pathways with unprecedented details. The microreactor channel length can be approximated to a given reaction time when operated in continuous mode and steady state. As a result, the system can be statically investigated, eliminating technique-dependent probing time constraints and local inhomogeneities caused by mixing issues. We have studied Au(0) NP formation kinetics from Au(III) precursors complexed with oleylamine in organic media, using triisopropylsilane as a reducing agent. The existence of Au(III)/Au(I) prenucleation clusters and the formation of a transient Au(I) lamellar phase under certain conditions, before the onset of Au(0) formation, have been observed. Taking advantage of the high frequency time-resolved information, we propose and model two different reaction pathways associated with the presence or absence of the Au(I) lamellar phase. In both cases, non-classical pathways leading to the formation of NPs are discussed.

RNA helicase IGHMBP2 regulates THO complex to ensure cellular mRNA homeostasis.

RNA helicases constitute a large protein family implicated in cellular RNA homeostasis and disease development. Here, we show that the RNA helicase IGHMBP2, linked to the neuromuscular disorder spinal muscular atrophy with respiratory distress type 1 (SMARD1), associates with polysomes and impacts translation of mRNAs containing short, GC-rich, and structured 5' UTRs. The absence of IGHMBP2 causes ribosome stalling at the start codon of target mRNAs, leading to reduced translation efficiency. The main mRNA targets of IGHMBP2-mediated regulation encode for components of the THO complex (THOC), linking IGHMBP2 to mRNA production and nuclear export. Accordingly, failure of IGHMBP2 regulation of THOC causes perturbations of the transcriptome and its encoded proteome, and ablation of THOC subunits phenocopies these changes. Thus, IGHMBP2 is an upstream regulator of THOC. Of note, IGHMBP2-dependent regulation of THOC is also observed in astrocytes derived from patients with SMARD1 disease, suggesting that deregulated mRNA metabolism contributes to SMARD1 etiology and may enable alternative therapeutic avenues.

Contribution of lowered hemoglobin threshold value in the diagnosis of polycythemia vera: Comparison of 2016 and 2008 WHO criteria.

Polycythemia vera (PV) diagnosis remains a difficult task despite various updates in the 2016 World Health Organization (WHO) diagnostic criteria compared to 2008 criteria. This study aimed to examine the biochemical and clinical features of patients diagnosed with PV using the WHO 2016 criteria but would have been missed by the WHO 2008 criteria, and to ascertain the impact of the lowered thresholds on PV diagnosis. A total of 229 patients with suspected myeloproliferative neoplasms were included in this cross sectional study. The study group was divided with regard to hemoglobin values. Group A consisted of 126 patients with hemoglobin values of ≤ 18.5 g/dL in males and ≤ 16.5 g/dL in females. Group B comprised 103 patients with hemoglobin values of > 18.5 g/dL in males and > 16.5 g/dL in females. The number of PV diagnoses increased to 145 from 87 (increased by 66.67%) when the 2016 diagnostic criteria were employed rather that the 2008 criteria. Mean age and the frequency of female subjects were lower in Group A compared to Group B. The groups were similar in terms of chronic obstructive pulmonary disease/obstructive sleep apnea syndrome, spleen status, smoking status, and mean corpuscular volume, white blood count, neutrophil, eosinophil and platelet values. red blood cells and lactate dehydrogenase values were significantly higher, while lymphocyte counts were significantly lower in Group B. With the introduction of WHO 2016 criteria, we found a significant increase in the number of patients who were candidates for PV testing and were ultimately diagnosed with PV. These findings support the diagnostic value of the 2016 WHO criteria, and by extension, the lowered thresholds for detection of patients requiring further analysis.

Plastin 3 rescues cell surface translocation and activation of TrkB in spinal muscular atrophy.

Plastin 3 (PLS3) is an F-actin-bundling protein that has gained attention as a modifier of spinal muscular atrophy (SMA) pathology. SMA is a lethal pediatric neuromuscular disease caused by loss of or mutations in the Survival Motor Neuron 1 (SMN1) gene. Pathophysiological hallmarks are cellular maturation defects of motoneurons prior to degeneration. Despite the observed beneficial modifying effect of PLS3, the mechanism of how it supports F-actin-mediated cellular processes in motoneurons is not yet well understood. Our data reveal disturbed F-actin-dependent translocation of the Tropomyosin receptor kinase B (TrkB) to the cell surface of Smn-deficient motor axon terminals, resulting in reduced TrkB activation by its ligand brain-derived neurotrophic factor (BDNF). Improved actin dynamics by overexpression of hPLS3 restores membrane recruitment and activation of TrkB and enhances spontaneous calcium transients by increasing Cav2.1/2 "cluster-like" formations in SMA axon terminals. Thus, our study provides a novel role for PLS3 in supporting correct alignment of transmembrane proteins, a key mechanism for (moto)-neuronal development.

The role of pre-nucleation clusters in the crystallization of gold nanoparticles.

The syntheses of metal nanoparticles by reduction in apolar solvents in the presence of long chain surfactants have proven to be extremely effective in the control of the particle size and shape. Nevertheless, the elucidation of the nucleation/growth mechanism is not straightforward because of the multiple roles played by surfactants. The nucleation stage, in particular, is very difficult to describe precisely and requires in situ and time-resolved techniques. Here, relying on in situ small angle X-ray scattering (SAXS), X-ray absorption spectroscopy (XAS) and high-energy X-ray diffraction (HE-XRD), we propose that ultra-small gold particles prepared by reduction of gold chloride in a solution of oleylamine (OY) in hexane with triisopropylsilane do not follow a classical nucleation process but result from pre-nucleation clusters (PNCs). These PNCs contain Au(iii) and Au(i) precursors; they are almost stable in size during the induction stage, as shown by SAXS, prior to undergoing a very fast shrinkage during the nucleation stage. The gold speciation as a function of time deduced from the XAS spectra has been analyzed through multi-step reaction pathways comprising both highly reactive species, involved in the nucleation and growth stages, and poorly reactive species acting as a reservoir for the reactive species. The duration of the induction period is related to the reactivity of the gold precursors, which is tuned by the coordination of OY to the gold complexes, while the nucleation stage was found to depend on the size and reactivity of the PNCs. The role of the PNCs in determining the final particle size and structure is also discussed in relation to previous studies. The multiple roles of OY, as the solubilizing agent of the gold salt, the ligand of the gold complexes determining both the size of the PNCs and the reactivity of the gold precursors, and finally the capping agent of the final gold particles as oleylammonium chloride, have been clearly established. This work opens new perspectives to synthesize metal NPs via metal-organic PNCs and to define new synthesis routes for nanoparticles that may present structure and morphologies different from those obtained by the classical nucleation routes.

In Vitro Wound Healing Activity of Wheat-Derived Nanovesicles.

Triticum aestivum plant extracts are often used as a natural healer in traditional medicine but which particles mainly have role in these processes are not scientifically proven. In other words, no attempts have been made to investigate the effects of wheat exosomes in regenerative medicine applications or drug development up to now. The current study was first time performed to demonstrate the activity of wheat exosomes in wound healing process using in vitro approaches. Although its fundamental wound healing process remains a mystery, in the current study, the efficiency of wheat grass juice-derived exosomes on cell viability and migration was examined. Increasing concentrations up to 200 µg/mL of the wheat exosome have yielded astonishing proliferative and migratory effects on endothelial, epithelial, and dermal fibroblast cells. RT-PCR analysis also showed collagen type I; mRNA levels were approximately twofold higher in expression after treating with 200 µg/mL wheat exosome. Additionally, Annexin V staining of apoptotic cells accompanied with the cell cycle analysis resulted with the reduction of the apoptotic cell number with no dispersion to the cell cycle analysis while plant exosomes have also increased tube-like structure formation of the endothelial cells. All in all, this research suggests a brand-new opening for skin wound healing therapy strategy by using wheat-derived exosomes due to its proliferative and migratory characteristics. Plant exosomes require a further research both clinically and in in vivo for wound healing drug development. Moreover, plant exosome therapy strategies would be safer and economical alternative for clinical wound healing.