Characterization of vestibular schwannoma tissues using liquid chromatography-tandem mass spectrometry analysis of specific peptide fragments separated by in-sample tryptic protein digestion followed by mathematical analysis

Vestibular schwannoma is the most common benign neoplasm of the cerebellopontine angle. Its first symptoms include hearing loss, tinnitus, and vestibular symptoms, followed by cerebellar and brainstem symptoms, along with palsy of the adjacent cranial nerves. However, the clinical picture has unpredictable dynamics and currently, there are no reliable predictors of tumor behavior. Hence, it is desirable to have a fast routine method for analysis of vestibular schwannoma tissues at the molecular level. The major objective of this study was to verify whether a technique using in-sample specific protein digestion with trypsin would have the potential to provide a proteomic characterization of these pathological tissues. The achieved results showed that the use of this approach with subsequent liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis of released peptides allowed a fast identification of a considerable number of proteins in two differential parts of vestibular schwannoma tissue as well as in tissues of control healthy samples. Furthermore, mathematical analysis of MS data was able to discriminate between pathological vestibular schwannoma tissues and healthy tissues. Thus, in-sample protein digestion combined with LC-MS/MS separation and identification of released specific peptides followed by mathematical analysis appears to have the potential for routine characterization of vestibular schwannomas at the molecular level. Data are available via ProteomeXchange with identifier PXD045261.     

Development of a Silk Fibroin-Small Intestinal Submucosa Small-Diameter Vascular Graft with Sequential VEGF and TGF-β1 Inhibitor Delivery for In Situ Tissue Engineering

Proper endothelialization and limited collagen deposition on the luminal surface after graft implantation plays a crucial role to prevent the occurrence of stenosis. To achieve these conditions, a biodegradable graft with adequate mechanical properties and the ability to sequentially deliver therapeutic agents isfabricated. In this study, a dual-release system is constructed through coaxial electrospinning by incorporating recombinant human vascular endothelial growth factor (VEGF) and transforming growth factor β1 (TGF-β1) inhibitor into silk fibroin (SF) nanofibers to form a bioactive membrane. The functionalized SF membrane as the inner layer of the graft is characterized by the release profile, cell proliferation and protein expression. It presents excellent biocompatibility and biodegradation, facilitating cell attachment, proliferation, and infiltration. The core-shell structure enables rapid VEGF release within 10 days and sustained plasmid delivery for 21 days. A 2.0-mm-diameter vascular graft is fabricated by integrating the SF membrane with decellularized porcine small intestinal submucosa (SIS), aiming to facilitate the integration process under a stable extracellular matrix structure. The bioengineered graft is functionalized with the sequential administration of VEGF and TGF-β1, and with the reinforced and compatible mechanical properties, thereby offers an orchestrated solution for stenosis with potential for in situ vascular tissue engineering applications.     

Lipid nanotubule fabrication by microfluidic tweezing

There is currently great interest in the development of lipid enclosed systems with complex geometrical arrangements that mimic cellular compartments. With biochemical functionalization, these soft matter devices can be used to probe deeper into life's transport dominated biochemical operations. In this paper, we present a novel tool for machining lipid nanotubules by microfluidic tweezing. A bilayer poly(dimethylsiloxane) (PDMS) device was designed with a lipid reservoir that was loaded by capillary action for lipid film deposition. The lipid reservoir is vertically separated from an upper flow for controlled material wetting and the formation of giant tubule bodies. Three fluidic paths are interfaced for introduction of the giant tubules into the high velocity center of a parabolic flow profile for exposure to hydrodynamic shear stresses. At local velocities approximating 2 mm s (-1), a 300-500 nm diameter jet of lipid material was tweezed from the giant tubule body and elongated with the flow. The high velocity flow provides uniform drag for the rapid and continuous fabrication of lipid nanotubules with tremendous axial ratios. Below a critical velocity, a remarkable shape transformation occurred and the projected lipid tubule grew until a constant 3.6 mum diameter tubule was attained. These lipid tubules could be wired for the construction of advanced lifelike bioreactor systems.     

Crystalline inclusion compounds of lower rim propyl substituted calix[4]arenes featuring different number and positions of the modifying groups

Three lower rim n-propyl substituted calix[4]arenes (1–3) with varied number and position of the modifying groups have been prepared. Inclusion compounds (five species) involving different kinds of guest solvents have been isolated. Their X-ray crystal structures were determined and comparatively discussed using isostructurality calculations. Two of the inclusion compounds obtained (1a and 1b) are polymorphs containing the same host and guest molecules in equal stoichiometric ratio but different Z′ values caused by a phase transition around 140 K. The inclusion compounds 2a and 2b refer to the interesting case of a mixed solvent complex while 3a allows studying the effect of full lower rim n-propyl substitution.     

Determination of N-glycosylation sites and site heterogeneity in a monoclonal antibody by electrospray quadrupole ion-mobility time-of-flight mass spectrometry

This paper presents an improved analytical method for glycosylation structural characterizations of a monoclonal antibody (mAb) using a newly developed quadrupole ion-mobility time-of-flight (ESI-Q-IM-TOF) mass spectrometer. Using this method, high-resolution mass spectra were acquired to produce the overall glycosylation profile of the mAb. Additionally, the light and heavy chains from the reduced antibody were separated in the gas phase by the ion mobility functionality of the instrument, allowing accurate mass measurement of each subunit. Furthermore, the glycan sequences, as well as the glycosylation site, were determined by a two-step sequential fragmentation process using the unique dual-collision-cell design of the instrument, thus providing detailed characterizations of the glycan structures.     

Applications of solvent extraction in the high-yield multi-process reduction/separation of Eu from excess Sm

A novel multi-process method for separating Eu from neighbouring lanthanides (Ln) has been developed that chemically reduces Eu(III) to Eu(II) prior to solvent extraction of Ln(III) with thenoyltrifluoroacetone in benzene. This method is capable of achieving higher purities (>99%) and separation yields than previously published multi-process methods that stabilize and separate the reduced Eu(II) as a sulphate solid and is ideal for enriching materials of high-value. Results from a variety of combinations of a chemical or electrochemical reduction process preceding a separation process using either ion-exchange chromatography, reversed phase chromatography, or solvent extraction are discussed.     

Analysis of single blood cells for CSF diagnostics via a combination of fluorescence staining and micro-Raman spectroscopy

This contribution provides a new approach for single blood cell analysis in cerebrospinal fluid (CSF) with the possibility of utilizing simultaneously on the same sample the unique capabilities of the two methods fluorescence staining and Raman spectroscopy. By doing so this technique enables the potential of accurate and rapid cell identification in order to determine cell parameters immediately (e.g. the study of the level of activation or phagocytosis activity of single blood cells). Fluorescence labeling of blood cells offers the great possibility of differentiating easily between the subtypes of white blood cells, while Raman spectroscopy reveals molecular fingerprint information with a spatial resolution down to the diffraction limit. Compared to an unstained cell, the presented results nicely demonstrate that the selected fluorescence dye does not influence the Raman spectrum of a labeled blood cell notably. By the combined application of Raman spectroscopy and statistical data analysis a distinction between white blood cell substructures could be performed. Since several blood cell types also differ in the amount of their cell components, differentiation between several blood cell types is also possible when one blood cell is described in the database by several Raman spectra according their presented sub-microscopic structures. This capability with the possibility of accurate and rapid blood cell identification in cerebrospinal fluid is extremely promising for implementation in clinical diagnostics.