Bone Marrow-Derived Mononuclear Cells in the Treatment of Neurological Diseases: Knowns and Unknowns.

Bone marrow-derived mononuclear cells (BMMNCs) have been used for decades in preclinical and clinical studies to treat various neurological diseases. However, there is still a knowledge gap in the understanding of the underlying mechanisms of BMMNCs in the treatment of neurological diseases. In addition, prerequisite factors for the efficacy of BMMNC administration, such as the optimal route, dose, and number of administrations, remain unclear. In this review, we discuss known and unknown aspects of BMMNCs, including the cell harvesting, administration route and dose; mechanisms of action; and their applications in neurological diseases, including stroke, cerebral palsy, spinal cord injury, traumatic brain injury, amyotrophic lateral sclerosis, autism spectrum disorder, and epilepsy. Furthermore, recommendations on indications for BMMNC administration and the advantages and limitations of BMMNC applications for neurological diseases are discussed. BMMNCs in the treatment of neurological diseases. BMMNCs have been applied in several neurological diseases. Proposed mechanisms for the action of BMMNCs include homing, differentiation and paracrine effects (angiogenesis, neuroprotection, and anti-inflammation). Further studies should be performed to determine the optimal cell dose and administration route, the roles of BMMNC subtypes, and the indications for the use of BMMNCs in neurological conditions with and without genetic abnormalities.

Chemical Conformation Induced Transport Carrier Switching in Molecular Junction based on Carboxylic-Terminated Thiol Molecules.

The paper demonstrates the effect of the chemical conformation of the -COOH group on the transport characteristic including conductance, rectification, and length effect in molecular junctions (MJs) formed by self-assembled monolayers of carboxylic-terminated thiol molecules. For an alkyl chain shorter than C11, the transport mechanism was attributed to a direct off-resonant tunneling of a hole carrier, located at the Au-S interface, whereas a hopping mechanism was assigned to the alkyl chain longer than the C11 chain located at the -COOH group. The hopping mechanism may be operated by electron transport associated with the breaking of the -OH bonding likely driven by a voltage. Importantly, at the C11 alkyl chain, we observed that the transport carrier operating in MJs could change from a hole carrier into an electron carrier. The result strongly proves that the chemical conformation should be considered in analyzing molecular electronics and provides a basis for the rational design of molecular electronic devices.

Discovery of a SHP2 Degrader with In Vivo Anti-Tumor Activity.

Src homology 2 domain-containing phosphatase 2 (SHP2) is an attractive target for cancer therapy due to its multifaceted roles in both tumor and immune cells. Herein, we designed and synthesized a novel series of proteolysis targeting chimeras (PROTACs) using a SHP2 allosteric inhibitor as warhead, with the goal of achieving SHP2 degradation both inside the cell and in vivo. Among these molecules, compound P9 induces efficient degradation of SHP2 (DC50 = 35.2 ± 1.5 nM) in a concentration- and time-dependent manner. Mechanistic investigation illustrates that the P9-mediated SHP2 degradation requires the recruitment of the E3 ligase and is ubiquitination- and proteasome-dependent. P9 shows improved anti-tumor activity in a number of cancer cell lines over its parent allosteric inhibitor. Importantly, administration of P9 leads to a nearly complete tumor regression in a xenograft mouse model, as a result of robust SHP2 depletion and suppression of phospho-ERK1/2 in the tumor. Hence, P9 represents the first SHP2 PROTAC molecule with excellent in vivo efficacy. It is anticipated that P9 could serve not only as a new chemical tool to interrogate SHP2 biology but also as a starting point for the development of novel therapeutics targeting SHP2.

Religiosity and Social Support Predict Resilience in Older Adults After a Flood.

In this study, we examined religiosity and social support as predictors of resilience after a devastating flood. Three flood exposure groups of primarily middle-aged and older adults were compared: (1) non-flooded adults as controls, (2) once-flooded adults with structural damage to homes and property in the 2016 flood, and (3) twice-flooded adults who had relocated inland because of prior catastrophic losses in the 2005 Hurricanes Katrina and Rita and then flooded again in 2016. Resilience was assessed using the Connor-Davidson Resilience Scale (CD-RISC). Correlation analyses confirmed that older age was correlated with higher religiosity, charitable work done for others, and resilience. Regression analyses indicated that religious beliefs and coping, social support, and charitable work done for others were associated with higher levels of resilience, whereas flood damage was unrelated to resilience. Implications for current views on post-disaster adversity and resilience in later life are discussed.

Small Molecule Degraders of Protein Tyrosine Phosphatase 1B and T-Cell Protein Tyrosine Phosphatase for Cancer Immunotherapy.

Protein tyrosine phosphatase 1B (PTP1B) and T-cell protein tyrosine phosphatase (TC-PTP) play non-redundant negative regulatory roles in T-cell activation, tumor antigen presentation, insulin and leptin signaling, and are potential targets for several therapeutic applications. Here, we report the development of a highly potent and selective small molecule degrader DU-14 for both PTP1B and TC-PTP. DU-14 mediated PTP1B and TC-PTP degradation requires both target protein(s) and VHL E3 ligase engagement and is also ubiquitination- and proteasome-dependent. DU-14 enhances IFN-γ induced JAK1/2-STAT1 pathway activation and promotes MHC-I expression in tumor cells. DU-14 also activates CD8+ T-cells and augments STAT1 and STAT5 phosphorylation. Importantly, DU-14 induces PTP1B and TC-PTP degradation in vivo and suppresses MC38 syngeneic tumor growth. The results indicate that DU-14, as the first PTP1B and TC-PTP dual degrader, merits further development for treating cancer and other indications.

Establishing pre-analytical requirements and maximizing peptide recovery in the analytical phase for mass spectrometric quantification of amyloid-ß peptides 1-42 and 1-40 in CSF.

OBJECTIVES: Amyloid-ß (Aß) peptides in cerebrospinal fluid (CSF), including Aß42 (residues 1-42) and Aß40 (residues 1-40), are utilized as biomarkers in the diagnostic workup of Alzheimer's disease. Careful consideration has been given to the pre-analytical and analytical factors associated with measurement of these peptides via immunoassays; however, far less information is available for mass spectrometric methods. As such, we performed a comprehensive evaluation of pre-analytical and analytical factors specific to Aß quantification using mass spectrometry. METHODS: Using our quantitative mass spectrometry assay for Aß42 and Aß40 in CSF, we investigated the potential for interference from hemolysate, bilirubin, lipids, and anti-Aß-antibodies. We also optimized the composition of the calibrator surrogate matrix and Aß recovery during and after solid phase extraction (SPE). RESULTS: There was no interreference observed with total protein up to 12 g/L, hemolysate up to 10% (v/v), bilirubin up to 0.5% (v/v), intralipid up to 1% (v/v), or anti-Aß-antibodies at expected therapeutic concentrations. For hemolysate, bilirubin and lipids, visual CSF contamination thresholds were established. In the analytical phase, Aß recovery was increased by ∼50% via SPE solvent modifications and by over 150% via modification of the SPE collection plate, which also extended analyte stability in the autosampler. CONCLUSIONS: Attention to mass spectrometric-specific pre-analytical and analytical considerations improved analytical sensitivity and reproducibility, as well as, established CSF specimen acceptance and rejection criteria for use by the clinical laboratory.

Hopping Conductance in Molecular Wires Exhibits a Large Heavy-Atom Kinetic Isotope Effect.

We report a large kinetic isotope effect (KIE) for intramolecular charge transport in π-conjugated oligophenyleneimine (OPI) molecules connected to Au electrodes. 13C and 15N substitution on the imine bonds produces a conductance KIE of ∼2.7 per labeled atom in long OPI wires >4 nm in length, far larger than typical heavy-atom KIEs for chemical reactions. In contrast, isotopic labeling in shorter OPI wires <4 nm does not produce a conductance KIE, consistent with a direct tunneling mechanism. Temperature-dependent measurements reveal that conductance for a long 15N-substituted OPI wire is activated, and we propose that the exceptionally large conductance KIEs imply a thermally assisted, through-barrier polaron tunneling mechanism. In general, observation of large conductance KIEs opens up considerable opportunities for understanding microscopic conduction mechanisms in π-conjugated molecules.

Reproducibility and accuracy of microscale thermophoresis in the NanoTemper Monolith: a multi laboratory benchmark study.

Microscale thermophoresis (MST), and the closely related Temperature Related Intensity Change (TRIC), are synonyms for a recently developed measurement technique in the field of biophysics to quantify biomolecular interactions, using the (capillary-based) NanoTemper Monolith and (multiwell plate-based) Dianthus instruments. Although this technique has been extensively used within the scientific community due to its low sample consumption, ease of use, and ubiquitous applicability, MST/TRIC has not enjoyed the unambiguous acceptance from biophysicists afforded to other biophysical techniques like isothermal titration calorimetry (ITC) or surface plasmon resonance (SPR). This might be attributed to several facts, e.g., that various (not fully understood) effects are contributing to the signal, that the technique is licensed to only a single instrument developer, NanoTemper Technology, and that its reliability and reproducibility have never been tested independently and systematically. Thus, a working group of ARBRE-MOBIEU has set up a benchmark study on MST/TRIC to assess this technique as a method to characterize biomolecular interactions. Here we present the results of this study involving 32 scientific groups within Europe and two groups from the US, carrying out experiments on 40 Monolith instruments, employing a standard operation procedure and centrally prepared samples. A protein-small molecule interaction, a newly developed protein-protein interaction system and a pure dye were used as test systems. We characterized the instrument properties and evaluated instrument performance, reproducibility, the effect of different analysis tools, the influence of the experimenter during data analysis, and thus the overall reliability of this method.

The Chimeric Scapulodorsal Vascularized Latissimus Dorsi Nerve Flap for Immediate Reconstruction of Total Parotidectomy Defects With Facial Nerve Sacrifice: Building a New Program and Preliminary Results From 25 Cases.

BACKGROUND: Total parotidectomy with facial nerve sacrifice creates 2 challenging reconstructive problems: restoration of facial contour and facial nerve rehabilitation. Strong evidence suggesting that vascularized nerve grafts are superior to nonvascularized nerve grafts motivated our team to develop a chimeric scapulodorsal flap combining the usual harvestable local tissues with the vascularized latissimus dorsi motor nerve (SD-LDVxN). We present our experiences developing a new program at University of California, San Diego, highlighting our first case here, and present preliminary retrospective results focusing on the functional outcomes of facial nerve reanimation. MATERIALS AND METHODS: The first case performed in the United States was a 57-year-old woman with stage IVA left parotid adenoid cystic carcinoma and House-Brackmann grade 6 facial palsy. She underwent total parotidectomy with facial nerve sacrifice and a free chimeric SD-LDVxN flap reconstruction. She had an unremarkable postoperative course, and 3- and 6-month follow-up functional results are reported. Preliminary functional results from our total series of 25 patients were reported. RESULTS: At her 3-month follow-up, she was a House-Brackmann 5 with a static eFACE score of 37, dynamic eFACE score of 31, and smile eFACE score of 48. At her 6-month follow-up, she was a House-Brackmann 5 with a static eFACE score of 50, dynamic eFACE score of 27, and smile eFACE score of 53. Preliminary results from our total series of 25 patients with an average of 5 years of follow-up were a House-Brackmann 2.5 and eFACE scores of 83.1 for static facial symmetry, 67.5 for dynamic facial symmetry, and 77.7 for smile score. Twenty of the 25 patients had postoperative radiotherapy. No local tumor recurrence had been reported. The average reinnervation time was 9 months and ranged from 3 to 15 months. CONCLUSIONS: The SD-LDVxN flap is a highly resourceful solution to reconstruct complex parotid defects, especially those that sacrifice the facial nerve. The vascularized nerve graft allows for primary facial reanimation. Nerve recovery may be superior to what could be expected with a conventional nerve graft.

Fluorescence Imaging of Nerves During Surgery.

OBJECTIVE: This review details the agents for fluorescence-guided nerve imaging in both preclinical and clinical use to identify factors important in selecting nerve-specific fluorescent agents for surgical procedures. BACKGROUND: Iatrogenic nerve injury remains a significant cause of morbidity in patients undergoing surgical procedures. Current real-time identification of nerves during surgery involves neurophysiologic nerve stimulation, which has practical limitations. Intraoperative fluorescence-guided imaging provides a complimentary means of differentiating tissue types and pathology. Recent advances in fluorescence-guided nerve imaging have shown promise, but the ideal agent remains elusive. METHODS: In February 2018, PubMed was searched for articles investigating peripheral nerve fluorescence. Key terms used in this search include: "intraoperative, nerve, fluorescence, peripheral nerve, visualization, near infrared, and myelin." Limits were set to exclude articles exclusively dealing with central nervous system targets or written in languages other than English. References were cross-checked for articles not otherwise identified. RESULTS: Of the nonspecific agents, tracers that rely on axonal transport showed the greatest tissue specificity; however, neurovascular dyes already enjoy wide clinical use. Fluorophores specific to nerve moieties result in excellent nerve to background ratios. Although noteworthy findings on tissue specificity, toxicity, and route of administration specific to each fluorescent agent were reported, significant data objectively quantifying nerve-specific fluorescence and toxicity are lacking. CONCLUSIONS: Fluorescence-based nerve enhancement has advanced rapidly over the past 10 years with potential for continued utilization and progression in translational research. An ideal agent would be easily administered perioperatively, would not cross the blood-brain barrier, and would fluoresce in the near-infrared spectrum. Agents administered systemically that target nerve-specific moieties have shown the greatest promise. Based on the heterogeneity of published studies and methods for reporting outcomes, it appears that the development of an optimal nerve imaging agent remains challenging.

FAIRDOMHub: a repository and collaboration environment for sharing systems biology research.

The FAIRDOMHub is a repository for publishing FAIR (Findable, Accessible, Interoperable and Reusable) Data, Operating procedures and Models (https://fairdomhub.org/) for the Systems Biology community. It is a web-accessible repository for storing and sharing systems biology research assets. It enables researchers to organize, share and publish data, models and protocols, interlink them in the context of the systems biology investigations that produced them, and to interrogate them via API interfaces. By using the FAIRDOMHub, researchers can achieve more effective exchange with geographically distributed collaborators during projects, ensure results are sustained and preserved and generate reproducible publications that adhere to the FAIR guiding principles of data stewardship.

Laminin targeting of a peripheral nerve-highlighting peptide enables degenerated nerve visualization.

Target-blind activity-based screening of molecular libraries is often used to develop first-generation compounds, but subsequent target identification is rate-limiting to developing improved agents with higher specific affinity and lower off-target binding. A fluorescently labeled nerve-binding peptide, NP41, selected by phage display, highlights peripheral nerves in vivo. Nerve highlighting has the potential to improve surgical outcomes by facilitating intraoperative nerve identification, reducing accidental nerve transection, and facilitating repair of damaged nerves. To enable screening of molecular target-specific molecules for higher nerve contrast and to identify potential toxicities, NP41's binding target was sought. Laminin-421 and -211 were identified by proximity-based labeling using singlet oxygen and by an adapted version of TRICEPS-based ligand-receptor capture to identify glycoprotein receptors via ligand cross-linking. In proximity labeling, photooxidation of a ligand-conjugated singlet oxygen generator is coupled to chemical labeling of locally oxidized residues. Photooxidation of methylene blue-NP41-bound nerves, followed by biotin hydrazide labeling and purification, resulted in light-induced enrichment of laminin subunits α4 and α2, nidogen 1, and decorin (FDR-adjusted P value < 10-7) and minor enrichment of laminin-γ1 and collagens I and VI. Glycoprotein receptor capture also identified laminin-α4 and -γ1. Laminins colocalized with NP41 within nerve sheath, particularly perineurium, where laminin-421 is predominant. Binding assays with phage expressing NP41 confirmed binding to purified laminin-421, laminin-211, and laminin-α4. Affinity for these extracellular matrix proteins explains the striking ability of NP41 to highlight degenerated nerve "ghosts" months posttransection that are invisible to the unaided eye but retain hollow laminin-rich tubular structures.

Silica Coated Iron/Iron Oxide Nanoparticles as a Nano-Platform for T2 Weighted Magnetic Resonance Imaging.

The growing concern over the toxicity of Gd-based contrast agents used in magnetic resonance imaging (MRI) motivates the search for less toxic and more effective alternatives. Among these alternatives, iron-iron oxide (Fe@FeOx) core-shell architectures have been long recognized as promising MRI contrast agents while limited information on their engineering is available. Here we report the synthesis of 10 nm large Fe@FeOx nanoparticles, their coating with a 11 nm thick layer of dense silica and functionalization by 5 kDa PEG chains to improve their biocompatibility. The nanomaterials obtained have been characterized by a set of complementary techniques such as infra-red and nuclear magnetic resonance spectroscopies, transmission electron microscopy, dynamic light scattering and zetametry, and magnetometry. They display hydrodynamic diameters in the 100 nm range, zetapotential values around -30 mV, and magnetization values higher than the reference contrast agent RESOVIST®. They display no cytotoxicity against 1BR3G and HCT116 cell lines and no hemolytic activity against human red blood cells. Their nuclear magnetic relaxation dispersion (NMRD) profiles are typical for nanomaterials of this size and magnetization. They display high r2 relaxivity values and low r1 leading to enhanced r2/r1 ratios in comparison with RESOVIST®. All these data make them promising contrast agents to detect early stage tumors.

Highly Efficient Long-Range Electron Transport in a Viologen-Based Molecular Junction.

Thin layers of viologen-based oligomers with thicknesses between 3 and 14 nm were deposited on gold electrodes by electrochemical reduction of a diazonium salt, and then a Ti/Au top contact was applied to complete a solid-state molecular junction (MJ). MJs show symmetric J- V curves and highly efficient long-range transport, with an attenuation factor as small as 0.25 nm-1. This is attributed both to the fact that the viologen LUMO energy lies between the energies of the Fermi levels of the two contacts and to strong electronic coupling between molecules and contacts. As a consequence, resonant tunneling is likely to be the dominant transport mechanism within these MJs, but the temperature dependence of the transport properties suggests that activated redox hopping plays a role at high temperature.

Visualization of macrophage recruitment in head and neck carcinoma model using fluorine-19 magnetic resonance imaging.

PURPOSE: To evaluate the role of infiltrating macrophages in murine models of single and double mutation head and neck tumors using a novel fluorine-19 (19 F) MRI technology. METHODS: Tumor cell lines single-hit/SCC4 or double-hit/Cal27, with mutations of TP53 and TP53 & FHIT, respectively, were injected bilaterally into the flanks of (n = 10) female mice. With tumors established, perfluorocarbon nanoemulsion was injected intravenously, which labels in situ predominantly monocytes and macrophages. Longitudinal spin density-weighted 19 F MRI data enabled quantification of the macrophage burden in tumor and surrounding tissue. RESULTS: The average number of 19 F atoms within the tumors was twice as high in the Cal27 group compared with SCC4 (3.9 × 1019 and 2.0 × 101919 F/tumor, respectively; P = 0.0034) two days after contrast injection, signifying increased tumor-associated macrophages in double-hit tumors. The difference was still significant 10 days after injection. Histology stains correlated with in vivo results, exhibiting numerous perfluorocarbon-labeled macrophages in double-hit tumors and to a lesser extent in single-hit tumors. CONCLUSIONS: This study helps to establish 19 F MRI as a method for quantifying immune cells in the tumor microenvironment, allowing distinction between double and single-hit head and neck tumors. This technique would be extremely valuable in the clinic for pretreatment planning, prognostics, and post-treatment surveillance. Magn Reson Med 79:1972-1980, 2018. © 2017 International Society for Magnetic Resonance in Medicine.

Rapid continuous 3D printing of customizable peripheral nerve guidance conduits.

Engineered nerve guidance conduits (NGCs) have been demonstrated for repairing peripheral nerve injuries. However, there remains a need for an advanced biofabrication system to build NGCs with complex architectures, tunable material properties, and customizable geometrical control. Here, a rapid continuous 3D-printing platform was developed to print customizable NGCs with unprecedented resolution, speed, flexibility, and scalability. A variety of NGC designs varying in complexity and size were created including a life-size biomimetic branched human facial NGC. In vivo implantation of NGCs with microchannels into complete sciatic nerve transections of mouse models demonstrated the effective directional guidance of regenerating sciatic nerves via branching into the microchannels and extending toward the distal end of the injury site. Histological staining and immunostaining further confirmed the progressive directional nerve regeneration and branching behavior across the entire NGC length. Observational and functional tests, including the von Frey threshold test and thermal test, showed promising recovery of motor function and sensation in the ipsilateral limbs grafted with the 3D-printed NGCs.

Robust Bipolar Light Emission and Charge Transport in Symmetric Molecular Junctions.

Molecular junctions consisting of a Ru(bpy)3 oligomer between conducting carbon contacts exhibit an exponential dependence of junction current on molecular layer thickness (d) similar to that observed for other aromatic devices when d < 4 nm. However, when d > 4 nm, a change in transport mechanism occurs which coincides with light emission in the range of 600-900 nm. Unlike light emission from electrochemical cells or solid-state films containing Ru(bpy)3, emission is bipolar, occurs in vacuum, has rapid rise time (<5 ms), and persists for >10 h. Light emission directly indicates simultaneous hole and electron injection and transport, possibly resonant due to the high electric field present (>3 MV/cm). Transport differs fundamentally from previous tunneling and hopping mechanisms and is a clear "molecular signature" relating molecular structure to electronic behavior.

Control of Rectification in Molecular Junctions: Contact Effects and Molecular Signature.

Thin layers of oligomers with thickness between 7 and 9 nm were deposited on flat gold electrode surfaces by electrochemical reduction of diazonium reagents, then a Ti(2 nm)/Au top contact was applied to complete a solid-state molecular junction. The molecular layers investigated included donor molecules with relatively high energy HOMO, molecules with high HOMO-LUMO gaps and acceptor molecules with low energy LUMO and terminal alkyl chain. Using an oligo(bisthienylbenzene) based layer, a molecule whose HOMO energy level in a vacuum is close to the Fermi level of the gold bottom electrode, the devices exhibit robust and highly reproducible rectification ratios above 1000 at low voltage (2.7 V). Higher current is observed when the bottom gold electrode is biased positively. When the molecular layer is based on a molecule with a high HOMO-LUMO gap, i.e., tetrafluorobenzene, no rectification is observed, while the direction of rectification is reversed if the molecular layer consists of naphtalene diimides having low LUMO energy level. Rectification persisted at low temperature (7 K), and was activationless between 7 and 100 K. The results show that rectification is induced by the asymmetric contact but is also directly affected by orbital energies of the molecular layer. A "molecular signature" on transport through layers with thicknesses above those used when direct tunneling dominates is thus clearly observed, and the rectification mechanism is discussed in terms of Fermi level pinning and electronic coupling between molecules and contacts.

Expression in grasses of multiple transgenes for degradation of munitions compounds on live-fire training ranges.

The deposition of toxic munitions compounds, such as hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine (RDX), on soils around targets in live-fire training ranges is an important source of groundwater contamination. Plants take up RDX but do not significantly degrade it. Reported here is the transformation of two perennial grass species, switchgrass (Panicum virgatum) and creeping bentgrass (Agrostis stolonifera), with the genes for degradation of RDX. These species possess a number of agronomic traits making them well equipped for the uptake and removal of RDX from root zone leachates. Transformation vectors were constructed with xplA and xplB, which confer the ability to degrade RDX, and nfsI, which encodes a nitroreductase for the detoxification of the co-contaminating explosive 2, 4, 6-trinitrotoluene (TNT). The vectors were transformed into the grass species using Agrobacterium tumefaciens infection. All transformed grass lines showing high transgene expression levels removed significantly more RDX from hydroponic solutions and retained significantly less RDX in their leaf tissues than wild-type plants. Soil columns planted with the best-performing switchgrass line were able to prevent leaching of RDX through a 0.5-m root zone. These plants represent a promising plant biotechnology to sustainably remove RDX from training range soil, thus preventing contamination of groundwater.