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Mesenchymal stem cells (MSCs) are very attractive for regenerative medicine due to their relatively easy derivation and broad range of differentiation capabilities, either naturally or induced through cell engineering. However, efficient methods of delivery to diseased tissues and the long-term survival of grafted cells still need improvement. Here, we review genetic engineering approaches designed to enhance the migratory capacities of MSCs, as well as extend their survival after transplantation by the modulation of prosurvival approaches, including prevention of senescence and apoptosis. We highlight some of the latest examples that explore these pivotal points, which have great relevance in cell-based therapies.

Myelin insulates neuronal axons and enables fast signal transmission, constituting a key component of brain development, aging and disease. Yet, myelin-specific imaging of macroscopic samples remains a challenge. Here, we exploit myelin’s nanostructural periodicity, and use small-angle X-ray scattering tensor tomography (SAXS-TT) to simultaneously quantify myelin levels, nanostructural integrity and axon orientations in nervous tissue. Proof-of-principle is demonstrated in whole mouse brain, mouse spinal cord and human white and gray matter samples. Outcomes are validated by 2D/3D histology and compared to MRI measurements sensitive to myelin and axon orientations. Specificity to nanostructure is exemplified by concomitantly imaging different myelin types with distinct periodicities. Finally, we illustrate the method’s sensitivity towards myelin-related diseases by quantifying myelin alterations in dysmyelinated mouse brain. This non-destructive, stain-free molecular imaging approach enables quantitative studies of myelination within and across samples during development, aging, disease and treatment, and is applicable to other ordered biomolecules or nanostructures. Small-angle X-ray scattering (SAXS) combines the high tissue penetration of X-rays with specificity to periodic nanostructures. The authors use SAXS tensor tomography (SAXS-TT) on intact mouse and human brain tissue samples, to quantify myelin levels and determine myelin integrity, myelinated axon orientation, and fibre tracts non-destructively.

The blood–brain barrier (BBB) is the main obstacle to the effective delivery of therapeutic agents to the brain, compromising treatment efficacy for a variety of neurological disorders. Intra-arterial (IA) injection of hyperosmotic mannitol has been used to permeabilize the BBB and improve parenchymal entry of therapeutic agents following IA delivery in preclinical and clinical studies. However, the reproducibility of IA BBB manipulation is low and therapeutic outcomes are variable. We demonstrated that this variability could be highly reduced or eliminated when the procedure of osmotic BBB opening is performed under the guidance of interventional MRI. Studies have reported the utility and applicability of this technique in several species. Here we describe a protocol to open the BBB by IA injection of hyperosmotic mannitol under the guidance of MRI in mice. The procedures (from preoperative preparation to postoperative care) can be completed within ~1.5 h, and the skill level required is on par with the induction of middle cerebral artery occlusion in small animals. This MRI-guided BBB opening technique in mice can be utilized to study the biology of the BBB and improve the delivery of various therapeutic agents to the brain. This protocol describes a method for blood–brain barrier opening (BBBO) under the guidance of interventional MRI in mice. The method, which considerably reduces BBBO variability, can be used to improve delivery of therapeutics to the brain.

Studies using large animal models are essential for better understanding the molecular processes underlying neurological diseases, including ischemic stroke, and serve as a robust foundation for evaluating potential therapies. To better understand the complex role of damage-associated molecular pattern molecules (DAMPs) after ischemia, we aimed to determine their expression in the porcine brain affected by ischemic stroke at four time points: 6 h, 24 h, 3 days and 7 days post-stroke. Within the first 24 h after the stroke, we observed the increased expression of several key factors, including calcium-binding proteins, peroxiredoxins, heat shock proteins and interleukins (1α and 1β, IL10, IL17α). Moreover, by day 7, multiple DAMPs were up-regulated, coinciding with an enhanced expression of vascular endothelial growth factor A (VEGFA) in the affected hemisphere. The effects of ischemic stroke were also evident systemically, as indicated by the altered serum levels of both pro- and anti-inflammatory interleukins, reflecting dynamic inflammatory response. To conclude, our findings provide new insights about the time-dependent DAMP activity in a large animal model of ischemic stroke, highlighting the simultaneous occurrence of an ongoing inflammatory response and the possible initiation of vascular remodeling as early as one week after stroke onset.

Hydrogels are one of the most attractive biomaterials, used in both three-dimensional (3D) and in vivo cultures. They facilitate the reconstruction of tissue microenvironments by preserving the spatial arrangement of cells, cell–cell interactions, and functional dynamics in the tissue. In this work, the long-term effect of alginate hydrogel on cell culture and the possibility of rapid cell recovery by dissolving the hydrogel were investigated. Mouse glial-restricted progenitors (GRPs) and porcine mesenchymal stem cells (MSCs) were suspended in hydrogels; their metabolic activity, viability, and expression of genes, which are involved in oxidative stress, apoptosis, proliferation, migration, and differentiation, were assessed using quantitative polymerase chain reaction (qPCR). The concentration that was able to dissolve the hydrogel and was the least harmful to the cells was 0.005 M ethylenediaminetetraacetic acid (EDTA). The metabolism of both cell types was reduced from the beginning of the experiment to day 3. From day 7 to the end of the experiment, the normalization of the GRP metabolism was observed, in contrast to the MSCs. For the apoptosis-related genes, caspase 3, 7, and B-cell leukemia (Casp3, Casp 7, Bcl2) were increased in GRPs and MSCs on days 0 and 1. After 3 and 7 days, an increase in the expression of oxidative stress genes (nuclear factor of activated T-cells 5—NFAT5 and autophagy-related 14-ATG14) was observed in cells cultured in calcium chloride (CaCl2). GRPs cultured in calcium alginate (CaM) were not affected and, remarkably, showed increased Antigen Kiel 67 (Ki67) levels after 30 days. In conclusion, alginate hydrogels provide an excellent environment for stem cell culture in 3D for a longer period of time, but this is dependent on the cell type. Therefore, an individual approach to cell culture is necessary, taking into account the requirements of the cells to be used.

Stem cell transplantation proved promising in animal models of neurological diseases; however, in conditions with disseminated pathology such as ALS, delivery of cells and their broad distribution is challenging. To address this problem, we explored intra-arterial (IA) delivery route, of stem cells. The goal of this study was to investigate the feasibility and safety of MRI-guided transplantation of glial restricted precursors (GRPs) and mesenchymal stem cells (MSCs) in dogs suffering from ALS-like disease, degenerative myelopathy (DM). Canine GRP transplantation in dogs resulted in rather poor retention in the brain, so MSCs were used in subsequent experiments. To evaluate the safety of MSC intraarterial transplantation, naïve pigs (n = 3) were used as a pre-treatment control before transplantation in dogs. Cells were labeled with iron oxide nanoparticles. For IA transplantation a 1.2-French microcatheter was advanced into the middle cerebral artery under roadmap guidance. Then, the cells were transplanted under real-time MRI with the acquisition of dynamic T2*-weighted images. The procedure in pigs has proven to be safe and histopathology has demonstrated the successful and predictable placement of transplanted porcine MSCs. Transplantation of canine MSCs in DM dogs resulted in their accumulation in the brain. Interventional and follow-up MRI proved the procedure was feasible and safe. Analysis of gene expression after transplantation revealed a reduction of inflammatory factors, which may indicate a promising therapeutic strategy in the treatment of neurodegenerative diseases.

Many surgeries are complicated by the need to anastomose, or reconnect, micrometre-scale vessels. Although suturing remains the gold standard for anastomosing vessels, it is difficult to place sutures correctly through collapsed lumen, making the procedure prone to failure. Here, we report a multiphase transitioning peptide hydrogel that can be injected into the lumen of vessels to facilitate suturing. The peptide, which contains a photocaged glutamic acid, forms a solid-like gel in a syringe and can be shear-thin delivered to the lumen of collapsed vessels (where it distends the vessel) and the space between two vessels (where it is used to approximate the vessel ends). Suturing is performed directly through the gel. Light is used to initiate the final gel–sol phase transition that disrupts the hydrogel network, allowing the gel to be removed and blood flow to resume. This gel adds a new tool to the armamentarium for micro- and supermicrosurgical procedures. Suturing of ultrasmall blood vessels is now simplified through the use of a hydrogel that can act as a temporary stent on injection and can be removed through light irradiation.

Addressing multiple neurodegenerative mechanisms may be a more viable strategy. [...]the restoration of lost brain tissue turned out to be a very complex endeavor. Since many of the mechanistic investigations reviewed were conducted in animal models of Alzheimer's disease, the translatability of these findings to human patients remains to be shown. [...]the role of Tc17 cells in multiple sclerosis needs to be elucidated in future translational studies.

Development of a novel, animal model for multiple sclerosis (MS) with reproducible and predictable lesion placement would enhance the discovery of effective treatments. Therefore, we would like to combine the advantages of the demyelination model with experimental autoimmune encephalomyelitis (EAE) to provide a local autoimmune encephalomyelitis (LAE) inside rat brain. We induced a demyelinating lesion by immunizing male Wistar rats, followed by blood-brain barrier opening protein (vascular endothelial growth factor) by stereotactic injection. We confirmed the immunization against myelin epitopes and minor neurological impairment. Histological assessment confirmed the lesion development after both 3- and 7 days post-injection. Our approach was sufficient to develop a demyelinating lesion with high reproducibility and low morbidity.