Hydrogels as delivery systems for spinal cord injury regeneration.

Spinal cord injury is extremely debilitating, both at physiological and psychological levels, changing completely the patient's lifestyle. The introduction of biomaterials has opened a new window to develop a therapeutic approach to induce regeneration after injury due to similarities with extracellular matrix. Particularly, hydrogels have the ability to support axonal growth and endogenous regeneration. Moreover, they can also act as potential matrixes in which to load and deliver therapeutic agents at injury site. In this review, we highlight some important characteristics to be considered when designing hydrogels as delivery systems (DS), such as rheology, mesh size, swelling, degradation, gelation temperature and surface charge. Additionally, affinity-based release systems, incorporation of nanoparticles, or ion-mediated interactions are also pondered. Overall, hydrogel DS aim to promote a sustained, controlled and prolonged release at injury site, allowing a targeted oriented action of the therapeutic agent that will be used.

Unilateral accumbal dopamine depletion affects decision-making in a side-specific manner.

Mechanisms underlying affective and cognitive deficits in Parkinson's disease (PD) remain less studied than motor symptoms. Nucleus accumbens (NAc) is affected in PD and due to its well-known involvement in motivation is an interesting target in this context. Furthermore, PD is frequently asymmetrical, with side-specific deficits aligning with evidences of accumbal laterality. We therefore used a 6-hydroxydopamine (6-OHDA) model to study the role of left and right NAc dopamine depletion in a battery of behavioral tasks. 2 months old male rats were used in all experiments. Habitual-based and goal-directed decision-making, impulsivity, anxiety- and depressive-like behavior and motor performance were tested 3 weeks after left (6-OHDA L) or right (6-OHDA R) NAc lesion was induced. Upon contingency degradation, 6-OHDA R decrease their lever press rate less than Sham and 6-OHDA L, indicating an impairment in the shift from habit-based to goal-directed strategies. On the other hand, 6-OHDA L lesions lead to increased rates of premature responding when delays where increased in the variable delay-to-signal test. Importantly, in both paradigms task acquisition was similar between groups. In the same line we found no differences in the amount of sugared pellets eaten when freely available as well as in both general and fine motor behaviors. In conclusion, left and right NAc play distinct roles in the contingency degradation and impulsivity. More studies are needed to understand the mechanisms behind this functional lateralization and its implications for PD patients.

The impact of Mesenchymal Stem Cells and their secretome as a treatment for gliomas.

In recent years, we have witnessed a significant increase in the amount of studies using Mesenchymal Stem Cells (MSCs) for cancer therapy, mostly as vectors for drug or gene delivery strategies. This is because of their intrinsic capacity of homing into tumor niches. However, the interactions between MSCs themselves and tumor cells is not fully understood, with contradictory results frequently being observed regarding their effects on cancer cell invasion and proliferation. This poses an important question of safety in respect to the application of these cells. The source of the MSC population used, as well as the type of cancer cells under study might strongly influence this interaction. Moreover, differences in isolation protocols, culture media compositions, time of culture and conditioned media collection, or even timing and mode of MSCs administration to in vivo models of cancer may also affect the interaction MSC-tumor cells. In this review, we drive our focus into malignant brain tumors, particularly gliomas, one of the deadliest forms of cancer. Moreover, we look with some detail into different studies using MSCs as a treatment for brain tumors and compare them, highlighting the main deviations and similarities among them.

Influence of Different ECM-Like Hydrogels on Neurite Outgrowth Induced by Adipose Tissue-Derived Stem Cells.

Mesenchymal stem cells (MSCs) have been proposed for spinal cord injury (SCI) applications due to their capacity to secrete growth factors and vesicles-secretome-that impacts important phenomena in SCI regeneration. To improve MSC survival into SCI sites, hydrogels have been used as transplantation vehicles. Herein, we hypothesized if different hydrogels could interact differently with adipose tissue-derived MSCs (ASCs). The efficacy of three natural hydrogels, gellan gum (functionalized with a fibronectin peptide), collagen, and a hydrogel rich in laminin epitopes (NVR-gel) in promoting neuritogenesis (alone and cocultured with ASCs), was evaluated in the present study. Their impact on ASC survival, metabolic activity, and gene expression was also evaluated. Our results indicated that all hydrogels supported ASC survival and viability, being this more evident for the functionalized GG hydrogels. Moreover, the presence of different ECM-derived biological cues within the hydrogels appears to differently affect the mRNA levels of growth factors involved in neuronal survival, differentiation, and axonal outgrowth. All the hydrogel-based systems supported axonal growth mediated by ASCs, but this effect was more robust in functionalized GG. The data herein presented highlights the importance of biological cues within hydrogel-based biomaterials as possible modulators of ASC secretome and its effects for SCI applications.

Tips on How to Collect and Administer the Mesenchymal Stem Cell Secretome for Central Nervous System Applications.

Human mesenchymal stem cells (hMSCs) have been proposed as possible therapeutic agents for central nervous system (CNS) disorders. Recently, it has been suggested that their effects are mostly mediated through their secretome, which contains a number of neuroregulatory molecules capable of increasing cell proliferation, differentiation, and survival in different physiological conditions. Here, we present an overview of the hMSC secretome as a possible candidate in the creation of new cell-free therapies, demonstrating the process of its collection and route of administration, focusing our attention on their effects in CNS regenerative medicine.

Animal model for chronic massive rotator cuff tear: behavioural and histologic analysis.

PURPOSE: Massive rotator cuff tears (MRCT) are usually chronic lesions that present associated degenerative changes of the myotendinous unit that have been implicated in limitations for surgical repair. In order to develop effective therapies, it is important to establish animal models that mimic the hallmarks of the injury itself. Therefore, in the present work, we aimed to (1) optimize a rodent animal model of MRCT that closely reproduces the fatty infiltration of the cuff muscles seen in humans and (2) describe the effects of unilateral or bilateral lesion in terms of histology and behaviour. METHODS: Massive tear was defined as two rotator cuff tendons-supraspinatus and infraspinatus-section. Twenty-one Wistar rats were randomly assigned to four groups: bilateral lesion (five animals), right-sided unilateral lesion (five animals), left-sided unilateral lesion (five animals) and control (six animals). Behaviour was analyzed with open field and staircase test, 16 weeks after lesion. After that, animals were killed, and the supraspinatus and infraspinatus muscles were processed. RESULTS: Histologic analysis revealed adipocytes, fatty infiltration and atrophy in the injured side with a greater consistency of these degenerative changes in the bilateral lesion group. Behaviour analysis revealed a significant functional impairment of the fine motor control of the forepaw analyzed in staircase test where the number of eaten pellets was significantly higher in sham animals (sham = 7 ± 5.0; left unilateral = 2.6 ± 3.0; right unilateral = 0 ± 0; and bilateral = 0 ± 0, p < 0.05). A trend to reach a lower level of steps, in more injured animals, was also observed (sham animals = 3 ± 1.6 > left unilateral = 2 ± 2.1 > right unilateral = 0.8 ± 1.3 > bilateral = 0.8 ± 1.1). CONCLUSIONS: The present study has been able to establish an animal model that disclosed the hallmarks of MRCT. This can now be used as a valuable, cost-effective, pre-clinical instrument to assist in the development of advanced tissue engineered strategies. Moreover, this animal model overcomes some of the limitations of those that have been reported so far and thus represents a more reliable source for the assessment of future therapeutic strategies with potential clinical relevance.

Unveiling the effects of the secretome of mesenchymal progenitors from the umbilical cord in different neuronal cell populations.

It has been previously shown that the secretome of Human Umbilical Cord Perivascular Cells (HUCPVCs), known for their mesenchymal like stem cell character, is able to increase the metabolic viability and hippocampal neuronal cell densities. However, due to the different micro-environments of the distinct brain regions it is important to study if neurons isolated from different areas have similar, or opposite, reactions when in the presence of HUCPVCs secretome (in the form of conditioned media-CM). In this work we: 1) studied how cortical and cerebellar neuronal primary cultures behaved when incubated with HUCPVCs CM and 2) characterized the differences between CM collected at two different conditioning time points. Primary cultures of cerebellar and cortical neurons were incubated with HUCPVCs CM (obtained 24 and 96 h after three days of culturing). HUCPVCs CM had a higher impact on the metabolic viability and proliferation of cortical cultures, than the cerebellar ones. Regarding neuronal cell densities it was observed that with 24 h CM condition there were higher number MAP-2 positive cells, a marker for fully differentiated neurons; this was, once again, more evident in cortical cultures. In an attempt to characterize the differences between the two conditioning time points a proteomics approach was followed, based on 2D Gel analysis followed by the identification of selected spots by tandem mass spectrometry. Results revealed important differences in proteins that have been previously related with phenomena such as neurl cell viability, proliferation and differentiation, namely 14-3-3, UCHL1, hsp70 and peroxiredoxin-6. In summary, we demonstrated differences on how neurons isolated from different brain regions react to HUCPVCs secretome and we have identified different proteins (14-3-3 and hsp70) in HUCPVCs CM that may explain the above-referred results.

Peripheral mineralization of a 3D biodegradable tubular construct as a way to enhance guidance stabilization in spinal cord injury regeneration.

Spinal cord injuries (SCI) present a major challenge to therapeutic development due to its complexity. Combinatorial approaches using biodegradable polymers that can simultaneously provide a tissue scaffold, a cell vehicle, and a reservoir for sustained drug delivery have shown very promising results. In our previous studies we have developed a novel hybrid system consisting of starch/poly-e-caprolactone (SPCL) semi-rigid tubular porous structure, based on a rapid prototyping technology, filled by a gellan gum hydrogel concentric core for the regeneration within spinal-cord injury sites. In the present work we intend to promote enhanced osteointegration on these systems by pre-mineralizing specifically the external surfaces of the SPCL tubular structures, though a biomimetic strategy, using a sodium silicate gel as nucleating agent. The idea is to create two different cell environments to promote axonal regeneration in the interior of the constructs while inducing osteogenic activity on its external surface. By using a Teflon cylinder to isolate the interior of the scaffold, it was possible to observe the formation of a bone-like poorly crystalline carbonated apatite layer continuously formed only in the external side of the tubular structure. This biomimetic layer was able to support the adhesion of Bone Marrow Mesenchymal Stem Cells, which have gone under cytoskeleton reorganization in the first hours of culture when compared to cells cultured on uncoated scaffolds. This strategy can be a useful route for locally stimulate bone tissue regeneration and facilitating early bone ingrowth.

The secretome of bone marrow mesenchymal stem cells-conditioned media varies with time and drives a distinct effect on mature neurons and glial cells (primary cultures).

Transplantation of bone marrow mesenchymal stem cells (BM-MSCs) has been shown to ameliorate the injured central nervous system (CNS). Although these effects were initially attributed to the putative differentiation of MSCs towards the neural lineage, it is now known that most of them are mediated by the secretome. Up to now most in vitro reports have dealt with the effects of the secretome on neural stem cells and their differentiation. Consequently, there is a lack of information regarding the role of the secretome on the viability and survival of pre-existent matured differentiated cell populations. Moreover, it is also not known how the time points of conditioned media (CM) collection affect such parameters. In the present study, primary cultures of hippocampal neurons and glial cells were incubated with CM obtained from MSCs. To determine how the temporal profiles of CM collection impact on post-natal neurons and glial cells, we collected MSCs CM at 24, 48, 72 and 96 h of conditioning. MTS test revealed that for the hippocampal cultures the incubation with CM increased cell viability for all time points, with significant increases in the percentage of neurons in culture incubated with CM 24 h. For glial cells only the later time point of CM collection (96 h) increased cell viability. Fluorescence microscopy observations also revealed that CM 48 h and 72 h increased astrocytes percentages, while CM 24 h decreased microglial cell and oligodendrocytes values. These results revealed that post-natal neuronal and glial cells respond differently to MSCs CM; moreover, there are specific temporal variations in the composition of the CM of MSCs collected at different time points that trigger different effects on mature neurons and the distinct glial cell populations (astrocytes, oligodendrocytes and microglial cells).

In vivo response to starch-based scaffolds designed for bone tissue engineering applications.

Our purpose was to evaluate the in vivo endosseous response to three starch-based scaffolds implanted in rats (n = 54). We implanted the three scaffold groups; a 50/50 (wt %) blend of corn starch and ethylene-vinyl alcohol (SEVA-C), the same composition coated with a biomimetic calcium phosphate (Ca-P) layer (SEVA-C/CaP), and a 50/50 (wt %) blend of corn starch and cellulose acetate (SCA), all produced by extrusion with blowing agents, into distal femurs proximal to the epiphyseal plate, for 1, 3, or 6 weeks. Our results showed that at 1 week considerable reparative bone formed around all scaffold groups, although the bone was separated from the scaffold by an intervening soft tissue interfacial zone that comprised two distinct compartments: the surface of the scaffold was occupied by multinucleate giant cells and the compartment between these cells and the surrounding bone was occupied by a streaming fibrous-like tissue. The extracellular matrix of the latter was continuous with the extracellular bone matrix itself, labeled positively for osteocalcin and appeared mineralized by back-scattered electron imaging. All three scaffolds showed a similar tissue response, with the soft tissue interface diminishing with time. No bone contact was observed with SEVA-C at any time point, only transitory bone contact was observed with SEVA-C/CaP at 3 weeks, but SCA exhibited direct bone contact at 6 weeks where 56.23 +/- 6.46% of the scaffold surface was occupied by bone. We conclude that all materials exhibited a favorable bony response and that the rapidly forming initial "connective tissue" seen around all scaffolds was a very early form of bone formation.