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Solid Lipid Nanoparticles (SLN) are colloidal drug delivery systems characterized by higher entrapment efficiency, good scalability of the preparation process and increased sustained release of the payload. Surface functionalization of SLN with ligands to achieve a site specific targeting makes them attractive to overcome the limited BloodBrain Barrier (BBB) penetration of therapeutic compounds. SLN are prepared for brain targeting by exploiting the adaptability of warm microemulsion process for the covalent surface modification with an Apolipoprotein Ederived peptide (SLNmApoE). Furthermore, the influence of the administration route on SLNmApoE brain bioavailability is here evaluated by means of Fluorescence Molecular Tomography, an advanced optical imaging technology that uses the NearInfrared Spectrum (NIR) (600–900 nm) for noninvasive in vivo imaging and ThreeDimensional (3D) quantification of the fluorescent probes. Fluorescent labelled SLNmApoE are able to cross intact a BBB in vitro model. The pulmonary administration of SLNmApoE is related to a higher confinement in the brain of Balb/c mice compared to the intravenous and intraperitoneal administration routes, without inducing any acute inflammatory reaction in the lungs. These results promote the pulmonary administration of braintargeted SLN as a feasible strategy for improving brain delivery of therapeutics as well as the FMT’s ability of quantitative assessment in vivobiodistribution studies.

The advent of the new revolutionary approach based on regenerative medicine is progressively reshaping the therapeutic scenario of many different diseases, such as cardiovascular diseases and immune diseases, with encouraging results. During the last 10 years, many studies have also proposed the use of mesenchymal stem cells (MSCs), adult stem cells with several interesting properties already used in different experimental models, for the treatment of diabetes, however, reporting conflicting outcomes. These reasons have given rise to a question: are these cells a real trump card for the biomedical field? Are they really able to outclass the traditional therapies, or at least able to give an advantage ove them? In this review, we will discuss the most promising results obtained with MSCs for the treatment of diabetes and its complications, we will compare the different therapeutic treatments applied as well as the most likely mechanisms of action, and overall we will give an in-depth overview of the pros and the cons of the use of MSCs for the therapy of both type-1 and type-2 diabetes.

In northern Italy, biomass burning-derived (BB) particles and diesel exhaust particles (DEP) are considered the most significant contributors to ultrafine particle (UFP) emission. However, a comparison between their impact on different brain regions was not investigated until now. Therefore, male BALB/c mice were treated with a single or three consecutive intratracheal instillations using 50 µg of UFPs in 100 µL of isotonic saline solution or 100 µL of isotonic saline solution alone, and brains were collected and analyzed. Proteins related to oxidative stress and inflammation, as well as Alzheimer’s disease markers, were examined in the hippocampus, cerebellum, and the rest of the brain (RoB). Histopathological examination of the brain was also performed. Moreover, correlations among different brain, pulmonary, and cardiovascular markers were performed, allowing us to identify the potentially most stressful UFP source. Although both acute exposures induced inflammatory pathways in mouse brain, only DEP showed strong oxidative stress. The sub-acute exposure also induced the modulation of APP and BACE1 protein levels for both UFPs. We observed that DEP exposure is more harmful than BB, and this different response could be explained by this UFP’s different chemical composition and reactivity.

Mesenchymal Stem Cells (MSCs) are adult multipotent cells able to increase sensory neuron survival: direct co-culture of MSCs with neurons is pivotal to observe a neuronal survival increase. Despite the identification of some mechanisms of action, little is known about how MSCs physically interact with neurons. The aim of this paper was to investigate and characterize the main mechanisms of interaction between MSCs and neurons. Morphological analysis showed the presence of gap junctions and tunneling nanotubes between MSCs and neurons only in direct co-cultures. Using a diffusible dye, we observed a flow from MSCs to neurons and further analysis demonstrated that MSCs donated mitochondria to neurons. Treatment of co-cultures with the gap junction blocker Carbenoxolone decreased neuronal survival, thus demonstrating the importance of gap junctions and, more in general, of cell communication for the MSC positive effect. We also investigated the role of extracellular vesicles; administration of direct co-cultures-derived vesicles was able to increase neuronal survival. In conclusion, our study demonstrates the presence and the importance of multiple routes of communication between MSCs and neurons. Such knowledge will allow a better understanding of the potential of MSCs and how to maximize their positive effect, with the final aim to provide the best protective treatment.

We report the rational design, synthesis, and in vitro preliminary evaluation of a new small library of non-peptide ligands of Gastrin Releasing Peptide Receptor (GRP-R), able to antagonize its natural ligand bombesin (BN) in the nanomolar range of concentration. GRP-R is a transmembrane G-protein coupled receptor promoting the stimulation of cancer cell proliferation. Being overexpressed on the surface of different human cancer cell lines, GRP-R is ideal for the selective delivery to tumor cells of both anticancer drug and diagnostic devices. What makes very challenging the design of non-peptide BN analogues is that the 3D structure of the GRP-R is not available, which is the case for many membrane-bound receptors. Thus, the design of GRP-R ligands has to be based on the structure of its natural ligands, BN and GRP. We recently mapped the BN binding epitope by NMR and here we exploited the same spectroscopy, combined with MD, to define BN conformation in proximity of biological membranes, where the interaction with GRP-R takes place. The gained structural information was used to identify a rigid C-galactosidic scaffold able to support pharmacophore groups mimicking the BN key residues’ side chains in a suitable manner for binding to GRP-R. Our BN antagonists represent hit compounds for the rational design and synthesis of new ligands and modulators of GRP-R. The further optimization of the pharmacophore groups will allow to increase the biological activity. Due to their favorable chemical properties and stability, they could be employed for the active receptor-mediated targeting of GRP-R positive tumors.

Exposure to ultrafine particles (UFPs) leads to adverse e_ects on health caused by an unbalanced ratio between UFPs deposition and clearance e_cacy. Since air pollution toxicity is first direct to cardiorespiratory system, we compared the acute and sub-acute e_ects of diesel exhaust particles (DEP) and biomass burning-derived particles (BB) on bronchoalveolar Lavage Fluid (BALf), lung and heart parenchyma. Markers of cytotoxicity, oxidative stress and inflammation were analysed in male BALB/c mice submitted to single and repeated intra-tracheal instillations of 50µg UFPs. This in-vivo study showed the activation of inflammatory response (COX-2 and MPO) after exposure to UFPs, both in respiratory and cardiovascular systems. Exposure to DEP results also in pro- and anti-oxidant (HO-1, iNOS, Cyp1b1, Hsp70) protein levels increase, although, stress persist only in cardiac tissue under repeated instillations. Statistical correlations suggest that stress marker variation was probably due to soluble components and/or mediators translocation of from first deposition site. This mechanism, appears more important after repeated instillations, since inflammation and oxidative stress endure only in heart. In summary, chemical composition of UFPs influenced the activation of di_erent responses mediated by their components or pro-inflammatory and pro-oxidative molecules, indicating DEP as the most damaging pollutant in the comparison.

Many potential therapeutic compounds for brain diseases fail to reach their molecular targets due to the impermeability of the blood-brain barrier, limiting their clinical development. Nanotechnology-based approaches might improve compounds pharmacokinetics by enhancing binding to the cerebrovascular endothelium and translocation into the brain. Adsorption of apolipoprotein E4 onto polysorbate 80-stabilized nanoparticles to produce a protein corona allows the specific targeting of cerebrovascular endothelium. This strategy increased nanoparticle translocation into brain parenchyma, and improved brain nanoparticle accumulation 3-fold compared to undecorated particles (119.8 vs 40.5 picomoles). Apolipoprotein decorated nanoparticles have high clinical translational potential and may improve the development of nanotechnology-based medicine for a variety of neurological diseases.

Mesenchymal stem cells (MSCs) are adult bone marrow-derived stem cells actually proposed indifferently for the therapy of neurological diseases of both the Central (CNS) and the Peripheral Nervous System (PNS), as a panacea able to treat so many different diseases by their immunomodulatory ability and supportive action on neuronal survival. However, the identification of the exact mechanism of MSC action in the different diseases, although mandatory to define their real and concrete utility, is still lacking. Moreover, CNS and PNS neurons present many different biological properties, and it is still unclear if they respond in the same manner not only to MSC treatment, but also to injuries. For these reasons, in this study we compared the susceptibility of cortical and sensory neurons both to toxic drug exposure and to MSC action, in order to verify if these two neuronal populations can respond differently. Our results demonstrated that Cisplatin (CDDP), Glutamate, and Paclitaxel-treated sensory neurons were protected by the co-culture with MSCs, in different manners: through direct contact able to block apoptosis for CDDP- and Glutamate-treated neurons, and by the release of trophic factors for Paclitaxel-treated ones. A possible key soluble factor for MSC protection was Glutathione, spontaneously released by these cells. On the contrary, cortical neurons resulted more sensitive than sensory ones to the toxic action of the drugs, and overall MSCs failed to protect them. All these data identified for the first time a different susceptibility of cortical and sensory neurons, and demonstrated a protective action of MSCs only against drugs in peripheral neurotoxicity.

Solid lipid nanoparticles (SLN) are colloidal drug delivery systems characterized by higher entrapment efficiency, good scalability of the preparation process and increased sustained prolonged release of the payload compared to other nanocarriers. The possibility to functionalize the surface of SLN with ligands to achieve a site specific targeting makes them attractive to overcome the limited blood-brain barrier (BBB) penetration of therapeutic compounds. SLN are prepared for brain targeting by exploiting the adaptability of warm microemulsion process for the covalent surface modification with an Apolipoprotein E-derived peptide (SLN-mApoE). Furthermore, the influence of the administration route on SLN-mApoE brain bioavailability is here evaluated. SLN-mApoE are able to cross intact a BBB in vitro model. The pulmonary administration of SLN-mApoE is related to a higher confinement in the brain of Balb/c mice compared to the intravenous and intraperitoneal administration routes, without inducing any acute inflammatory reaction in the lungs. These results promote the pulmonary administration of brain-targeted SLN as a feasible strategy for improving brain delivery of therapeutics.

Type-1 Diabetes is generally treated with exogenous insulin administration. Despite treatment, a very common long term consequence of diabetes is the development of a disabling and painful peripheral neuropathy. The transplantation of pancreatic islets is an advanced alternative therapeutic approach, but its clinical application is still very limited, mainly because of the great number of islets required to complete the procedure and of their short-term survival. An intriguing method to improve the performance of pancreatic islets transplantation is the co-transplantation of Mesenchymal Stem Cells (MSCs), adult stem cells already known to support the survival of different cellular populations. In this proof-of-concept study, we demonstrated using an in vivo model of diabetes, the ability of allogenic MSCs to reduce the number of pancreatic islets necessary to achieve glycemic control in diabetic rats, and overall their positive effect on diabetic neuropathy, with the reduction of all the neuropathic signs showed after disease induction. The cutback of the pancreatic islet number required to control glycemia and the regression of the painful neuropathy make MSC co-transplantation a very promising tool to improve the clinical feasibility of pancreatic islet transplantation for diabetes treatment

The difficulties encountered in the treatment of brain diseases with conventional pharmacological tools have created the need for innovative strategies. The combination of nanocarriers and alternative administration routes could represent an efficient approach to reach the brain. Intranasal administration (IN) provides a non-invasive option to deliver drugs to the brain, bypassing the BBB, reducing the first-pass effect and enhancing patient compliance. The objective of the present study was to investigate the biodistribution and bioavailability to the brain of polymeric nanoparticles (PNPs) after IN administration in healthy mice. PNPs were prepared with poly-lactide-co-glycolide polymer using nanoprecipitation method. PNPs had a polymodal distribution around 350 nm. The biodistribution of DiR-loaded PNPs was evaluated by means of 3D fluorescence tomography imaging. Our results show that 3h after a single IN administration, more than 5% of the injected dose was detectable in the brain. PNPs were quickly cleared from the thorax and the abdominal cavity, while the brain fluorescence slowly decreased ranging from 3.7% to 2.3% between 24h and 96h. Repeated IN administrations (2 administrations, 24h apart) provided a significant increment of PNPs-associated fluorescence in the brain, without affecting PNPs accumulation in other organs. These findings support nose-to-brain translocation of PNPs as a noninvasive strategy to enhance the bioavailability of therapeutics to the brain

Background and aim Cerebral ischemia is characterized by both acute and delayed neuronal injuries. Neuro-protection is a major issue that should be properly addressed from a pharmacological point of view, and cell-based treatment approaches are of interest due to their potential pleiotropic effects. Endothelial progenitor cells have the advantage of being mobilized from the bone marrow into the circulation, but have been less studied than other stem cells, such as mesenchymal stem cells. Therefore, the comparison between human endothelial progenitor cells (hEPC) and human mesenchymal progenitor cells (hMSC) in terms of efficacy in rescuing neurons from cell death after transitory ischemia is the aim of the current study, in the effort to address further directions. Materials and methods In vitro model of oxygen-glucose deprivation (OGD) on a primary culture of rodent cortical neurons was set up with different durations of exposure: 1, 2 and 3 hrs with assessment of neuron survival. The 2 hrs OGD was chosen for the subsequent experiments. After 2 hrs OGD neurons were either placed in indirect co-culture with hMSC or hEPC or cultured in hMSC or hEPC conditioned medium and cell viability was evaluated by MTT assay. Results At day 2 after 2 hrs OGD exposure, mean neuronal survival was 47.9 ± 24.2%. In contrast, after treatment with hEPC and hMSC indirect co-culture was 74.1 ± 27.3%; and 69.4 ± 18.8%, respectively. In contrast, treatment with conditioned medium did not provide any advantage in terms of survival to OGD neurons Conclusion The study shows the efficacy of hEPC in indirect co-culture to rescue neurons from cell death after OGD, comparable to that of hMSC. hEPC deserve further studies given their potential interest for ischemia.

Gingival augmentation techniques proposed in the international literature do not exclude a surgical component, which determines consequent post-surgical discomfort and results are not always predictable. In recent years, the introduction of laser biostimulation has led to a less invasive approach, particularly in the treatment of periodontally compromised patients, limiting the surgical phase to seriously compromised cases, with regeneration techniques for the restoration of a correct periodontal tissue anatomy. The aim of this in vitro study is to establish the validity of laser biostimulation in order to develop the epithelial keratinized layer of the tissue by stimulating fibroblasts-keratinocytes organotypic cultures and fibroblasts and keratinocytes mono-cultures. We created two groups (test and control), each one composed of 3 fibroblast cultures, 3 keratinocyte cultures and 3 organotypic cultures. We performed laser irradiation of test group with Wiser Doctor Smile Lambda, Flat Top Handpiece, at 50 J/cm2 of fluency with one application every 40 h for a total of 5 applications. Forty-eight h after the last laser application, we investigated the presence and amount of keratins 5 and 8 with citofluorymetric and western blotting analyses. Analyses showed an increase in keratin synthesis in test group cultures, showing a remarkable increase in production of keratin 8 in co-cultures test. Laser biostimulation can considerably enhance keratin synthesis when applied with high energy doses and repeated applications to keratinocytes-fibroblasts co-cultures.

Multiple Sclerosis (MS) is a neuroinflammatory and immune-mediated chronic disease of the Central Nervous System which progressively damages the axonal myelin sheath, leading to axonal transmission impairment and to the development of neurological symptoms. Most MS cases are characterized by a relapsing-remitting course, and current therapies rely only on the use of immunomodulating drugs which are, however, unable to reverse disease progression. Among the newly proposed alternative therapies, Mesenchymal Stem Cells (MSCs) are considered suitable for MS treatment due to their capacity to modulate the immune response and to modify the pattern of the released cytokines. So far, encouraging results have been obtained with the administration of MSCs before disease onset, mainly in animal models of acute Experimental Autoimmune Encephalomyelitis (EAE) in which MSCs were able to reduce inflammation, thus ameliorating also the disease’s clinical symptoms. On the contrary, only a very small number of studies have investigated the effect of MSCs on relapsing-remitting models of the disease. Here, we investigated the therapeutic potential of MSC administration, both before and after the disease’s onset, in an animal model of MS represented by Dark Agouti rats affected by chronic Relapsing-Remitting EAE. Our results demonstrated that in chronic Relapsing-Remitting EAE the administration of MSCs after the clinical disease’s appearance is able to completely abrogate the relapsing phase and to strongly reduce spinal cord demyelination. These encouraging results have demonstrated that MSCs can provide a protective and reparative strategy for MS treatment.

Introduction and hypothesis: To test in vitro and in vivo the capability of mesh materials to act as scaffolds for rat-derived mesenchymal stem cells (rMSCs) and to compare inflammatory response and collagen characteristics of implant materials, either seeded or not with rMSCs. Methods: rMSCs isolated from rat bone marrow were seeded and cultured in vitro on four different implant materials. Implants showing the best rMSC proliferation rate were selected for the in vivo experiment. Forty-eight adult female Sprague–Dawley rats were randomly divided into two treatment groups. The implant of interest—either seeded or not with rMSCs—was laid and fixed over the muscular abdominal wall. Main outcome measures were: in vitro, proliferation of rMSCs on selected materials; in vivo, the occurrence of topical complications, the evaluation of systemic and local inflammatory response and examination of the biomechanical properties of explants. Results: Surgisis and Pelvitex displayed the best cell growth in vitro. At 90 days in the rat model, rMSCs were related to a lower count of neutrophil cells for Pelvitex and a greater organisation and collagen amount for Surgisis. At 7 days Surgisis samples seeded with rMSCs displayed higher breaking force and stiffness. Conclusions: The presence of rMSCs reduced the systemic inflammatory response on synthetic implants and improved collagen characteristics at the interface between biological grafts and native tissues. rMSCs enhanced the stripping force on biological explants.