Perspectives on Stem Cell Therapy in Diabetic Neuropathic Pain.

Diabetes mellitus-related morbidity and mortality are primarily caused by long-term complications such as retinopathy, nephropathy, cardiomyopathy, and neuropathy. Diabetic neuropathy (DN) involves the progressive degeneration of axons and nerve fibers due to chronic exposure to hyperglycemia. This metabolic disturbance leads to excessive activation of the glycolytic pathway, inducing oxidative stress and mitochondrial dysfunction, ultimately resulting in nerve damage. There is no specific treatment for painful DN, and new approaches should aim not only to relieve pain but also to prevent oxidative stress and reduce inflammation. Given that existing therapies for painful DN are not effective for diabetic patients, mesenchymal stromal cells (MSCs)-based therapy shows promise for providing immunomodulatory and paracrine regulatory functions. MSCs from various sources can improve neuronal dysfunction associated with DN. Transplantation of MSCs has led to a reduction in hyperalgesia and allodynia, along with the recovery of nerve function in diabetic rats. While the pathogenesis of diabetic neuropathic pain is complex, clinical trials have demonstrated the importance of MSCs in modulating the immune response in diabetic patients. MSCs reduce the levels of inflammatory factors and increase anti-inflammatory cytokines, thereby interfering with the progression of DM. Further investigation is necessary to ensure the safety and efficacy of MSCs in preventing or treating neuropathic pain in diabetic patients.

Long-term recruitment of peripheral immune cells to brain scars after a neonatal insult.

Although brain scars in adults have been extensively studied, there is less data available regarding scar formation during the neonatal period, and the involvement of peripheral immune cells in this process remains unexplored in neonates. Using a murine model of neonatal hypoxic-ischemic encephalopathy (HIE) and confocal microscopy, we characterized the scarring process and examined the recruitment of peripheral immune cells to cortical and hippocampal scars for up to 1 year post-insult. Regional differences in scar formation were observed, including the presence of reticular fibrotic networks in the cortex and perivascular fibrosis in the hippocampus. We identified chemokines with chronically elevated levels in both regions and demonstrated, through a parabiosis-based strategy, the recruitment of lymphocytes, neutrophils, and monocyte-derived macrophages to the scars several weeks after the neonatal insult. After 1 year, however, neutrophils and lymphocytes were absent from the scars. Our data indicate that peripheral immune cells are transient components of HIE-induced brain scars, opening up new possibilities for late therapeutic interventions.

Mesenchymal stem cell- and extracellular vesicle-based therapies for Alzheimer's disease: progress, advantages, and challenges.

Alzheimer's disease is a severe, highly disabling neurodegenerative disease, clinically characterized by a progressive decline in cognitive functions, and is the most common form of dementia in the elderly. For decades, the search for disease-modifying therapies has focused on the two main Alzheimer's disease histopathological hallmarks, seeking to prevent, mitigate, or clear the formation of extracellular aggregates of ß-amyloid peptide and intracellular neurofibrillary tangles of tau protein, although without clinical success. Mesenchymal stem cell-based therapy has emerged as a promising alternative for the treatment of Alzheimer's disease, especially because it also targets other crucial players in the pathogenesis of the disease, such as neuroinflammation, synaptic dysfunction/loss, oxidative stress, and impaired neurogenesis. Herein, we review current knowledge of the therapeutic potential of mesenchymal stem cells and their extracellular vesicles for Alzheimer's disease, discussing the most recent findings in both preclinical and clinical trials as well as how advanced technologies have helped to overcome some limitations and contributed to stimulate the development of more effective treatments.

Hyperacute transplantation of umbilical cord mesenchymal stromal cells in a model of severe intracerebral hemorrhage.

Aim: Intracerebral hemorrhage (ICH) has limited therapeutic options. We have shown that an intravenous injection of human umbilical cord-derived mesenchymal stromal cells (hUC-MSC) 24 h after an ICH in rats reduced the residual hematoma volume after a moderate hemorrhage but was inefficient in severe ICH. Here, we investigated whether a treatment in the hyperacute phase would be more effective in severe ICH. Materials & methods: Wistar rats were randomly selected to receive an intravenous injection of hUC-MSC or the vehicle 1 h after a severe ICH. Results: The hyperacute treatment with hUC-MSC did not affect the 22-day survival rate, the motor function or the residual hematoma volume. Conclusion: These results indicate the need for optimization of hUC-MSC-based therapies for severe ICH.

Mesenchymal Stem Cell Therapy in Diabetic Cardiomyopathy.

The incidence and prevalence of diabetes mellitus (DM) are increasing worldwide, and the resulting cardiac complications are the leading cause of death. Among these complications is diabetes-induced cardiomyopathy (DCM), which is the consequence of a pro-inflammatory condition, oxidative stress and fibrosis caused by hyperglycemia. Cardiac remodeling will lead to an imbalance in cell survival and death, which can promote cardiac dysfunction. Since the conventional treatment of DM generally does not address the prevention of cardiac remodeling, it is important to develop new alternatives for the treatment of cardiovascular complications induced by DM. Thus, therapy with mesenchymal stem cells has been shown to be a promising approach for the prevention of DCM because of their anti-apoptotic, anti-fibrotic and anti-inflammatory effects, which could improve cardiac function in patients with DM.

Subacute AMD3100 Treatment Is Not Efficient in Neonatal Hypoxic-Ischemic Rats.

BACKGROUND AND PURPOSE: Despite the advances in treating neonatal hypoxic-ischemic encephalopathy (HIE) with induced hypothermia, the rates of severe disability are still high among survivors. Preclinical studies have indicated that cell therapies with hematopoietic stem/progenitor cells could improve neurological outcomes in HIE. In this study, we investigated whether the administration of AMD3100, a CXCR4 antagonist that mobilizes hematopoietic stem/progenitor cells into the circulation, has therapeutic effects in HIE. METHODS: P10 Wistar rats of both sexes were subjected to right common carotid artery occlusion or sham procedure, and then were exposed to hypoxia for 120 minutes. Two subcutaneous injections of AMD3100 or vehicle were given on the third and fourth day after HIE. We first assessed the interindividual variability in brain atrophy after experimental HIE and vehicle treatment in a small cohort of rats. Based on this exploratory analysis, we designed and conducted an experiment to test the efficacy of AMD3100. Brain atrophy on day 21 after HIE was defined as the primary end point. Secondary efficacy end points were cognitive (T-water maze) and motor function (rotarod) on days 17 and 18 after HIE, respectively. RESULTS: AMD3100 did not decrease the brain atrophy in animals of either sex. Cognitive impairments were not observed in the T-water maze, but male hypoxic-ischemic animals exhibited motor coordination deficits on the rotarod, which were not improved by AMD3100. A separate analysis combining data from animals of both sexes also revealed no evidence of the effectiveness of AMD3100 treatment. CONCLUSIONS: These results indicate that the subacute treatment with AMD3100 does not improve structural and functional outcomes in a rat HIE model.

Mesenchymal Stem Cells Therapies on Fibrotic Heart Diseases.

Stem cell therapy is a promising alternative approach to heart diseases. The most prevalent source of multipotent stem cells, usually called somatic or adult stem cells (mesenchymal stromal/stem cells, MSCs) used in clinical trials is bone marrow (BM-MSCs), adipose tissue (AT-MSCs), umbilical cord (UC-MSCs) and placenta. Therapeutic use of MSCs in cardiovascular diseases is based on the benefits in reducing cardiac fibrosis and inflammation that compose the cardiac remodeling responsible for the maintenance of normal function, something which may end up causing progressive and irreversible dysfunction. Many factors lead to cardiac fibrosis and failure, and an effective therapy is lacking to reverse or attenuate this condition. Different approaches have been shown to be promising in surpassing the poor survival of transplanted cells in cardiac tissue to provide cardioprotection and prevent cardiac remodeling. This review includes the description of pre-clinical and clinical investigation of the therapeutic potential of MSCs in improving ventricular dysfunction consequent to diverse cardiac diseases.

Human mesenchymal stem cell therapy promotes retinal ganglion cell survival and target reconnection after optic nerve crush in adult rats.

BACKGROUND: Optic-nerve injury results in impaired transmission of visual signals to central targets and leads to the death of retinal ganglion cells (RGCs) and irreversible vision loss. Therapies with mesenchymal stem cells (MSCs) from different sources have been used experimentally to increase survival and regeneration of RGCs. METHODS: We investigated the efficacy of human umbilical Wharton's jelly-derived MSCs (hWJ-MSCs) and their extracellular vesicles (EVs) in a rat model of optic nerve crush. RESULTS: hWJ-MSCs had a sustained neuroprotective effect on RGCs for 14, 60, and 120 days after optic nerve crush. The same effect was obtained using serum-deprived hWJ-MSCs, whereas transplantation of EVs obtained from those cells was ineffective. Treatment with hWJ-MSCs also promoted axonal regeneration along the optic nerve and reinnervation of visual targets 120 days after crush. CONCLUSIONS: The observations showed that this treatment with human-derived MSCs promoted sustained neuroprotection and regeneration of RGCs after optic nerve injury. These findings highlight the possibility to use cell therapy to preserve neurons and to promote axon regeneration, using a reliable source of human MSCs.

Iso-Oncotic Albumin Mitigates Brain and Kidney Injury in Experimental Focal Ischemic Stroke.

Background: There is widespread debate regarding the use of albumin in ischemic stroke. We tested the hypothesis that an iso-oncotic solution of albumin (5%), administered earlier after acute ischemic stroke (3 h), could provide neuroprotection without causing kidney damage, compared to a hyper-oncotic albumin (20%) and saline. Objective: To compare the effects of saline, iso-oncotic albumin, and hyper-oncotic albumin, all titrated to similar hemodynamic targets, on the brain and kidney. Methods: Ischemic stroke was induced in anesthetized male Wistar rats (n = 30; weight 437 ± 68 g) by thermocoagulation of pial blood vessels of the primary somatosensory, motor, and sensorimotor cortices. After 3 h, animals were anesthetized and randomly assigned (n = 8) to receive 0.9% NaCl (Saline), iso-oncotic albumin (5% ALB), and hyper-oncotic albumin (20% ALB), aiming to maintain hemodynamic stability (defined as distensibility index of inferior vena cava <25%, mean arterial pressure >80 mmHg). Rats were then ventilated using protective strategies for 2 h. Of these 30 animals, 6 were used as controls (focal ischemic stroke/no fluid). Results: The total fluid volume infused was higher in the Saline group than in the 5% ALB and 20% ALB groups (mean ± SD, 4.3 ± 1.6 vs. 2.7 ± 0.6 and 2.6 ± 0.5 mL, p = 0.03 and p = 0.02, respectively). The total albumin volume infused (g/kg) was higher in the 20% ALB group than in the 5% ALB group (1.4 ± 0.6 vs. 0.4 ± 0.2, p < 0.001). Saline increased neurodegeneration (Fluoro-Jade C staining), brain inflammation in the penumbra (higher tumor necrosis factor-alpha expression), and blood-brain barrier damage (lower gene expressions of claudin-1 and zona occludens-1) compared to both iso-oncotic and hyper-oncotic albumins, whereas it reduced the expression of brain-derived neurotrophic factor (a marker of neuroregeneration) compared only to iso-oncotic albumin. In the kidney, hyper-oncotic albumin led to greater damage as well as higher gene expressions of kidney injury molecule-1 and interleukin-6 than 5% ALB (p < 0.001). Conclusions: In this model of focal ischemic stroke, only iso-oncotic albumin had a protective effect against brain and kidney damage. Fluid therapy thus requires careful analysis of impact not only on the brain but also on the kidney.

Erratum to "Preconditioning of Rat Bone Marrow-Derived Mesenchymal Stromal Cells with Toll-Like Receptor Agonists".

[This corrects the article DOI: 10.1155/2019/7692973.].

Therapeutic Benefit of the Association of Lodenafil with Mesenchymal Stem Cells on Hypoxia-induced Pulmonary Hypertension in Rats.

Pulmonary arterial hypertension (PAH) is characterized by the remodeling of pulmonary arteries, with an increased pulmonary arterial pressure and right ventricle (RV) overload. This work investigated the benefit of the association of human umbilical cord mesenchymal stem cells (hMSCs) with lodenafil, a phosphodiesterase-5 inhibitor, in an animal model of PAH. Male Wistar rats were exposed to hypoxia (10% O2) for three weeks plus a weekly i.p. injection of a vascular endothelial growth factor receptor inhibitor (SU5416, 20 mg/kg, SuHx). After confirmation of PAH, animals received intravenous injection of 5.105 hMSCs or vehicle, followed by oral treatment with lodenafil carbonate (10 mg/kg/day) for 14 days. The ratio between pulmonary artery acceleration time and RV ejection time reduced from 0.42 ± 0.01 (control) to 0.24 ± 0.01 in the SuHx group, which was not altered by lodenafil alone but was recovered to 0.31 ± 0.01 when administered in association with hMSCs. RV afterload was confirmed in the SuHx group with an increased RV systolic pressure (mmHg) of 52.1 ± 8.8 normalized to 29.6 ± 2.2 after treatment with the association. Treatment with hMSCs + lodenafil reversed RV hypertrophy, fibrosis and interstitial cell infiltration in the SuHx group. Combined therapy of lodenafil and hMSCs may be a strategy for PAH treatment.

Paracrine signaling of human mesenchymal stem cell modulates retinal microglia population number and phenotype in vitro.

PURPOSE: Cellular therapy with mesenchymal stem cells (MSC) is emerging as an effective option to treat optic neuropathies. In models of retinal degeneration, MSC injected in the vitreous body protects injured retinal ganglion cells and stimulate their regeneration, however the mechanism is still unknown. Considering the immunomodulating proprieties of MSC and the controversial role of microglial contribution on retinal regeneration, we developed an in vitro co-culture model to analyze the effect of MSC on retinal microglia population. METHODS: We used whole adult rat retinal explants in co-culture with human Wharton's jelly mesenchymal stem cells (hMSC) separated by a transwell membrane and analyzed hMSC effect on both retinal ganglion cells (RGCs) and retinal microglia. RESULTS: hMSC in co-culture protected RGCs after 3 days in vitro by paracrine signaling. In addition, hMSC reduced microglia population and inhibited the pro-inflammatory phenotype of the remaining microglia. CONCLUSIONS: Using a co-culture model, we demonstrated the paracrine effect of hMSC on RGC survival after injury concomitant with a reduction of microglial population. Paracrine signaling of hMSC also changed microglia phenotype and the expression of antiinflammatory factors in the retina. Our results are consistent with a detrimental effect of microglia on RGC survival and regeneration after injury.

Therapy with Cardiomyocytes Derived from Pluripotent Cells in Chronic Chagasic Cardiomyopathy.

Chagas disease discovered more than a century ago remains an incurable disease. The objective of this work was to investigate the therapeutic potential of cardiomyocytes derived from mouse embryonic stem cells (CM-mESC) in a model of chronic Chagasic cardiomyopathy (CCC). Mouse embryonic stem cells (mESC) were characterized, transduced with luciferase, and submitted to cardiac differentiation. CM-mESC were labeled with superparamagnetic iron oxide particles. To induce CCC, mice were infected with Brazil strain trypomastigotes. At 150 days post-infection (dpi), infected animals were treated with CM-mESC or PBS. Cells were detected by magnetic resonance imaging (MRI) and bioluminescence. Cardiac function was evaluated by MRI and electrocardiogram at 150 and 196 dpi. CCC mice showed significant differences in MRI and ECG parameters compared to non-infected mice. However, no differences were observed in contractile and electrical parameters between cell and PBS injected groups, 45 days after cell transplantation. Cells were detected 24 h after transplantation by MRI. CM-mESC bioluminescence tracking demonstrated over 90% decrease in signal 8 days after treatment. Nevertheless, the Infected + CM-mESC group showed a significant reduction in the percentage of collagen fibers when compared to the Infected + PBS group. In conclusion, CM-mESC therapy was not effective in reversing cardiac functional changes induced by Chagas disease despite some improvement in myocardial fibrosis.

Intravenous Human Umbilical Cord-Derived Mesenchymal Stromal Cell Administration in Models of Moderate and Severe Intracerebral Hemorrhage.

Intracerebral hemorrhage (ICH) is as a life-threatening condition that can occur in young adults, often causing long-term disability. Recent preclinical data suggest mesenchymal stromal cell (MSC)-based therapies as promising options to minimize brain damage after ICH. However, therapeutic evidence and mechanistic insights are still limited, particularly when compared with other disorders such as ischemic stroke. Herein, we employed a model of collagenase-induced ICH in young adult rats to investigate the potential therapeutic effects of an intravenous injection of human umbilical cord Wharton's jelly-derived MSCs (hUC-MSCs). Two doses of collagenase were used to cause moderate or severe hemorrhages. Magnetic resonance imaging showed that animals treated with hUC-MSCs after moderate ICH had smaller residual hematoma volumes than vehicle-treated rats, whereas the cell therapy failed to decrease the hematoma volume in animals with a severe ICH. Functional assessments (rotarod and elevated body swing tests) were performed for up to 21 days after ICH. Enduring neurological impairments were seen only in animals subjected to severe ICH, but the cell therapy did not induce statistically significant improvements in the functional recovery. The biodistribution of Technetium-99m-labeled hUC-MSCs was also evaluated, showing that most cells were found in organs such as the spleen and lungs 24 h after transplantation. Nevertheless, it was possible to detect a weak signal in the brain, which was higher in the ipsilateral hemisphere of rats subjected to a severe ICH. These data indicate that hUC-MSCs have moderately beneficial effects in cases of less severe brain hemorrhages in rats by decreasing the residual hematoma volume, and that optimization of the therapy is still necessary.

Effects of a combinatorial treatment with gene and cell therapy on retinal ganglion cell survival and axonal outgrowth after optic nerve injury.

After an injury, axons in the central nervous system do not regenerate over large distances and permanently lose their connections to the brain. Two promising approaches to correct this condition are cell and gene therapies. In the present work, we evaluated the neuroprotective and neuroregenerative potential of pigment epithelium-derived factor (PEDF) gene therapy alone and combined with human mesenchymal stem cell (hMSC) therapy after optic nerve injury by analysis of retinal ganglion cell survival and axonal outgrowth. Overexpression of PEDF by intravitreal delivery of AAV2 vector significantly increased Tuj1-positive cells survival and modulated FGF-2, IL-1ß, Iba-1, and GFAP immunostaining in the ganglion cell layer (GCL) at 4 weeks after optic nerve crush, although it could not promote axonal outgrowth. The combination of AAV2.PEDF and hMSC therapy showed a higher number of Tuj1-positive cells and a pronounced axonal outgrowth than unimodal therapy after optic nerve crush. In summary, our results highlight a synergistic effect of combined gene and cell therapy relevant for future therapeutic interventions regarding optic nerve injury.

Tempol improves neuroinflammation and delays motor dysfunction in a mouse model (SOD1G93A) of ALS.

BACKGROUND: The development of new therapeutic strategies to treat amyotrophic lateral sclerosis (ALS) is of utmost importance. The use of cyclic nitroxides such as tempol may provide neuroprotection and improve lifespan. We investigated whether tempol (50 mg/kg) presents therapeutic potential in SOD1G93A transgenic mice. METHODS: Tempol treatment began at the asymptomatic phase of the disease (10th week) and was administered every other day until week 14, after which it was administered twice a week until the final stage of the disease. The animals were sacrificed at week 14 (initial stage of symptoms-ISS) and at the end stage (ES) of the disease. The lumbar spinal cord of the animals was dissected and processed for use in the following techniques: Nissl staining to evaluate neuronal survival; immunohistochemistry to evaluate astrogliosis and microgliosis (ISS and ES); qRT-PCR to evaluate the expression of neurotrophic factors and pro-inflammatory cytokines (ISS); and transmission electron microscopy to evaluate the alpha-motoneurons (ES). Behavioral analyses considering the survival of animals, bodyweight loss, and Rotarod motor performance test started on week 10 and were performed every 3 days until the end-stage of the disease. RESULTS: The results revealed that treatment with tempol promoted greater neuronal survival (23%) at ISS compared to untreated animals, which was maintained until ES. The intense reactivity of astrocytes and microglia observed in vehicle animals was reduced in the lumbar spinal cords of the animals treated with tempol. In addition, the groups treated with tempol showed reduced expression of proinflammatory cytokines (IL1ß and TNFα) and a three-fold decrease in the expression of TGFß1 at ISS compared with the group treated with vehicle. CONCLUSIONS: Altogether, our results indicate that treatment with tempol has beneficial effects, delaying the onset of the disease by enhancing neuronal survival and decreasing glial cell reactivity during ALS progression in SOD1G93A mice.

Extracellular vesicles derived from human Wharton's jelly mesenchymal stem cells protect hippocampal neurons from oxidative stress and synapse damage induced by amyloid-ß oligomers.

BACKGROUND: Mesenchymal stem cells (MSCs) have been explored as promising tools for treatment of several neurological and neurodegenerative diseases. MSCs release abundant extracellular vesicles (EVs) containing a variety of biomolecules, including mRNAs, miRNAs, and proteins. We hypothesized that EVs derived from human Wharton's jelly would act as mediators of the communication between hMSCs and neurons and could protect hippocampal neurons from damage induced by Alzheimer's disease-linked amyloid beta oligomers (AßOs). METHODS: We isolated and characterized EVs released by human Wharton's jelly mesenchymal stem cells (hMSC-EVs). The neuroprotective action of hMSC-EVs was investigated in primary hippocampal cultures exposed to AßOs. RESULTS: hMSC-EVs were internalized by hippocampal cells in culture, and this was enhanced in the presence of AßOs in the medium. hMSC-EVs protected hippocampal neurons from oxidative stress and synapse damage induced by AßOs. Neuroprotection by hMSC-EVs was mediated by catalase and was abolished in the presence of the catalase inhibitor, aminotriazole. CONCLUSIONS: hMSC-EVs protected hippocampal neurons from damage induced by AßOs, and this was related to the transfer of enzymatically active catalase contained in EVs. Results suggest that hMSC-EVs should be further explored as a cell-free therapeutic approach to prevent neuronal damage in Alzheimer's disease.

Reduction of cardiac and renal dysfunction by new inhibitor of DPP4 in diabetic rats.

BACKGROUND: Increased mortality due to type 2 diabetes mellitus (T2DM) has been associated with renal and/or cardiovascular dysfunction. Dipeptidyl dipeptidase-4 inhibitors (iDPP-4s) may exert cardioprotective effects through their pleiotropic actions via glucagon-like peptide 1-dependent mechanisms. In this study, the pharmacological profile of a new iDPP-4 (LASSBio-2124) was investigated in rats with cardiac and renal dysfunction induced by T2DM. METHODS: T2DM was induced in rats by 2 weeks of a high-fat diet followed by intravenous injection of streptozotocin. Metabolic disturbance and cardiac, vascular, and renal dysfunction were analyzed in the experimental groups. RESULTS: Sitagliptin and LASSBio-2124 administration after T2DM induction reduced elevated glucose levels to 319.8 ±â€¯13.2 and 279.7 ±â€¯17.8 mg/dL, respectively (p < 0.05). LASSBio-2124 also lowered the cholesterol and triglyceride levels from 76.8 ±â€¯8.0 to 42.7 ±â€¯3.2 mg/dL and from 229.7 ±â€¯25.4 to 100.7 ±â€¯17.1 mg/dL, in diabetic rats. Sitagliptin and LASSBio-2124 reversed the reduction of the plasma insulin level. LASSBio-2124 recovered the increased urinary flow in diabetic animals and reduced 24-h proteinuria from 23.7 ±â€¯1.5 to 13.3 ±â€¯2.8 mg (p < 0.05). It also reduced systolic and diastolic left-ventricular dysfunction in hearts from diabetic rats. CONCLUSION: The effects of LASSBio-2124 were superior to those of sitagliptin in the cardiovascular systems of T2DM rats. This new prototype showed promise for the avoidance of comorbidities in a T2DM experimental model, and thus may constitute an innovative therapeutic agent for the treatment of these conditions in the clinical field in future.

Preconditioning of Rat Bone Marrow-Derived Mesenchymal Stromal Cells with Toll-Like Receptor Agonists.

Bone marrow-derived mesenchymal stromal cells (BM-MSCs) are dynamic cells that can sense the environment, adapting their regulatory functions to different conditions. Accordingly, the therapeutic potential of BM-MSCs can be modulated by preconditioning strategies aimed at modifying their paracrine action. Although rat BM-MSCs (rBM-MSCs) have been widely tested in preclinical research, most preconditioning studies have employed human and mouse BM-MSCs. Herein, we investigated whether rBM-MSCs modify their phenotype and paracrine functions in response to Toll-like receptor (TLR) agonists. The data showed that rBM-MSCs expressed TLR3, TLR4, and MDA5 mRNA and were able to internalize polyinosinic-polycytidylic acid (Poly(I:C)), a TLR3/MDA5 agonist. rBM-MSCs were then stimulated with Poly(I:C) or with lipopolysaccharide (LPS, a TLR4 agonist) for 1 h and were grown under normal culture conditions. LPS or Poly(I:C) stimulation did not affect the viability or the morphology of rBM-MSCs and did not modify the expression pattern of key cell surface markers. Poly(I:C) did not induce statistically significant changes in the release of several inflammatory mediators and VEGF by rBM-MSCs, although it tended to increase IL-6 and MCP-1 secretion, whereas LPS increased the release of IL-6, MCP-1, and VEGF, three factors that were constitutively secreted by unstimulated cells. The neurotrophic activity of the conditioned medium from unstimulated and LPS-preconditioned rBM-MSCs was investigated using dorsal root ganglion explants, showing that soluble factors produced by unstimulated and LPS-preconditioned rBM-MSCs can stimulate neurite outgrowth similarly, in a VEGF-dependent manner. LPS-preconditioned cells, however, were slightly more efficient in increasing the number of regrowing axons in a model of sciatic nerve transection in rats. In conclusion, LPS preconditioning boosted the production of constitutively secreted factors by rBM-MSCs, without changing their mesenchymal identity, an effect that requires further investigation in exploratory preclinical studies.

Long-term neuronal survival, regeneration, and transient target reconnection after optic nerve crush and mesenchymal stem cell transplantation.

BACKGROUND: Retina and/or optic nerve injury may cause irreversible blindness, due to degeneration of retinal ganglion cells. We and others have previously shown that the intravitreal injection of mesenchymal stem cells (MSCs) protects injured retinal ganglion cells and stimulates their regeneration after optic nerve injury, but the long-term effects of this therapy are still unknown. METHODS: We injected rat MSC (rMSC) intravitreally in adult (3-5 months) Lister Hooded rats of either sex after optic nerve crush. Retinal ganglion cell survival, axonal regeneration, and reconnection were analyzed 60 and 240 days after crush by immunohistochemistry for Tuj1, anterograde labeling with cholera-toxin B and by immunohistochemistry for nerve growth factor-induced gene A (NGFI-A, driven by light stimulation) in the superior colliculus after a cycle of light deprivation-stimulation. Visual behaviors (optokinetic reflex, looming response, and preference for dark) were analyzed 70 days after crush. RESULTS: rMSC treatment doubled the number of surviving retinal ganglion cells, preferentially of a larger subtype, and of axons regenerating up to 0.5 mm. Some axons regenerated to the lateral geniculate nucleus and superior colliculus. NGFI-A+ cells were doubled in rMSC-treated animals 60 days after crush, but equivalent to vehicle-injected animals 240 days after crush, suggesting that newly formed synapses degenerated. Animals did not recover visual behaviors. CONCLUSIONS: We conclude that rMSC-induced neuroprotection is sustained at longer time points. Although rMSCs promoted long-term neuroprotection and long-distance axon regeneration, the reconnection of retinal ganglion cells with their targets was transitory, indicating that they need additional stimuli to make stable reconnections.