To evaluate the feasibility, safety, and immunological effects of intrathecal and intravenous administration of autologous mesenchymal stem cells (MSCs) (also called mesenchymal stromal cells) in patients with multiple sclerosis (MS) and amyotrophic lateral sclerosis (ALS). Design: A phase 1/2 open-safety clinical trial. Patients: Fifteen patients with MS (mean [SD] Expanded Disability Status Scale [EDSS] score, 6.7 [1.0]) and 19 with ALS (mean [SD] Amyotrophic Lateral Sclerosis Functional Rating Scale [ALSFRS] score, 20.8 [8.0]) were enrolled. Intervention: After culture, a mean (SD) of 63.2ϫ 10 6(2.5ϫ10 6 ) MSCs was injected intrathecally (n =34) and intravenously (n = 14). In 9 cases, MSCs were magnetically labeled with the superparamagnetic iron oxide ferumoxides (Feridex). Main Outcome Measures:The main outcome measure was the recording of side effects. Follow-up (Յ25 months) included adverse events evaluation, neurological disability assessment by means of the EDSS, magnetic resonance imaging to exclude unexpected pathologies and track the labeled stem cells, and immunological tests to assess the short-term immunomodulatory effects of MSC transplantation.Results: Twenty-one patients had injection-related adverse effects consisting of transient fever, and 15 reported headache. No major adverse effects were reported during follow-up. The mean ALSFRS score remained stable during the first 6 months of observation, whereas the mean (SD) EDSS score improved from 6.7 (1.0) to 5.9 (1.6). Magnetic resonance imaging visualized the MSCs in the occipital horns of the ventricles, indicating the possible migration of ferumoxides-labeled cells in the meninges, subarachnoid space, and spinal cord. Immunological analysis revealed an increase in the proportion of CD4 ϩ CD25 ϩ regulatory T cells, a decrease in the proliferative responses of lymphocytes, and the expression of CD40 ϩ , CD83 ϩ , CD86 ϩ , and HLA-DR on myeloid dendritic cells at 24 hours after MSC transplantation. Conclusion:Transplantation of MSCs in patients with MS and ALS is a clinically feasible and relatively safe procedure and induces immediate immunomodulatory effects.

To investigate the therapeutic potential of mesenchymal stromal cells (MSCs) in the chronic model of experimental autoimmune encephalomyelitis (EAE). Design: Mesenchymal stromal cells were obtained from the bone marrow of naïve C57BL and green fluorescent protein-transgenic mice and cultured with Eagle minimum essential medium/alpha medium after removal of adhering cells. Following 2 to 3 passages, MSCs were injected intraventricularly or intravenously into mice in which chronic EAE had been induced with myelin oligodendrocyte glycoprotein 35-55 peptide. Results: In 8 separate experiments, the intravenously and intraventricularly injected green fluorescent proteinpositive MSCs were attracted to the areas of central nervous system inflammation and expressed galactocerebroside, O4, glial fibrillary acidic protein, and ␤-tubulin type III. The clinical course of chronic EAE was ameliorated in MSC-treated animals (0% mortality; mean [SE] maximal EAE score, 1.76 [1.01] and 1.8 [0.46] in the intraventricular and intravenous groups, respectively, vs 13% and 21% mortality and 2.80 [0.79] and 3.42 [0.54]

Adult mesenchymal stem cells (MSC) that are able to differentiate into various mesenchymal cell types are typically isolated from bone marrow, but their significant presence in human peripheral blood (PB) is controversial. Fibrin microbeads (FMB) that bind matrix-dependent cells were used to isolate MSC from the mononuclear fraction of mobilized PB of adult healthy human donors treated with a granulocyte colony-stimulating factor. Isolation by plastic adherence resulted in a negligible number of MSC in all samples tested, whereas FMBbased isolation yielded spindle-shaped cell samples that could further expand on plastic or on FMB in eight out of the 11 samples. The yield of these cells at days 17-18 after the harvest was B0.5% of the initial cell number. The isolated cells were grown on plastic and characterized by FACS analysis and immunohistochemistry for specific markers. Following culturing and first passage, the FMBisolated cells stained positive for mesenchymal stromal cell markers CD90 and CD105, expressed vimentin and fibronectin and were negative for hematopoietic markers CD45 and CD34. These cells could differentiate into osteoblasts, adipocytes and chondrocytes. This study indicates that FMB may have special advantage in isolating MSC from sources such as mobilized PB, where the number of such cells is scarce.

IMPORTANCE Preclinical studies have shown that neurotrophic growth factors (NTFs) extend the survival of motor neurons in amyotrophic lateral sclerosis (ALS) and that the combined delivery of these neurotrophic factors has a strong synergistic effect. We have developed a culture-based method for inducing mesenchymal stem cells (MSCs) to secrete neurotrophic factors. These MSC-NTF cells have been shown to be protective in several animal models of neurodegenerative diseases. OBJECTIVE To determine the safety and possible clinical efficacy of autologous MSC-NTF cells transplantation in patients with ALS. DESIGN, SETTING, AND PARTICIPANTS In these open-label proof-of-concept studies, patients with ALS were enrolled between June 2011 and October 2014 at the Hadassah Medical Center in Jerusalem, Israel. All patients were followed up for 3 months before transplantation and 6 months after transplantation. In the phase 1/2 part of the trial, 6 patients with early-stage ALS were injected intramuscularly (IM) and 6 patients with more advanced disease were transplanted intrathecally (IT). In the second stage, a phase 2a dose-escalating study, 14 patients with early-stage ALS received a combined IM and IT transplantation of autologous MSC-NTF cells. INTERVENTIONS Patients were administered a single dose of MSC-NTF cells. MAIN OUTCOMES AND MEASURES The primary end points of the studies were safety and tolerability of this cell therapy. Secondary end points included the effects of the treatment on various clinical parameters, such as the ALS Functional Rating Scale-Revised score and the respiratory function. RESULTS Among the 12 patients in the phase 1/2 trial and the 14 patients in the phase 2a trial aged 20 and 75 years, the treatment was found to be safe and well tolerated over the study follow-up period. Most of the adverse effects were mild and transient, not including any treatment-related serious adverse event. The rate of progression of the forced vital capacity and of the ALS Functional Rating Scale-Revised score in the IT (or IT+IM)-treated patients was reduced (from −5.1% to −1.2%/month percentage predicted forced vital capacity, P < .04 and from −1.2 to 0.6 ALS Functional Rating Scale-Revised points/month, P = .052) during the 6 months following MSC-NTF cell transplantation vs the pretreatment period. Of these patients, 13 (87%) were defined as responders to either ALS Functional Rating Scale-Revised or forced vital capacity, having at least 25% improvement at 6 months after treatment in the slope of progression. CONCLUSIONS AND RELEVANCE The results suggest that IT and IM administration of MSC-NTF cells in patients with ALS is safe and provide indications of possible clinical benefits, to be confirmed in upcoming clinical trials.

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In this study (trial registration: NCT02166021), we aimed to evaluate the optimal way of administration, the safety and the clinical efficacy of mesenchymal stem cell (MSC) transplantation in patients with active and progressive multiple sclerosis. Forty-eight patients (28 males and 20 females) with progressive multiple sclerosis (Expanded Disability Status Scale: 3.0–6.5, mean : 5.6 ± 0.8, mean age: 47.5 ± 12.3) and evidence of either clinical worsening or activity during the previous year, were enrolled (between 2015 and 2018). Patients were randomized into three groups and treated intrathecally (IT) or intravenously (IV) with autologous MSCs (1 × 106/kg) or sham injections. After 6 months, half of the patients from the MSC-IT and MSC-IV groups were retreated with MSCs, and the other half with sham injections. Patients initially assigned to sham treatment were divided into two subgroups and treated with either MSC-IT or MSC-IV. The study duration was 14 months. No serious treatment-related safety issues were detected. Significantly fewer patients experienced treatment failure in the MSC-IT and MSC-IV groups compared with those in the sham-treated group (6.7%, 9.7%, and 41.9%, respectively, P = 0.0003 and P = 0.0008). During the 1-year follow-up, 58.6% and 40.6% of patients treated with MSC-IT and MSC-IV, respectively, exhibited no evidence of disease activity compared with 9.7% in the sham-treated group (P &lt; 0.0001 and P &lt; 0.0048, respectively). MSC-IT transplantation induced additional benefits on the relapse rate, on the monthly changes of the T2 lesion load on MRI, and on the timed 25-foot walking test, 9-hole peg test, optical coherence tomography, functional MRI and cognitive tests. Treatment with MSCs was well-tolerated in progressive multiple sclerosis and induced short-term beneficial effects regarding the primary end points, especially in the patients with active disease. The intrathecal administration was more efficacious than the intravenous in several parameters of the disease. A phase III trial is warranted to confirm these findings.

Mesenchymal stromal cells (MSC) are part of the bone marrow stem cells repertoire which also includes the main stem cells population of the bone marrow, the hematopoietic stem cells. The main role of MSCs is to support hematopoiesis but they can also give rise to cells of the mesodermal layers. Recently, significant interactions between MSCs and cells from the immune system have been demonstrated: MSCs were found to downregulate T and B lymphocytes, natural killer cells (NK) and antigen presenting cells through various mechanisms, including cell-to cell interaction and soluble factor production. Besides the immunomodulatory effects, MSCs were shown to possess additional stem cells features, such as the self-renewal potential and multipotency. Their debatable transdifferentiation potential to cells of the endo- and exo-dermal layer, including cells of the CNS, may explain in part their reported neuroprotective effects. Studies in vitro and in vivo (in cells cultures and in animal models) have indicated neuroprotective effects. MSCs are believed to promote functional recovery following CNS injury or inflammation, by producing trophic factors that may facilitate the mobilization of endogenous neural stem cells and promote the regeneration or the survival of the affected neurons. These immunomodulatory and neuroprotective features could make MSCs potential candidates for future therapeutic modalities in immune-mediated and neurodegenerative diseases.

Fibrin microbeads (FMB), made of extensively cross-linked dense and partially denatured fibrin, were used as a matrix for efficient isolation of mesenchymal stem cells (MSC) from rat bone marrow (BM). After 2 days of incubation of FMB with whole BM in suspension, a high number of cells of mesenchymal origin attached to the FMB. On the 14th day after their transfer to plastic, the yield of the cells isolated via FMB was approximately 3-4 times higher than that obtained by currently used protocols based solely on plastic adhesion. This implies that the number of MSC in BM may be higher than previously reported. FACS analyses and immunostaining showed the mesenchymal characteristics of these cells by positive staining for fibronectin, vimentin, CD49E, and CD29. Immediately after isolation, less than 20% of the cells still expressed the hematopoietic markers CD11b and CD45. Most of these cells were eventually eliminated after further expansion of the isolated cells on plastic. Cells isolated via FMB were expanded in culture for more than 4 months and could be defined as MSC along this time period based on their ability to differentiate into precursors of mesenchymal tissues, such as osteogenic, adipogenic, and chondrogenic cells. Similar differentiation plasticity was observed in clones derived from single cells from whole MSC populations isolated via FMB. Based on our results we propose that FMB can serve as a 3-dimensional biodegradable matrix for isolation, differentiation, and possibly implantation of MSC for tissue regeneration.