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Effect of neuroglobin genetically modified bone marrow mesenchymal stem cells transplantation on spinal cord injury in rabbits.

ABSTRACT: OBJECTIVE: This study aims to investigate the potentially protective effect of neuroglobin (Ngb) gene-modified bone marrow mesenchymal stem cells (BMSCs) on traumatic spinal cord injury (SCI) in rabbits. METHODS: A lentiviral vector containing an Ngb gene was constructed and used to deliver Ngb to BMSCs. Ngb gene-modified BMSCs were then injected at the SCI sites 24 hours after SCI. The motor functions of the rabbits were evaluated by the Basso-Beattie-Bresnahan rating scale. Fluorescence microscopy, quantitative real-time PCRs, Western blots, malondialdehyde (MDA) tests, and terminal deoxynucleotidyltransferase-mediated UTP end labeling assays were also performed. RESULTS: Ngb expression in the Ngb-BMSC group increased significantly. A more significant functional improvement was observed in the Ngb-BMSC group compared with those in the other groups. Traumatic SCI seemingly led to an increase in MDA level and number of apoptotic cells, which can be prevented by Ngb-BMSC treatment. CONCLUSION: This study demonstrates that Ngb gene-modified BMSCs can strengthen the therapeutic benefits of BMSCs in reducing secondary damage and improving the neurological outcome after traumatic SCI. Therefore, the combined strategy of BMSC transplantation and Ngb gene therapy can be used to treat traumatic SCI.

SUBMITTER: Lin WP

PROVIDER: S-EPMC3642116 | biostudies-other | 2013

REPOSITORIES: biostudies-other

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Effect of neuroglobin genetically modified bone marrow mesenchymal stem cells transplantation on spinal cord injury in rabbits.

Lin Wen-Ping WP Chen Xuan-Wei XW Zhang Li-Qun LQ Wu Chao-Yang CY Huang Zi-Da ZD Lin Jian-Hua JH

PloS one 20130502 5

<h4>Objective</h4>This study aims to investigate the potentially protective effect of neuroglobin (Ngb) gene-modified bone marrow mesenchymal stem cells (BMSCs) on traumatic spinal cord injury (SCI) in rabbits.<h4>Methods</h4>A lentiviral vector containing an Ngb gene was constructed and used to deliver Ngb to BMSCs. Ngb gene-modified BMSCs were then injected at the SCI sites 24 hours after SCI. The motor functions of the rabbits were evaluated by the Basso-Beattie-Bresnahan rating scale. Fluore ...[more]

PMID: 23658829

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Project description:The administration of stem cells holds promise as a potential therapy for spinal cord injury (SCI). Mesenchymal stem cells have advantages for clinical applications, since they can be easily obtained, are suitable for autologous transplantation and have been previously shown to induce regeneration of the spinal cord in experimental settings. Here we evaluated the feasibility, safety and potential efficacy of autologous transplantation of mesenchymal stem cells in subjects with chronic complete SCI.We conducted a phase I, non-controlled study in 14 subjects of both genders aging between 18 to 65 years, with chronic traumatic SCI (>6 months), at thoracic or lumbar levels, classified as American Spinal Injury Association (ASIA) A - complete injury. Baseline somatosensory evoked potentials (SSEP), spinal magnetic resonance imaging (MRI) and urodynamics were assessed before and after treatment. Pain rating was performed using the McGill Pain Questionnaire and a visual analogue score scale. Bone marrow-derived mesenchymal stem cells were cultured and characterized by flow cytometry, cell differentiation assays and G-band karyotyping. Mesenchymal stem cells were injected directly into the lesion following laminectomy and durotomy.Cell transplantation was an overall safe and well-tolerated procedure. All subjects displayed variable improvements in tactile sensitivity and eight subjects developed lower limbs motor functional gains, principally in the hip flexors. Seven subjects presented sacral sparing and improved American Spinal Injury Association impairment scale (AIS) grades to B or C - incomplete injury. Nine subjects had improvements in urologic function. One subject presented changes in SSEP 3 and 6 months after mesenchymal stem cells transplantation. Statistically significant correlations between the improvements in neurological function and both injury size and level were found.Intralesional transplantation of autologous mesenchymal stem cells in subjects with chronic, complete spinal cord injury is safe, feasible, and may promote neurological improvements.ClinicalTrials.gov NCT01325103 - Registered 28 March 2011.

Project description:Currently, there is no effective strategy to promote functional recovery after a spinal cord injury. Collagen scaffolds can not only provide support and guidance for axonal regeneration, but can also serve as a bridge for nerve regeneration at the injury site. They can additionally be used as carriers to retain mesenchymal stem cells at the injury site to enhance their effectiveness. Hence, we hypothesized that transplanting human umbilical cord-mesenchymal stem cells on collagen scaffolds would enhance healing following acute complete spinal cord injury. Here, we test this hypothesis through animal studies and a phase I clinical trial. (1) Animal experiments: Models of completely transected spinal cord injury were established in rats and canines by microsurgery. Mesenchymal stem cells derived from neonatal umbilical cord tissue were adsorbed onto collagen scaffolds and surgically implanted at the injury site in rats and canines; the animals were observed after 1 week-6 months. The transplantation resulted in increased motor scores, enhanced amplitude and shortened latency of the motor evoked potential, and reduced injury area as measured by magnetic resonance imaging. (2) Phase I clinical trial: Forty patients with acute complete cervical injuries were enrolled at the Characteristic Medical Center of Chinese People's Armed Police Force and divided into two groups. The treatment group (n = 20) received collagen scaffolds loaded with mesenchymal stem cells derived from neonatal umbilical cord tissues; the control group (n = 20) did not receive the stem-cell loaded collagen implant. All patients were followed for 12 months. In the treatment group, the American Spinal Injury Association scores and activities of daily life scores were increased, bowel and urinary functions were recovered, and residual urine volume was reduced compared with the pre-treatment baseline. Furthermore, magnetic resonance imaging showed that new nerve fiber connections were formed, and diffusion tensor imaging showed that electrophysiological activity was recovered after the treatment. No serious complication was observed during follow-up. In contrast, the neurological functions of the patients in the control group were not improved over the follow-up period. The above data preliminarily demonstrate that the transplantation of human umbilical cord-mesenchymal stem cells on a collagen scaffold can promote the recovery of neurological function after acute spinal cord injury. In the future, these results need to be confirmed in a multicenter, randomized controlled clinical trial with a larger sample size. The clinical trial was approved by the Ethics Committee of the Characteristic Medical Center of Chinese People's Armed Police Force on February 3, 2016 (approval No. PJHEC-2016-A8). All animal experiments were approved by the Ethics Committee of the Characteristic Medical Center of Chinese People's Armed Police Force on May 20, 2015 (approval No. PJHEC-2015-D5).

Project description:The objective of this study was to determine the potential of human bone marrow derived mesenchymal stem cells (hBMSCs) as gene carriers for improving the outcome of human islet transplantation. hBMSCs were characterized for the expression of phenotypic markers and transduced with Adv-hVEGF-hIL-1Ra to overexpress human vascular endothelial growth factor (hVEGF) and human interleukin-1 receptor antagonist (hIL-1Ra). Human islets were co-cultured with hBMSCs overexpressing hVEGF and hIL-1Ra. Islet viability was determined by membrane fluorescent method and glucose stimulation test. Transduced hBMSCs and human islets were co-transplanted under the kidney capsule of NOD.Cg-Prkdc(scid) Il2rg(tm1Wjl) /SzJ (NSG) diabetic mice and blood glucose levels were measured over time to demonstrate the efficacy of genetically modified hBMSCs. At the end of study, immunofluorescent staining of kidney section bearing islets was performed for insulin and von Willebrand Factor (vWF). hBMSCs were positive for the expression of CD73, CD90, CD105, CD146 and Stro-1 surface markers as determined by flow cytometry. Transduction of hBMSCs with adenovirus did not affect their stemness and differentiation potential as confirmed by mRNA levels of stem cell markers and adipogenic differentiation of transduced hBMSCs. hBMSCs were efficiently transduced with Adv-hVEGF-hIL-1Ra to overexpress hVEGF and hIL-1Ra. Live dead cell staining and glucose stimulation test have shown that transduced hBMSCs improved the viability of islets against cytokine cocktail. Co-transplantation of human islets with genetically modified hBMSCs improved the glycemic control of diabetic NSG mice as determined by mean blood glucose levels and intraperitoneal glucose tolerance test. Immunofluorescent staining of kidney sections was positive for human insulin and vWF. In conclusion, our results have demonstrated that hBMSCs may be used as gene carriers and nursing cells to improve the outcome of islet transplantation.

Project description:BackgroundSpinal cord injury (SCI) is an inflammatory condition, and excessive adenosine triphosphate (ATP) is released into the extracellular space, which can be catabolized into adenosine by CD73. Extracellular vesicles have been designed as nano drug carriers in many diseases. However, their impacts on delivery of CD73 after SCI are not yet known. We aimed to construct CD73 modified extracellular vesicles and explore the anti-inflammatory effects after SCI.MethodsCD73 engineered extracellular vesicles (CD73+ hucMSC-EVs) were firstly established, which were derived from human umbilical cord mesenchymal stem cells (hucMSCs) transduced by lentiviral vectors to upregulate the expression of CD73. Effects of CD73+ hucMSC-EVs on hydrolyzing ATP into adenosine were detected. The polarization of M2/M1 was verified by immunofluorescence. Furthermore, A2aR and A2bR inhibitors and A2bR knockdown cells were used to investigate the activated adenosine receptor. Biomarkers of microglia and levels of cAMP/PKA were also detected. Repetitively in vivo study, morphology staining, flow cytometry, cytokine analysis, and ELISA assay, were also applied for verifications.ResultsCD73+ hucMSC-EVs reduced concentration of ATP and promoted the level of adenosine. In vitro experiments, CD73+ hucMSC-EVs increased macrophages/microglia M2:M1 polarization, activated adenosine 2b receptor (A2bR), and then promoted cAMP/PKA signaling pathway. In mice using model of thoracic spinal cord contusion injury, CD73+ hucMSC-EVs improved the functional recovery after SCI through decreasing the content of ATP in cerebrospinal fluid and improving the polarization from M1 to M2 phenotype. Thus, the cascaded pro-inflammatory cytokines were downregulated, such as TNF-α, IL-1β, and IL-6, while the anti-inflammatory cytokines were upregulated, such as IL-10 and IL-4.ConclusionsCD73+ hucMSC-EVs ameliorated inflammation after spinal cord injury by reducing extracellular ATP, promoting A2bR/cAMP/PKA pathway and M2/M1 polarization. CD73+ hucMSC-EVs might be promising nano drugs for clinical application in SCI therapy.

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Effect of neuroglobin genetically modified bone marrow mesenchymal stem cells transplantation on spinal cord injury in rabbits.

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Effect of neuroglobin genetically modified bone marrow mesenchymal stem cells transplantation on spinal cord injury in rabbits.

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