Combined Transplantation of Human Neural Stem Cells and Human Mesenchymal Stem Cells Following Spinal Cord Injury

Combined Transplantation of Human Neural Stem Cells and Human Mesenchymal Stem Cells Following Spinal Cord Injury

Proceedings of the NASS 26th Annual Meeting / The Spine Journal 11 (2011) 1S–173S occurred early in the healing process with day 3, 7 and 14 levels ex...

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Proceedings of the NASS 26th Annual Meeting / The Spine Journal 11 (2011) 1S–173S occurred early in the healing process with day 3, 7 and 14 levels exhibiting control (non operated distant skeletal site) and treated sites with similar noggin responses. Bone morphogenic protein-3(BMP-3) - The temporal gene expression pattern was interesting in terms of a delayed upregulation not occurring until day 14 with exposure to BMP-7. In addition, the results suggest down-regulation of BMP-3 at day 3 and 7 when compared to control levels for all treatment groups. This biphasic initial down then up-regulation of BMP-3 was prominently noted with BMP-7 and less than BMP-6 (p5.0211). The down regulation at day 14 was noted significantly different with autograft and BMP-6. Sclerostin - The temporal sequence is similar to that seen with BMP-3 in that a delayed upregulation of Sclerostin is noted with exposure to BMP-7 at day 14 with down regulation when compared to control at all other time points. Sclerostin expression was significantly greater for BMP-7 than BMP-6 at day 14 (p5.0381). Follistatin – The most noteworthy response of follistatin to all treatments occurred at day 3. Autograft, BMP-6 and BMP-7 all induced comparable upregulation of follistatin while at the other time points relative expression levels were similar to control sites. Chordin - Chordin gene expression showed similar upregulation among each BMP and autograft with minimal increase exhibited at day one and maximizing at day 3. The elevated expression remained relatively high at day 7 except for BMP-7 which returned to near control levels. Slight nonsignificant increases were furthermore noted at day 14 with all treatment groups. Twisted Gastrulation Homologue (TSG) – Slight increase in expression of TSG was noted with both BMP treatments compared with autograft from day 3 through day 14. CONCLUSIONS: The results of the gene expression analysis confirm a characteristic temporal response of each of the antagonists as well as response to differing BMP and autogenous graft. There appears to be a highly regulated negative feedback process that interestingly is stimulated differently by BMP and autograft - greater upregulation of inhibitors is noted with osteoinductive proteins. In addition, there is suggestion that BMP’s may also stimulate this antagonistic response to differing degrees. These results will be critical when approaches are made to knock down any of these inhibitors of bone formation in terms of when they are present within a developing fusion mass. FDA DEVICE/DRUG STATUS: This abstract does not discuss or include any applicable devices or drugs. doi: 10.1016/j.spinee.2011.08.090

62. Combined Transplantation of Human Neural Stem Cells and Human Mesenchymal Stem Cells Following Spinal Cord Injury Ivan Cheng, MD; Stanford University, Menlo Park, CA, USA BACKGROUND CONTEXT: Spinal cord injury is a significant cause of morbidity and mortality in traumatized patients, leading to long-term functional impairment and disability. Functional recovery is limited due to the inability of the spinal cord to restore demyelinated axonal pathways. Current treatment modalities are highly debated given the lack of consistent improvement with substantial risks. The utilization of stem cell therapy is an attractive alternative to regenerate damaged axons given their pluripotent nature. PURPOSE: Acute transplantation of human fetal neural stem cells (hNSCs) has previously shown significant functional recovery after spinal cord contusion in a rat model. Other studies have indicated that human mesenchymal stem cells (hMSCs) can home to areas of damage, and in particular, they have been shown to be able to cross the blood brain barrier and migrate throughout the central nervous system. We hypothesized that acute administration of hMSCs would create a more hospitable environment at the site of injury, and when combined with subacute treatment using hNSCs would enhance functional recovery in the rat model. STUDY DESIGN/SETTING: This was a prospective randomized controlled trial. PATIENT SAMPLE: Subjects utilized were 24 adult female Long-Evans hooded rats.

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OUTCOME MEASURES: Subjects were assessed following injury and then weekly for 6 weeks. Functional assessment was measured using the BBB Locomotor Rating Score. Data were analyzed using ANCOVA and mixed effect regression analyzing the effect of experimental condition on outcome slope. METHODS: All subjects underwent a posterior exposure and laminectomy at the T10 level. A moderate spinal cord contusion at the T10 level was induced by use of the Multicenter Animal Spinal Cord Injury Study Impactor with a 10 g weight dropped from a height of 25 mm. 4 groups were identified for this study (see Table). Group 1 received hMSCs intravenously (IV) immediately after spinal cord injury (acute) and returned 1 week later (subacute) for injection of hNSC directly at the site of injury. Group 2 received hMSC IV acutely and cell media directly subacutely. Group 3 received cell media IV acutely and hNSC subacutely. Group 4 received cell media IV acutely and cell media subacutely. RESULTS: 6 subjects were included in each group. A statistically significant functional improvement was seen in the MSCþNSC group and the NSC-only group compared with control (p5.027 and 0.042 respectively), but the MSC-only group did not demonstrate a significant improvement over control (p5.145, see Figure). Comparing the MSCþNSC group and the NSC-only group, there was no significant difference (p5.357). CONCLUSIONS: The subacute transplantation of hNSCs into the contused spinal cord of a rat led to significant functional recovery of the spinal cord when injected either with or without the acute IV administration of hMSCs. Neither the addition of hMSC to hNSC treatment nor the delivery of hMSCs-alone resulted in significant functional improvement. Although the use of hMSCs in this study did not prove to be useful, future study may be necessary to determine if the route and timing of administration may make a significant difference. FDA DEVICE/DRUG STATUS: This abstract does not discuss or include any applicable devices or drugs. doi: 10.1016/j.spinee.2011.08.091

63. Interleukin 1-beta (IL-1) Exposure Significantly Upregulates Neurotrophin Gene Expression for Brain-Derived Neurotrophic Factor, Neurotrophin 3 and Neuropilin 2 During 3D Culture of Human Annulus Cells Helen Gruber, PhD1, Gretchen Hoelscher1, Synthia Bethea2, Edward Hanley, MD3; 1Carolinas Medical Center, Charlotte, NC, USA; 2 Orthopaedic Biology Research, Charlotte, NC, USA; 3Carolinas Medical Center, Department of Orthopedics, Charlotte, NC, USA BACKGROUND CONTEXT: Although the degenerating disc is considered to be the key source of pain in patients with discogenic low back pain, the relationship between disc cells, nerves, and pain production in the disc is poorly understood. Neurotrophins, signaling molecules involved in the survival, differentiation, migration, and neurite outgrowth of central and peripheral neurons, are now known to be expressed in a variety of non-neuronal tissues including the disc, and function in the adult with primary nociceptive neurons. PURPOSE: To test the hypothesis that in vitro exposure of human disc cells to the proinflammatory cytokine IL-1 will elevate neurotrophin expression levels. STUDY DESIGN/SETTING: Institutional Review Board-approved study using human disc cells in vitro. PATIENT SAMPLE: Disc specimens obtained at surgery. Cells were isolated from three Thompson grade IV discs, and one grade III disc. OUTCOME MEASURES: Gene expression levels from microarray including correction for false discovery rates. METHODS: Human annulus cells derived from surgical disc specimens were cultured in 3D within a collagen sponge for 9 days with media changes in the presence of control (minimal essential medium supplemented with 20% FBS (MEM20) (controls) or were treated with 102 pM IL-1 beta; cells then grew for 5 days without feeding. Experiments

All referenced figures and tables will be available at the Annual Meeting and will be included with the post-meeting online content.