OBJECTIVE: Functional and neural tissue recovery has been reported in many animal studies conducted with stem cells. However, the combined effect of cytokines and stem cells has not yet been adequately researched. Here, we analyzed the additive effects of granulocyte colony-stimulating factor (GCSF) on adipose-derived stem cells (ADSCs) infusion in the treatment of acute spinal cord injury (SCI) in rats. METHODS: Four days after intrathecal infusion tubes implantation in Sprague-Dawley rats, SCI was induced with an infinite horizon impactor. In the Sham group (n=5), phosphate-buffered saline was injected 3, 7, and 14 days after SCI. GCSF, ADSCs, and ADSCs with GCSF were injected at the same time in the GCSF (n=8), ADSC (n=8), and ADSC+GCSF groups (n=7), respectively. RESULTS: The ADSC and ADSC+GCSF groups, but not the GCSF group, showed significantly higher Basso-Beattie-Bresnahan scores than the Sham group during 8 weeks (p<0.01), but no significant difference between the ADSC and ADSC+GCSF groups. In the ladder rung test, all four groups were significantly different from each other, with the ADSC+GCSF group showing the best improvement (p<0.01). On immunofluorescent staining (GAP43, MAP2), western blotting (GAP43), and reverse transcription polymerase chain reaction (GAP43, nerve growth factor), the ADSC and ADSC+GCSF groups showed higher levels than the Sham and GCSF groups. CONCLUSION: Our analyses suggest that the combination of GCSF and ADSCs infusions in acute SCI in the rat does not have a significant additive effect. Hence, when combination agents for SCI stem cell therapy are considered, molecules other than GCSF, or modifications to the methodology, should be investigated.
Animals ; Blotting, Western ; Combined Modality Therapy ; Cytokines ; GAP-43 Protein ; Granulocyte Colony-Stimulating Factor ; Mesenchymal Stromal Cells ; Models, Animal* ; Polymerase Chain Reaction ; Rats* ; Rats, Sprague-Dawley ; Reverse Transcription ; Spinal Cord Injuries* ; Spinal Cord* ; Stem Cells*

OBJECTIVE: This study describes a method for inducing spinal cord injuries in dogs by using balloon catheters via laminectomy and the subsequent changes in the electrophysiological response. METHODS: Female Beagle (Orient Bio, Seongnam, Korea) dogs weighing 10 kg at the time of injury were used. Under inhalation anesthesia, a posterior midline approach laminectomy was performed. A silicone balloon catheter (size 6 Fr; Sewoon Medical, Cheonan, Korea) was then inserted into the vertebral canal at the center of T10. The balloon was inflated to the maximum volume for 1, 2, or 3 days. Open field testing was performed for evaluating motor functions of the hindlimbs. Motor evoked potentials (MEPs) induced by electrical and magnetic stimulation were recorded before and after spinal cord injury. RESULTS: Open field testing yielded locomotor scores of 0 or 1 for dogs subjected to compression for 3 days. These dogs showed no obvious improvement throughout the observation period, and the tonus of their hindlimbs was flaccid. In contrast, motor functions of dogs that had experienced compression for 1 or 2 days were variable, and all dogs showed spastic tonus in their hindlimbs. In dogs subjected to after compression for 3 days, electrically stimulated MEPs for the hindlimbs showed a significant amplitude reduction. Further, hindlimb movements were not evoked by magnetic stimulation of the cervical spine and vertex area. CONCLUSION: Compression for 3 days with a balloon catheter is a safe, reproducible, and reliable method for evaluating electrophysiological changes in a dog model of complete spinal cord injury.
Anesthesia, Inhalation ; Animals ; Catheters ; Chungcheongnam-do ; Dogs* ; Evoked Potentials, Motor ; Female ; Gyeonggi-do ; Hindlimb ; Humans ; Laminectomy ; Muscle Spasticity ; Silicones ; Spinal Cord Injuries* ; Spine

OBJECTIVE: Recent studies have shown encouraging progress toward the use of autogenic and allogenic mesenchymal stem cells (MSCs) to arrest, or even lead to partial regeneration in, intervertebral disc (IVD) degeneration. However, this technology is still in its infancy, and further development is required. The aim of this study was to analyze whether rat adipose-derived mesenchymal stem cells (ADMSC) can differentiate towards IVD-like cells after treatment with transforming growth factor beta3 (TGF-beta3) in vitro. We also performed quantitative analysis of gene expression for ADMSC only, ADMSCs treated with TGF-beta3, and co-cultured ADMSCs treated with TGF-beta3. METHODS: ADMSCs were sub-cultured to homogeneity and used in fluorocytometry assays for CD11, CD45, and CD90/Thy1. ADMSCs were differentiated in spheroid culture towards the chondrogenic lineage by the presence of TGF-beta3, dexamethasone, and ascorbate. We also co-cultured pure ADMSCs and nucleus pulposus cells in 24-well plates, and performed immunohistochemical staining, western blotting, and RT-PCR for quantitativeanalysis of gene expression. RESULTS: Results of fluorocytometry were positive for CD90/Thy1 and negative for CD11 and CD45. TGF-beta3-mediated induction of ADMSCs led to the expression of the differentiation markers of intervertebral disc-like cells, such as aggrecan, collagen II, and sox-9. Co-cultured ADMSCs treated with TGF-beta3 showed higher expression of differentiation markers and greater extracellular matrix production compared with ADMSCs treated with TGF-beta3 alone. CONCLUSION: ADMSC treated with TGF-beta3 may be an attractive source for regeneration therapy in degenerative IVD. These findings may also help elucidate the pathologic mechanism of MSC therapy in the degeneration of IVD in vivo.
Adipose Tissue ; Aggrecans ; Animals ; Antigens, Differentiation ; Blotting, Western ; Collagen ; Dexamethasone ; Extracellular Matrix ; Gene Expression ; Intervertebral Disc ; Intervertebral Disc Degeneration ; Mesenchymal Stromal Cells ; Rats ; Regeneration ; Transforming Growth Factor beta3