Introduction: Adipose tissue has been known for some time to contain regenerative cells. These regenerative cells are able to differentiate into a nucleus pulposus-like phenotype when exposed to environmental factors similar to disc. In an effort to develop a clinical option for cell placement and assess the response of the cells to the post-surgical milieu, adipose-derived cells were collected, concentrated, and transplanted under fluoroscopic guidance directly into a surgically damaged disc in a dog model.

Methods: After IACUC approval 12 dogs, 2 years of age, were obtained. Adipose cells were harvested from the super-scapular region of the neck (scruff) and adherent cells separated, collected, and labeled with DAPI. Three lumbar intervertebral disc levels in each dog underwent a partial nucleotomy; other levels served as non-operated controls. Levels of intervention as well as the regimen of treatment were dually randomized. Three interventions were used in this study; adipose-derived cells in hyaluronic acid (HA) carrier, HA alone, or no intervention. All deliveries were guided by fluoroscopy. The dogs were radiographed, received MRI scans and then euthanized by 12 months. The disc tissue was harvested from the lumbar spine in each dog. Cells labeled with DAPI prior to implantation were evident in the tissue. Matrix composition was assessed for aggrecan, Types I and II collagen by both RT-PCR (Table I) and ELISA (Table II) to assess and compare matrix regeneration. mRNA and protein from each level are presented with respect to normal values defined as the 100 percent expression.

Results: The results can be summarized as:

the regenerative cells are viable following implantation.

Discussion: This study provides evidence that adipose derived cells might offer a reliable source of regenerative potential capable of bio-restitution. The span of this study was sufficient to show that freshly isolated adipose-derived cells can be transplanted percutaneously and will survive the trauma associated with post-surgical inflammation to remain viable and produce appropriate, tissue-specific matrix.

Study Design: Experimental study in dogs:

Objective: To assess the efficiency of disc chondrocyte transplantation in a canine model.

Summary of Background Data: Conventional clinical treatments of intervertebral disc herniation and degeneration are focused on excision of damaged tissue, stabilization, and spinal fusion. The development and refinement of cell-based therapeutics for tissue regeneration and repair have spawned a multitude of applications including autologous disc chondrocyte transplantation. For clinical application the efficiency of disc chondrocyte transplantation was assessed using a pre-clinical canine model to show the technical feasibility and biological relevance for disc repair and retardation of disc degeneration. This report examined the protein expression of transplanted disc chondrocytes and their role in the clinically observed disc repair following autologous disc chondrocyte transplantation.

Methods: The nucleus and inner annulus were sampled from four skeletally-mature dogs by micro-discectomy. Disc chondrocytes were isolated and propagated under GMP validated conditions including completely autologous serum conditions. Two months later, the cultured cells were transplanted through the contralateral side of experimental discs after testing complete healing of the annulus by measuring intradiscal pressure stability. After seven months the animals were humanely killed. One half of the vertically halved lumbar spines were embedded in paraffin and sections were analysed histologically and immunohistochemically.

Results: Histological examinations revealed large clusters of cells within the nucleus area of the treated discs. Cells within these cell clones were found to be viable and surrounded by de novo synthesized matrix as evidenced by a distinct histological staining and immunohistochemical expression pattern. A disc-specific expression of collagen type I and II and hyaline-specific proteoglycans was observed indicating the regenerative and reconstructive capacity of the transplanted disc chondrocytes.

Conclusions: These results indicate the contribution of transplanted disc chondrocytes to the observed clinical success of this cell-based therapy.