GEORG-SCHMORL-PRIZE OF THE GERMAN SPINE SOCIETY (DWG) 2016: Comparison of in vitro osteogenic potential of iliac crest and degenerative facet joint bone autografts for intervertebral fusion in lumbar spinal stenosis.

PURPOSE: The promotion of spinal fusion using bone autografts is largely mediated by the osteoinductive potential of progenitors/mesenchymal stem cells (MSC) that reside in the marrow spaces of cancellous bone. Iliac crest is the common autograft donor site, but its use presents an increased risk for donor site pain, morbidity and infection. Degenerative bone samples harvested during facetectomy might provide an alternative viable source of osteoinductive autografts. In this study, we conducted an intra-individual comparison of the osteogenic potential of isolated low passage MSC from both sources. METHODS: Iliac crest and degenerative facet joints were harvested from eight consecutive patients undergoing transforaminal lumbar interspinal fusion due to lumbar spinal stenosis. MSC were isolated by collagenase digestion, selected by plastic adherence and minimally expanded for downstream assays. Clonogenic and osteogenic potential was evaluated by colony formation assays in control and osteogenic culture medium. Osteogenic properties, including alkaline phosphatase (ALP) induction, matrix mineralization and type I collagen mRNA and protein expression were characterized using quantitative histochemical staining and reverse transcription PCR. Spontaneous adipogenesis was analysed by adipocyte enumeration and gene expression analysis of adipogenic markers. RESULTS: Average colony-forming efficiency in osteogenic medium was equal between iliac crest (38 ± 12%) and facet joint (36 ± 11%). Osteogenic potential at the clonal level was 55 ± 26 and 68 ± 17% for iliac crest and facet joint MSC, respectively. Clonogenic and osteogenic potential were significantly negatively associated with donor age. Osteogenic differentiation led to significant induction of ALP activity in iliac crest (sixfold) and facet joint (eightfold) MSC. Matrix mineralization quantified by Alizarin red staining was increased by osteogenic differentiation, yet similar between both MSC sources. Protein expression of type I collagen was enhanced during osteogenesis and significantly greater in iliac crest MSC. Correspondingly, COL1A2 mRNA expression was higher in osteogenically differentiated MSC from iliac crest. Adipocyte numbers showed significant differences between iliac crest (63 ± 60) and facet joint (18 ± 15) MSC under osteogenic conditions. Negative (GREM1) and positive (FABP4) adipogenic markers were not differentially expressed between sources. CONCLUSION: MSC from iliac crest and degenerative facet joints largely display similar clonogenic and osteogenic properties in vitro. Differences at the molecular level are not likely to impair the osteoinductive capacity of facet joint MSC. Bone autografts from facetectomy would be viable alternatives as bone autografts for intervertebral spinal fusion in lumbar spinal stenosis.

Isolation and Characterization of Human Mesenchymal Stem Cells From Facet Joints and Interspinous Ligaments.

STUDY DESIGN: A descriptive in vitro study on isolation and differentiation of human mesenchymal stem cells (MSCs) derived from the facet joints and interspinous ligaments. OBJECTIVE: To isolate cells from the facet joints and interspinous ligaments and investigate their surface marker profile and differentiation potentials. SUMMARY OF BACKGROUND DATA: Lumbar spinal canal stenosis and ossification of the posterior longitudinal ligament are progressive conditions characterized by the hypertrophy and ossification of ligaments and joints within the spinal canal. MSCs are believed to play a role in the advancement of these diseases and the existence of MSCs has been demonstrated within the ligamentum flavum and posterior longitudinal ligament. The aim of this study was to investigate whether these cells could also be found within facet joints and interspinous ligaments. METHODS: Samples were harvested from 10 patients undergoing spinal surgery. The MSCs from facet joints and interspinous ligaments were isolated using direct tissue explant technique. Cell surface antigen profilings were performed via flow cytometry. Their lineage differentiation potentials were analyzed. RESULTS: The facet joints and interspinous ligaments-derived MSCs have the tri-lineage potential to be differentiated into osteogenic, adipogenic, and chondrogenic cells under appropriate inductions. Flow cytometry analysis revealed both cell lines expressed MSCs markers. Both facet joints and interspinous ligaments-derived MSCs expressed marker genes for osteoblasts, adipocytes, and chondrocytes. CONCLUSION: The facet joints and interspinous ligaments may provide alternative sources of MSCs for tissue engineering applications. The facet joints and interspinous ligaments-derived MSCs are part of the microenvironment of the human ligaments of the spinal column and might play a crucial role in the development and progression of degenerative spine conditions.