Conditioned medium of intervertebral disc cells inhibits osteogenesis on autologous bone-marrow-derived mesenchymal stromal cells and osteoblasts

Chen, Shuimu; Croft, Andreas S.; Bigdon, Sebastian; Li, Zhen; Albers, Christoph E; Crump, Katherine B.; Bermudez, Paola; Gantenbein, Benjamin (2024). Conditioned medium of intervertebral disc cells inhibits osteogenesis on autologous bone-marrow-derived mesenchymal stromal cells and osteoblasts (Unpublished). In: 50th International Society for the Study of the Lumbar Spine Annual Meeting 2024. Milano, Italy. 29 May 2024.

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INTRODUCTION: Low back pain (LBP) is a significant global burden and is associated with the degeneration of the spine and human intervertebral discs (hIVD). Current surgical treatment for hIVD degeneration is the removal of the affected tissue with a cage to promote spinal fusion and relieve discomfort. Despite progress in the treatment of LBP, however, in ~30% of all cases this procedure ends in non-fusion and painful pseudo-athrosis after operation. Previous research from our laboratory showed morphogenetic protein (BMP) antagonists secreted by the intervertebral disc have been potentially associated with inhibiting the process of osteogenesis. However, the co-cultured cells did not come from the same donor. In this study, we investigated the hypothesis that IVD cells secrete BMP inhibitors that inhibit osteogenesis in autologous osteoblast (hOB) and bone marrow mesenchymal stem cell (hMSC).
METHODS: Conditioned Medium (CM) collected from primary hIVD cells in 3D alginate culture was co-cultured with seven donor-matched hOB and hMSC in 2D culture. Osteogenesis after 10 days was then quantified at the transcript level using qPCR to measure the expression of bone makers and BMP antagonists, and at the protein level by alkaline phosphatase (ALP) activity. Additionally, they were evaluated histologically by alizarin red (ALZR) staining on Day 21. The relationship between ALP activity, osteogenesis and Noggin expression in hOB or hMSC or hIVD was investigated to uncover the potential causes.
RESULTS: ALP activity significantly decreased and the formation of calcium deposits in alizarin red staining was inhibited after culture with CM derived from hIVD. Interestingly, less changes of bone makers and BMP inhibitors’ expression was found in hOB or hMSC on Day 10. Noggin was relatively higher expressed (Average fold change: AF, 6.9; CEP, 10.0; NP, 6.3; relative to autologous hOB. AF, 2.3; CEP, 3.4; NP, 3.2; relative to autologous hMSC.) in hIVD compared to hOB or hMSC.
DISCUSSION: The up-regulation of Noggin mRNA (and possibly other BMP inhibitors) in residual hIVD tissue after spinal fusion surgery is potentially a potent reason for prevention of successful osteogenesis. Similar results were found previously with allogenic co-cultures. However, in previous study there were merely trends of inhibition. Here we show a significant decrease with autologous donor-matched samples.

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