BACKGROUND CONTEXT Biologic-based therapies for inhibiting or reversing intervertebral disc (IVD) degeneration have garnered considerable interest, however obtaining a stem cell source remains a resource-intensive hurdle. Strategies to recruit endogenous mesenchymal stem cells (MSCs) to the IVD would assist in circumventing these hurdles. PURPOSE The purpose of this study is to evaluate the ability of three homing chemokines to recruit marrow-derived MSCs to the IVD. METHODS Under IACUC approval, 24 female Lewis rats (n=6/group) underwent anterior, transperitoneal L5/L6 intradiscal injection of hydrogel containing homing chemokine (SDF-1, RANTES, or MCP-1) or hydrogel alone (control). Immediately postop, all rats were administered 1×107 MSCs labeled with CellVue Burgundy, via tail vein injection. In vivo near-infrared (NIR) fluorescence imaging was used to longitudinally track MSC migration to L5/L6 at 1, 2, 7, and 14days postinjection. At each time point, the dorsal lumbar region was shaved to minimize hair-related auto-fluorescence, rats were positioned prone on an imaging bed and NIR fluorescence imaging of the dorsal lumbar region was conducted at 700 nm. Following euthanasia at 14days, the lumbar spine was harvested, dissected, and the anterior aspect was re-imaged using 700 nm NIR. A consistently sized, circular region-of-interest was placed over L5/L6 and mean signal intensity was computed. In vivo and ex vivo imaging were normalized to skin and the L3/L4 disc, respectively, to account for nonspecific signal. Mean signal intensity of chemokine groups was compared to the control group using one-way ANOVA. Post-hoc tests were conducted using the Dunnett correction for multiple comparisons, and all reported P-values are compared to the control group. RESULTS Longitudinal in vivo NIR demonstrates measurable fluorescent signal at L5/L6 at 1day (control: 0.005±0.007, SDF-1: 0.008±0.008, P=0.918; RANTES: 0.017±0.011, P=0.074; MCP-1: 0.007±0.009, P=0.990); however, no significant differences were noted between groups. NIR signal was indistinguishable from skin at 3, 7, and 14day time points for all groups. Ex vivo NIR signal was appreciable at the L5/L6 disc level in all treatment groups, indicating successful migration of tagged MSCs to the target disc. RANTES and MCP-1 treatment groups exhibited significantly higher ex vivo signal compared to control (control: 0.0017±0.0006, SDF-1: 0.0025±0.0003, P=0.054; RANTES: 0.0030±0.0006, P=0.001; MCP-1: 0.0026±0.0006, P=0.017). CONCLUSIONS Following intra-discal injection, in vivo imaging demonstrates early recruitment of labeled MSCs to the L5/L6 IVD, with chemokine groups exhibiting anecdotally higher signal compared to control. Appreciable in vivo signal was only detected at 24hours, suggesting that the highest localization of MSCs to the L5/L6 disc level occurs acutely. Ex vivo imaging, which allows the removal of extraneous tissue and allows antero-posterior image acquisition, revealed notable fluorescent signal at L5/L6, demonstrating retention of labeled MSCs at the L5/L6 IVD at 14days following chemokine injection. Specifically, RANTES was most effective at recruiting MSCs to the IVD compared to control groups and will be used in future studies characterizing therapeutic effects of recruited MSCs in the setting of IVD degeneration. Ongoing histologic analysis will localize tagged cells within specific tissues.
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