197. In vitro and in vivo assessment of bone regeneration after local steroid administration using a rodent posterolateral fusion model

Abstract

BACKGROUND CONTEXT Local intraoperative steroid administration has been shown to reduce soft tissue swelling and dysphagia after cervical spinal fusion. However, the theoretical negative impact of steroid compounds on bone healing has prevented this practice from becoming routine. PURPOSE This study aims to evaluate the effect of local steroid administration on bone regeneration utilizing an established rodent posterolateral fusion model. STUDY DESIGN/SETTING Preclinical animal model PATIENT SAMPLE Female Sprague-Dawley rats (age 12-16 weeks) OUTCOME MEASURES Radiographs, Fusion Scoring, Micro-Computed Tomography, Histology, Gene Expression Analysis METHODS For in vitro assessment, human bone marrow derived mesenchymal stem cells (hBM-MSCs) were grown in media with INFUSETM and different amounts of steroid: vehicle control (no steroid), therapeutic triamcinolone (0.5 mg/kg), and supratherapeutic triamcinolone (2.5 mg/kg). Gene expression analysis of pro-osteogenic markers (RUNX2, Osterix, ALPL, COL1A1, Osteocalcin, Osteopontin), alkaline phosphatase (ALP) activity assays, and alizarin red staining were used to assess for differences in osteogenic differentiation among the treatment groups. For in vivo assessment, 45 female Sprague-Dawley rats underwent bilateral L4-L5 posterolateral fusion (PLF), receiving subtherapeutic doses of INFUSETM (1.0 mg per animal). Additionally, three groups (n = 15 animals each) received either: no steroid (vehicle control), therapeutic triamcinolone (0.5 mg/kg), or supratherapeutic triamcinolone (2.5 mg/kg), administered locally. Spines were harvested at 8 weeks postoperation. Blinded manual palpation by three independent reviewers was used to determine a fusion score, wherein 0 = no fusion, 1 = unilateral bridging bone, and 2 = bilateral bridging bone. The fusion rate was calculated based on the fusion score, in which a score ≥1 indicates successful fusion. New bone volume was quantified with microCT imaging. Representative samples underwent hematoxylin and eosin staining (H&E) for evaluation of fusion. RESULTS All pro-osteogenic transcripts were significantly elevated in samples treated with INFUSETM. Among INFUSETM groups, significant differences in RUNX2, Osterix, COL1A1, and Osteocalcin expression were noted between the different treatment groups: except for Osteocalcin, expression was significantly increased with therapeutic or supra-therapeutic triamcinolone relative to vehicle controls at one or more of the time points tested. ALP activity also increased with triamcinolone treatment relative to controls after 3 days, 3 week, and 3 weeks in culture, and mineral deposition was comparable between all groups. For in vivo assessment, there were no significant differences in average fusion score (FS), fusion rate (FR), or new bone volume (BV) between the control (FS: 0.62 ± 0.59, FR: 38%, BV: 32.4 ± 22.4 mm3), therapeutic steroid (FS: 1.09 ± 0.81, FR: 57%, BV: 40.1 ± 23.2 mm3), and supratherapeutic steroid (FS: 0.58 ± 0.58, FR: 33%, BV: 25.5 ± 21.8 mm3) groups. CONCLUSIONS Local intraoperative steroid, at both therapeutic and supratherapeutic doses, did not have a significant negative effect on multiple markers of bone regeneration in both in vitro and in vivo models. These data suggest that the benefits of local steroid may outweigh any theoretical risk of poor bone healing. FDA DEVICE/DRUG STATUS rhBMP-2 (INFUSE) [non-human use] (Investigational/Not approved)