Effect of C5a Receptor Antagonist on the Osteogenic Differentiation of Dental Pulp Stem Cells
Xiaohan Tan1,2, Junlong Hu3, Junhui Li1
1Department of Prosthodontics, the Second Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang, P.R.China.
Objective:
Complement component 5a (C5a) binding to its receptor (C5aR) can mediate pro-inflammatory immune responses, which impairs osteogenic differentiation of dental pulp stem cells (DPSCs). The present study aimed to evaluate the regulatory effect of C5aR antagonist (W54011) on the osteogenic differentiation of DPSCs in an in vitro inflammatory microenvironment and its reparative effect on in vivo LPS-induced inflammatory mandibular defects, to provide novel strategies for bone regeneration under inflammatory conditions.
Materials And Methods:
In vitro, a DPSCs inflammatory model was established with 1.0 μg/mL LPS for 48 h, and cells were divided into four groups (LPS + W54011 + OIM, OIM, W54011 only, blank) with W54011 at 1.0 μg/mL. Each assay included 3 biological and 3 technical replicates, and osteogenic differentiation was assessed via Alkaline Phosphatase (ALP) staining, alizarin red S (ARS) staining, RT-qPCR, and western blotting for OCN, Osterix, and RUNX2. In vivo, an LPS-induced inflammatory mandibular defect model was constructed in Sprague-Dawley (SD) rats; after 2 weeks of inflammation, defects were implanted with W54011-loaded Bio-Oss bone powder, and healing was evaluated by micro-CT, H-E staining, and immunohistochemistry for IL-6, OCN, and RUNX2.
Results:
In vitro, all 4 groups showed positive ALP and mineralized nodule staining. Molecular biology methods found differential OCN, Osterix and RUNX2 expression at day 7, 14 and 21, with the LPS + W54011 + OIM group having higher osteogenesis markers at certain times (p < 0.05, one-way ANOVA with Bonferroni post-hoc test). In vivo, Micro-CT and H&E staining revealed that the defect site implanted with W54011-loaded Bio-Oss (treatment group) exhibited higher bone density and denser new bone formation compared to the control groups. However, due to the small sample size, these results were considered preliminary qualitative observations without statistical analysis.
Conclusion:
1. LPS (1 μg/ml) stimulation for 48 h can be used to establish a DPSCs inflammation model. 2. W54011 can promote the osteogenic differentiation of DPSCs in an inflammatory environment in vitro. 3. Preliminary in vivo data suggest that W54011-loaded Bio-Oss may facilitate the healing of inflammatory mandibular defects, warranting further investigation with larger samples.
