The paradoxical effect of cobalt on biological processes has aroused controversy regarding the application of cobalt-based biomaterials in bone regeneration. Tuning the dose range of cobalt ions may be a valid strategy to resolve the controversies about cobalt use for orthopedic applications. Recent progress in bone biology has highlighted the effects of multisystem cooperation (especially of osteoimmune, skeletal, and vascular systems) on bone dynamics. Before the application of this dose-tuning strategy, a deeper understanding of its dose-dependent effect on the cooperation of osteoimmune, skeletal, and vascular systems is needed. However, due to the difficulties with investigating the interaction of multiple systems in vitro, the multimodal effects of cobalt on bone homeostasis were investigated here, in an in vivo scenario.

Methods: In vitro CCK8 assay and cytoskeletal staining were preformed to detecte the cell cytotoxic reaction in response to 0.1-100 ppm cobalt stimulation. Blood clot containing 0.1 to 5 ppm of cobalt were implanted in the rat calvarium defect. The gene profile of osteoimmune, skeletal, and vascular system as well as the systemic toxicity were evaluated via RT-qPCR, histological analysis and inductively coupled plasma mass spectrometry. The bone regeneration, osteoclastogenesis and vascularization were assessed by micro-ct and histological analysis.

Results: Cobalt concentration below 5 ppm did not cause cell toxicity in vitro. No systemic toxicity was observed in vivo at 0.1-5 ppm cobalt concentration. It was found that the early cytokine profiles of the multiple interacting systems were different in response to different cobalt doses. Most of the anti-inflammatory, osteogenic, and proangiogenic factors were upregulated in the 1 ppm cobalt group at the early stage. In the late stage, the 1ppm group was most superior in bone regenerative effect while the 5 ppm group displayed the strongest osteoclastogenesis activity.

Conclusions: The 1 ppm concentration of cobalt yielded the most favorable cooperation of the osteoimmune, skeletal, and vascular systems and subsequently optimal bone regeneration outcomes. Tuning the cobalt dose range to manipulate the cooperation of osteoimmune, skeletal, and vascular systems could be a promising and valuable strategy to prevent paradoxical effects of cobalt while preserving its beneficial effects.

钴对生物过程的矛盾作用引起了关于钴基生物材料在骨再生中应用的争议。调整钴离子的剂量范围可能是解决骨科应用中钴使用争议的有效策略。骨生物学的最新进展强调了多系统协作（特别是骨免疫、骨骼和血管系统）对骨动力学的影响。在应用这种剂量调整策略之前，需要更深入地了解其对骨免疫、骨骼和血管系统协作的剂量依赖性影响。然而，由于在体外研究多个系统的相互作用存在困难，因此在体内研究了钴对骨稳态的多模式影响。

方法：采用体外CCK8测定和细胞骨架染色来检测细胞对0.1-100 ppm钴刺激的细胞毒性反应。将含有 0.1 至 5 ppm 钴的血凝块植入大鼠颅骨缺损处。通过RT-qPCR、组织学分析和电感耦合等离子体质谱法评估骨免疫、骨骼和血管系统的基因谱以及全身毒性。通过显微CT和组织学分析评估骨再生、破骨细胞生成和血管化。

结果：钴浓度低于 5 ppm在体外不会引起细胞毒性。在 0.1-5 ppm 钴浓度下，体内未观察到全身毒性。研究发现，多个相互作用系统的早期细胞因子谱对不同钴剂量的反应是不同的。大多数抗炎、成骨和促血管生成因子在早期阶段在1 ppm钴组中上调。在后期，1ppm组的骨再生效果最为优异，而5ppm组的破骨细胞生成活性最强。

结论：1 ppm 浓度的钴可实现骨免疫、骨骼和血管系统最有利的配合，从而实现最佳的骨再生结果。调整钴剂量范围以操纵骨免疫、骨骼和血管系统的合作可能是一种有前途且有价值的策略，可以防止钴的矛盾效应，同时保留其有益作用。