Biodegradable polymer scaffolds combined with bioactive components which accelerate osteogenesis and angiogenesis have promise for use in clinical bone defect repair. The preclinical acute toxicity evaluation is an essential assay of implantable biomaterials to assess the biosafety for accelerating clinical translation. We have successfully developed magnesium (Mg) particles and beta-tricalcium phosphate (β-TCP) for incorporation into poly(lactic-co-glycolic acid) (PLGA) porous composite scaffolds (PTM) using low-temperature rapid prototyping three-dimensional-printing technology. The PTM scaffolds have been fully evaluated and found to exhibit excellent osteogenic capacity for bone defect repair. The preclinical evaluation of acute systemic toxicities is essential and important for development of porous scaffolds to facilitate their clinical translation. In this study, acute systemic toxicity of the PTM scaffolds was evaluated in mice by intraperitoneal injection of the extract solutions of the scaffolds. PTM composite scaffolds with different Mg and β-TCP content (denoted as PT5M, PT10M, and PT15M) were extracted with different tissue culture media, including normal saline, phosphate-buffered saline , and serum-free minimum essential medium , to create the extract solutions. The evaluation was carried out following the National Standard. The acute toxicity was fully evaluated through the collection of extensive data, including serum/organs ion concentration, fluorescence staining, and in vivo median lethal dose measurement. Mg in major organs (heart, liver, and lung), and Mg ion concentrations in serum of mice, after intraperitoneal injection of the extract solutions, were measured and showed that the extract solutions of PT15M caused significant elevation of serum Mg ion concentrations, which exceeded the safety threshold and led to the death of the mice. In contrast, the extract solutions of PT5M and PT10M scaffolds did not cause the death of the injected mice. The median lethal dose of Mg ions in vivo for mice was determined for the first time in this study to be 110.66 mg/kg, and the safety level of serum magnesium toxicity in mice is 5.4 mM, while the calcium serum safety level is determined as 3.4 mM. The study was approved by the Animal Care and Use Committee of Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences (approval No. SIAT-IRB-170401-YGS-LYX-A0346) on April 5, 2017. All these results showed that the Mg ion concentration of intraperitoneally-injected extract solutions was a determinant of mouse survival, and a high Mg ion concentration (more than 240 mM) was the pivotal factor contributing to the death of the mice, while changes in pH value showed a negligible effect. The comprehensive acute systemic toxicity evaluation for PTM porous composite scaffolds in this study provided a reference to guide the design and optimization of this composite scaffold and the results demonstrated the preclinical safety of the as-fabricated PTM scaffold with appropriate Mg content, strongly supporting the official registration process of the PTM scaffold as a medical device for clinical translation.
Keywords: acute systemic toxicity; clinical translation; magnesium; median lethal dose; porous composite scaffolds.