Extensive H(+) release by bone substitutes affects biocompatibility in vitro testing

Bone substitutes are widespread in orthopedic and trauma surgery to restore critical bony defects and/or promote local bone healing. Cell culture systems have been used for many years to screen biomaterials for their toxicity and biocompatibility. This study applies a human bone marrow cell culture system to evaluate the toxic in vitro effects of soluble components of different bone substitutes, which are already in clinical use. Different specimens of tricalcium phosphates (TCP) (Vitoss, Cerasorb), nondecalcified bovine bone (Lubboc), demineralized human bone matrices (DBM) (Grafton Flex/Putty), and collagen I/III matrix (ACI-Maix) were tested in Dulbecco's modified Eagle's medium (DMEM) and MesenCult culture solution and compared with a biomaterial-free cell culture. Biocompatibility parameters were cell viability evaluated by phase-contrast microscopy and laser flow cytometry, morphology, and the local H(+) release by bone substitutes. There were significant differences (p < 0.05) between the tested biomaterials and culture solutions. Collagen I/III, non-demineralized bovine bone, and TCP materials showed advantages for cell survival over other tested biomaterials (average values of vital cells/mL MesenCult/DMEM: Collagen I/III: 1090/1083; Vitoss: 893/483; Cerasorb: 471/523; Lubboc: 815/410; Grafton Putty: 61/44; Grafton Flex: 149/57). Especially the DBM materials lead to a significant decrease of pH, which is considered to be a major factor for cell death. DMEM culture solution supports cell survival for those bone substitutes that induce an alkaline reaction, whereas MesenCult media promotes cell vitality in biomaterials, which leads to an acidification of culture solution.