Abstract:

BACKGROUND: Xenograft bone materials have been limited in clinical application, mainly due to their strong immunogenicity, but immunogenicity evaluation on the xenograft bone materials is based on the clinical observation or through indirect determination by histopathological observation. Up to now, there is no quantitative indicator or industry "gold standard".
OBJECTIVE: To observe the in vitro activation and proliferation of peripheral blood lymphocytes during co-culturing with bone material and to prepare for the next specific immunogenicity experiments.
METHODS: The hydroxyapatite (2 g/mL) was selected as negative control, the phytohemagglutinin was considered as positive control. The CCK-8 method was used to detect the effect of 0.5, 1 and 2 g/mL cortical bone meal (treated with degreasing, acellular and multi-faceted antigen processing) and 1 g/mL disinfected cortical bone meal on the proliferation of lymphocytes; flow cytometry was used to detect the effect of CD69 labeled bone materials on the in vitro activation of lymphocytes.
RESULTS AND CONCLUSION: There was no significant difference of proliferation of lymphocytes between 0.5, 1 and 2 g/mL cortical bone meal groups and hydroxyapatite group (P > 0.05), while the proliferation of lymphocytes in disinfected cortical bone meal group and phytohemagglutinin positive group was higher than that in the hydroxyapatite group (P < 0.05, 0.01). The low expression rate of CD69 of lymphocytes in the hydroxyapatite group indicated that the T cells were in the static status; there was no significant difference in expression rate of CD69 of lymphocytes between 0.5, 1 and 2 g/mL cortical bone meal groups and hydroxyapatite group (P > 0.05); but the expression rate of CD69 of lymphocytes co-cultured with disinfected cortical bone meal was significantly increased; the expression of CD69 of lymphocytes in the phytohemagglutinin positive group was most obvious. The processed bone meal has no immunogenic effect on lymphocytes.