Aim: The purpose of this study was to investigate the effects of platelet-rich fibrin (PRF) modified with bioactive radiopacifiers–nanohydroxyapatite (nHA) and dentin chips (DC) on odontoblastic differentiation in human dental pulp cells (HDPCs). Subjects and Methods: PRF was modified with 50wt% of nHA (G bone-SHAG31, Surgiwear Company) and 50 wt% of DC. HDPSCs differentiation and mineralization by the groups ([Group A - Control (Dimethyl sulfoxide), Group B - PRF, Group C – PRF + nHA, Group D – PRF + DC]) were assessed. ELISA was done to quantify the interleukin (IL)-6 and IL-8 cytokine expression. The odontoblastic differentiation was determined by the expression of odontogenesis-related genes and the extent of mineralization using alizarin red S staining. Statistical Analysis Used: One-way ANOVA with post hoc Tukey-honestly significant difference tests were applied to assess the significance among various groups. Results: The level of inflammatory cytokines (IL-6 and IL-8) expression by Group D (PRF + 50 wt% DC) was higher compared to Group B (PRF) and Group C (PRF + 50 wt% DC). Group C (PRF + 50 wt% nHA) induced more mineralization nodules compared to other groups. The integrated density value for the DSPP and DMP-1 protein expression by Group C (PRF + 50 wt% nHA) and Group D (PRF + 50 wt% DC) was higher compared to Group B (PRF). Conclusions: The results suggest that the addition of bioactive radiopacifiers into PRF has a synergistic effect on the stimulation of odontoblastic differentiation of HDPCs, hence inducing mineralization.