Impact of Thermal Treatment on Growth Factor Levels in Injectable Horizontal Platelet‐Rich Fibrin: An In Vitro Study

ABSTRACT Background Platelet‐rich fibrin (PRF) is widely recognized for its regenerative properties in periodontal therapy, particularly through the release of endogenous growth factors. Injectable horizontal PRF (H‐PRF), a recent advancement, has gained interest due to its improved cell distribution and structural integrity. The aim of this study was to analyze and compare the levels of growth factors released from injectable H‐PRF subjected to different thermal treatments. Methods Venous blood samples from 16 healthy participants (aged 18–25 years) were collected and divided into four groups: Group I (unheated control), Group II (37°C), Group III (45°C), and Group IV (60°C). Liquid H‐PRF was prepared using horizontal centrifugation (700 RCF, 8 min), followed by controlled heating. Growth factor quantification was performed using the LEGENDplex Human Growth Factor Panel. Solidification and degradation profiles were also recorded. Statistical analyses included ANOVA and Bonferroni post hoc tests (significance at p < 0.05). Results The solidification time decreased significantly with increasing temperature, ranging from 24 min in Group I to 5 min in Group IV. After 24 days, heated H‐PRF gels (especially 45°C and 60°C) retained greater mass, indicating slower degradation. Significant intergroup differences were observed for angiopoietin‐2 ( p < 0.001), PDGF‐AA ( p = 0.002), TGF‐α ( p = 0.036), and VEGF ( p = 0.011), with Group II showing the highest levels of angiogenic factors. No significant differences were noted for EGF, FGF‐basic, HGF, and PDGF‐BB. Conclusion Thermal treatment of H‐PRF at moderate temperatures (particularly 37°C and 45°C) enhances the release of angiogenic growth factors while improving the mechanical stability of the gel. However, the heating protocol will need to be customized based on the intended clinical applications.