Evaluation of mechanical, histological and biological properties of generations of concentrated growth factor: an in vitro and ex vivo study

Abstract

Background
Concentrated Growth Factor (CGF) is an autologous platelet-rich biomaterial composed of fibrin, leukocytes and various growth factors and has gained increasing attention for its role in promoting tissue regeneration in dentistry. Despite its favourable bioactivity and clinical potential, its rapid degradation, typically occurring within two to three weeks, limits its effectiveness in applications requiring long-term scaffold integrity. In response, recent developments have introduced modified forms of CGF, including heat-denatured albumin (ALB-CGF) and silver nanoparticles (ALB-CGF-SNP), to enhance its degradation resistance and provide additional antimicrobial properties. This research integrates a systematic review to evaluate the efficacy of CGF in regenerative dentistry and experimental studies to investigate the growth factor release, degradation, mechanical, histological and antibacterial properties of CGF and its modified formulations.

Methods
A systematic review and meta-analysis were conducted, including randomised controlled clinical trials published between July 2013 and July 2023. Studies were identified from databases including MEDLINE, OVID, Scopus, Cochrane Library, Google Scholar, Web of Science and Elsevier. The quality of the trials was assessed using the Cochrane Risk of Bias tool and JADAD scale; data were synthesised using RevMan 5.4 software. In parallel, five in vitro and ex vivo experiments were carried out using blood samples collected from 15 healthy volunteers. CGF, ALB-CGF and ALB-CGF-SNP were prepared using the Medifuge MF200 centrifuge and albumin modification was performed via the APAG heating device. Growth factor release was measured using ELISA at intervals of 1, 7, 14, 30 and 60 days for PDGF-AB, VEGF, TGF-β1 and EGF. Degradation studies included both non-enzymatic and enzymatic conditions. Non-enzymatic degradation was conducted in DMEM and quantified via percentage weight loss and protein concentration using the BCA assay. Enzymatic degradation was induced using trypsin and EDTA, followed from day 1 to day 21 and evaluated through qualitative (subjective) assessment and protein concentration via BCA assay. Mechanical behaviour was assessed through tensile strength assays, which measured the Ultimate Tensile Strength (UTS), Modulus of Toughness, Young’s Modulus and strain at break. Cellular content, including WBCs, RBCs, platelets, neutrophils, eosinophils, basophils, 21lymphocytes and monocytes, was quantified using a Sysmex haematology analyser. Structural morphology and internal porosity were examined through surface and cross-sectional evaluation using Scanning Electron Microscopy (SEM). Antibacterial testing was performed against Staphylococcus aureus using colony-forming unit per millilitre (CFU mL⁻¹) counting at 0, 1, 4 and 24 hours. Statistical analysis was performed using one-way ANOVA and Welch ANOVA, with post hoc comparisons through Tukey’s and Dunnett’s tests, depending on the homogeneity of variance.

Results
The systematic review revealed that CGF, when combined with bone grafts, significantly improved periodontal intrabony defect outcomes, with a mean reduction in defect depth of 1.41 mm and a gain of 0.55 mm in clinical attachment level (P < 0.00001 and P = 0.002, respectively). In cases of gingival recession, CGF combined with CAF produced better outcomes than CAF alone, increasing keratinised tissue width by 0.41 mm and gingival thickness by 0.26 mm (P < 0.00001), although it remained less effective than CTG with CAF, with reductions in root coverage of -15.09% and gingival thickness of -0.50 mm (P < 0.0001). Additional clinical improvements were observed in GBR, alveolar ridge preservation, sinus elevation, dental implant and third molar post-extraction healing. In the experimental studies, CGF showed a characteristic burst release of growth factors, particularly on day 1, with TGF-β1 peaking at 15,398 pg mL⁻¹. By day 14, ALB-CGF demonstrated higher VEGF (448.4 pgmL⁻¹ vs. 296.4 pg mL⁻¹, P = 0.004) and TGF-β1 (5800 pg mL⁻¹ vs. 3519 pg mL⁻¹, P = 0.004) compared to CGF, while no significant differences were observed in PDGF-AB (P = 0.368) and EGF (P = 0.397). ALB-CGF-SNP exhibited a delayed release profile, maintaining the highest concentrations of VEGF and EGF by day 30 (P < 0.001), with TGF-β1 also significantly sustained (P < 0.00001). Non-enzymatic degradation analysis showed the highest degradation in CGF, with 84.2% weight loss by day 60, compared to 68.63% in ALB-CGF and 45.2% in ALB-CGF-SNP (P < 0.01). In enzymatic degradation, ALB-CGF and ALB-CGF-SNP retained significantly lower protein concentrations throughout the 21-day period (P < 0.001). In terms of mechanical performance, CGF outperformed other groups in UTS (95.6 kPa), Modulus of Toughness (55.55 kJ m⁻³) and Young’s Modulus (75.73 kPa), all with P < 0.01, while strain at break showed no significant variation (P > 0.90). Cellular analysis revealed that CGF had the highest platelet and WBCs content compared to the WB, with fold differences of 3.11 and 2.25, respectively, compared to other formulations (P < 0.01). SEM examination 22revealed that CGF exhibited dense fibrin matrices, ALB-CGF displayed more open and porous architecture in dense protein structure and ALB-CGF-SNP showed irregular fibrin structureswith less dense protein structure. Porosity and density differences were evident in both surface and cross-sectional imaging. The antibacterial evaluation demonstrated that ALB-CGF-SNP had the strongest inhibitory effect against Staphylococcus aureus compared with other CGF formulations, achieving a statistically significant reduction in CFU mL⁻¹ at 24 hours (P < 0.0001). However, it did not reach the outcomes of methicillin.

Conclusion
This research integrates systematic review and experimental evidence to comprehensively evaluate CGF and its modifications for use in regenerative dentistry. While CGF demonstrates superior mechanical strength and high initial growth factor release, its rapid degradation limits its sustained use. ALB-CGF and ALB-CGF-SNP modifications significantly improve scaffold longevity and bioactivity, with ALB-CGF-SNP showing the most favourable profile in terms of sustained growth factor release, degradation resistance and antibacterial efficacy. These findings support the use of modified CGF in extended applications such as periodontal regeneration, GBR and implantology, while also underscoring the continuing relevance of unmodified CGF in procedures where high mechanical integrity is immediately required. Further clinical and pre-clinical trials with standardised protocols are recommended to validate these findings and ensure broader clinical adoption.

Date of Award	Oct 2025
Original language	English
Supervisor	Deborah Lowry (Supervisor), Paul McCarron (Supervisor), Mohamed Elnaem (Supervisor) & Ziad Al-Ani (Supervisor)