3D printing nacre powder/sodium alginate scaffold loaded with PRF promotes bone tissue repair and regeneration.

Bone defects are a common complication of bone diseases, which often affect the quality of life and mental health of patients. The use of biomimetic bone scaffolds loaded with bioactive substances has become a focal point in the research on bone defect repair. In this study, composite scaffolds resembling bone tissue were created using nacre powder (NP) and sodium alginate (SA) through 3D printing. These scaffolds exhibit several physiological structural and mechanical characteristics of bone tissue, such as suitable porosity, an appropriate pore size, applicable degradation performance and satisfying the mechanical requirements of cancellous bone, etc. Then, platelet-rich fibrin (PRF), containing a mass of growth factors, was loaded on the NP/SA scaffolds. This was aimed to fully maximize the synergistic effect with NP, thereby accelerating bone tissue regeneration. Overall, this study marks the first instance of preparing a bionic bone structure scaffold containing NP by 3D printing technology, which is combined with PRF to further accelerate bone regeneration. These findings offer a new treatment strategy for bone tissue regeneration in clinical applications.

Effect of leukocyte-platelet-rich fibrin (L-PRF) on the rate of orthodontic tooth movement and expression of various biomarkers in gingival crevicular fluid.

OBJECTIVES: To assess the outcome of leukocyte-platelet-rich fibrin (L-PRF) on the rate of maxillary canine retraction and its correlation with the levels of Receptor activator of nuclear factor kappa-B ligand (RANKL), osteoprotegerin (OPG), and RANKL:OPG in the gingival crevicular fluid (GCF) during comprehensive orthodontic treatment. SUBJECTS AND METHODS: Eighteen females who required all 1st premolars extraction for the correction of their class I bimaxillary protrusion malocclusions were included. The L-PRF plugs were placed in the experimental side 1st premolar extraction sockets. Canine retraction was performed by sliding mechanics. Canine retraction was assessed from the maxillary study models prepared just before the extraction (T0) and then at 1 week (T1), 2 weeks (T2), 4 weeks (T3), and 8 weeks (T4) after the 1st premolar extraction and placement of L-PRF plugs. The concentrations of RANKL and OPG in the GCF were evaluated at T0, T1, T2, T3, and T4. RESULTS: In experimental sides, the amount of canine retraction was statistically more during the T0-T1, T1-T2, and T2-T3 periods. The mean concentration of RANKL at T1, T2, and T3 was significantly more in the experimental sides. The mean concentration of OPG was significantly less in the experimental sides at T2, T3, and T4. The RANKL:OPG was significantly more in the experimental sides at T1, T2, T3, and T4. No significant correlation was found between amount of canine retraction and concentration of RANKL and OPG and RANKL to OPG ratio in GCF. CONCLUSIONS: The L-PRF accelerated the rate of maxillary canine retraction by 0.28 mm over an 8-week period. The L-PRF favored the local osteoclastogenesis by enhancing the RANKL and suppressing the OPG concentrations. There was no significant correlation between the rate of maxillary canine retraction and expression of RANKL, OPG, and RANKL:OPG in GCF. TRIAL REGISTRATION: The Clinical Trials Registry of India (Reg. No. CTRI/2020/10/028390, Date-13.10.2020).

Bone density and implant stability with advanced platelet-rich fibrin in immediately loaded- implant assisted mandibular overdentures / Densidade óssea e estabilidade dos implantes com fibrina rica em plaquetas avançada em overdentures mandibulares com carga imediata

ABSTRACT Objective: The aim of this study is to determine the effect of Advanced Platelet-Rich Fibrin on bone density and implant stability in immediately loaded- implant-assisted mandibular overdentures (Split-mouth study). Material and Methods: Ten completely edentulous patients received two implants in the mandibular canine region and locator attachments were used to retain immediately loaded- implant mandibular overdentures. Each patient served in two Groups, one Group for each side. One side of the mandible received an implant with topical application of Advanced Platelet-Rich Fibrin in the implant osteotomy site (Group I) and the other site received an implant without application of Advanced platelet-rich fibrin (Group II). Each patient was examined clinically for implant stability using Osstell Mentor device and radiographically by ultra-low dose CT scan to measure bone density around the implant at baseline, three, six months, and one year. Results: There were no statistically significant differences (P>.05) in bone density and implant stability among the studied Groups during one year follow-up period. Conclusion : Advanced Platelet-Rich Fibrin has no effect on bone density and implant stability in immediately loaded implant-assisted mandibular overdenture.(AU)

RESUMO Objetivo: O objetivo deste estudo é determinar o efeito da Fibrina Rica em Plaquetas Avançada na densidade óssea e estabilidade dos implantes em Overdentures mandibulares com carga imediata (estudo de boca dividida). Material e Métodos: Dez pacientes edêntulos foram submetidos à instalação de dois implantes mandibulares na região dos caninos e pilares locator foram utilizados como sistema de retenção para as overdentures mandibulares com carga imediata. Cada paciente participou nos dois grupos, sendo um grupo para cada lado. Um lado da mandíbula recebeu implante com aplicação tópica de Fibrina Rica em Plaquetas Avançada no local do sítio cirúrgico do implante (Grupo I) e o outro local recebeu implante sem aplicação de Fibrina Rica em Plaquetas Avançada (Grupo II). Cada paciente foi examinado clinicamente quanto à estabilidade do implante usando o dispositivo Osstell Mentor e radiograficamente por tomografia computadorizada de ultrabaixa dose para medir a densidade óssea ao redor do implante no início do estudo, três, seis meses e um ano. Resultados: Não houve diferenças estatisticamente significativas (P>0,05) na densidade óssea e na estabilidade do implante entre os grupos estudados durante o período de acompanhamento de um ano. Conclusão: A Fibrina Rica em Plaquetas Avançada não tem efeito na densidade óssea e na estabilidade de implantes em Overdentures mandibulares com carga imediata (AU)

Exploration of proper heating protocol for injectable horizontal platelet-rich fibrin gel.

PURPOSE: Platelet-rich fibrin (PRF) has been proposed as promising biomaterials with the advantages of host accumulation of platelets and leukocytes with entrapment of growth factors and fibrin scaffold. However, limitations including fast resorption rate (~ 2 weeks) restricts its clinical application. Recent studies have demonstrated heating treatment can prolong PRF degradation. Current published articles used the method of 75 °C for 10 min to obtain longer degradation, while few studies investigated the most suitable temperature for heating horizontal PRF. Our present study was to discover and confirm the optimum temperature for heat treatment before obtaining H-PRF gels by investigating their structure, mechanical properties, and bioactivity of the H-PRF gels after heating treatment. METHODS: In the present study, 2-mL upper layer of horizontal PRF was collected and heated at 45 °C, 60 °C, 75 °C, and 90 °C to heat 2-mL upper layer of horizontal PRF for 10 min before mixing with the 2-mL lower layer horizontal PRF. The weight, solidification time and the degradation properties were subsequently recorded. Scanning electron microscopy (SEM) and rheologic tests were carried out to investigate the microstructure and rheologic properties of each H-PRF gel. The biological activity of each H-PRF gel was also evaluated using live/dead staining. RESULTS: H-PRF gel prepared at 75 °C for 10 min had the fast solidification period (over a tenfold increase than control) as well as the best resistance to degradation. The number of living cells in H-PRF gel is greater than 90%. SEM showed that H-PRF gel becomes denser as the heating temperature increases, and rheologic tests also revealed that the heat treatment improved the mechanical properties of H-PRF gels when compared to non-heated control group. Future clinical studies are needed to further support the clinical application of H-PRF gels in tissue regeneration procedures. CONCLUSIONS: Our results demonstrated that the H-PRF gel obtained at 75 °C for 10 min could produce a uniform, moldable gel with a short time for solidification time, great rheologic behavior and, high percent of live cells in PRF gel. A promising use of the commonly utilized PRF gel was achieved facilitating tissue regeneration and preventing degradation.

Lyophilised Platelet-Rich Fibrin: Physical and Biological Characterisation.

BACKGROUND: Platelet-rich fibrin (PRF) has gained popularity in craniofacial surgery, as it provides an excellent reservoir of autologous growth factors (GFs) that are essential for bone regeneration. However, the low elastic modulus, short-term clinical application, poor storage potential and limitations in emergency therapy use restrict its more widespread clinical application. This study fabricates lyophilised PRF (Ly-PRF), evaluates its physical and biological properties, and explores its application for craniofacial tissue engineering purposes. MATERIAL AND METHODS: A lyophilisation method was applied, and the outcome was evaluated and compared with traditionally prepared PRF. We investigated how lyophilisation affected PRF's physical characteristics and biological properties by determining: (1) the physical and morphological architecture of Ly-PRF using SEM, and (2) the kinetic release of PDGF-AB using ELISA. RESULTS: Ly-PRF exhibited a dense and homogeneous interconnected 3D fibrin network. Moreover, clusters of morphologically consistent cells of platelets and leukocytes were apparent within Ly-PRF, along with evidence of PDGF-AB release in accordance with previously reports. CONCLUSIONS: The protocol established in this study for Ly-PRF preparation demonstrated versatility, and provides a biomaterial with growth factor release for potential use as a craniofacial bioscaffold.

Platelet-rich fibrin-loaded PCL/chitosan core-shell fibers scaffold for enhanced osteogenic differentiation of mesenchymal stem cells.

Here, we fabricated the platelet-rich fibrin (PRF)-loaded PCL/chitosan (PCL/CS-PRF) core-shell nanofibrous scaffold through a coaxial electrospinning method. Our goal was to evaluate the effect of CS-RPF in the core layer of the nanofibrous on the osteogenic differentiation of human mesenchymal stem cells (HMSCs). The elastic modulus of PCL/CS-PRF core-shell scaffold (44 MPa) was about 1.5-fold of PCL/CS scaffold (25 MPa). The specific surface area of the scaffolds increased from 9.98 m2/g for PCL/CS scaffold to 16.66 m2/g for the PCL/CS-PRF core-shell nanofibrous scaffold. Moreover, the release rate of PRF from PCL/CS-PRF nanofibrous scaffold was measured to be 24.50% after 10 days which showed slow and sustained release of PRF from the nanofibrous. The formation of Ca-P on the surface of scaffold immersed in simulated body fluid solution indicated the suitable osteoconductivity of PCL/CS-PRF core-shell nanofibrous scaffold. Also, the value of ALP activity and calcium deposited on the surface of PCL/CS-PRF core-shell nanofibrous scaffold were 81.97 U/L and 40.33 µg/scaffold, respectively after 14 days, which confirmed the significantly higher amounts of ALP and calcium deposition on the scaffold containing PRF compared to PCL/CS scaffold. Due to higher hydrophilicity and porosity of PCL/CS-PRF core-shell nanofibrous scaffold compared to PCL/CS scaffold, a better bone cell growth on surface of PCL/CS-PRF scaffold was observed. The Alizarin red-positive area was significantly higher on PCL/CS-PRF scaffold compared to PCL/CS scaffold, indicating more calcium deposition and osteogenic differentiation of HMSCs in the presence of PRF. Our findings demonstrate that PCL/CS-PRF core-shell scaffolds can provide a strong construct with improved osteogenic for bone tissue engineering applications.

Prospective cohort study of regenerative potential of non vital immature permanent maxillary central incisors using platelet rich fibrin scaffold.

Regenerative endodontic procedures have gained momentum as a treatment modality of young immature permanent teeth. Literature reports reveal that platelet-rich fibrin (PRF) stimulates growth factors and induces regeneration. This study was undertaken to assess the regenerative potential of non-vital immature permanent maxillary central incisors using PRF with a follow-up for 2 yrs. 19 patients in the age group of 9-25 yrs with immature, non-vital permanent maxillary central incisors (n = 23) with/without signs and/or symptoms of periapical pathosis and open apex were included in this study. In the first appointment, access opening, canal disinfection and triple antibiotic paste placement were done. In the subsequent visit, PRF was prepared and placed inside the canal. Access was sealed with Mineral trioxide aggregate plug and composite. The patient was reviewed up to 24 mths. The mean difference was statistically analyzed using Friedman test followed by Dunn post hoc test and adjusted by Bonferroni correction (p < 0.05). As per AAE guidelines, the primary and secondary goals were achieved. A significant (p < 0.001) gradual increase in the root length, thickness of dentinal walls and decrease in apical diameter were observed. Within the limitations of this study, PRF placement was clinically and radiographically effective in inducing regeneration of non-vital immature permanent teeth.

Minimally invasive treatment of long bone non-unions with bone marrow concentrate, demineralized bone matrix and platelet-rich fibrin in 38 patients.

To determine the efficacy of percutaneous injection of autologous bone marrow concentrated (BMC), demineralized bone matrix (DBM), and platelet rich fibrin (PRF) in the treatment of long bone non-unions. From January 2011 to January 2018 patients with non-union of the lower limbs who were on the waiting list for open grafting with established tibial or femoral non-union and minimal deformity were eligible to participate in this study. Patients were treated with a single percutaneous injection of DBM, BMC and PRF. Our study group comprised 38 patients (26 males and 12 females; mean age 39, range 18 to 65). Non-unions were located in the femur (18 cases) and in the tibia (20 cases). Clinical and imaging follow-up ranged from 4 to 60 months (mean 20 months). Bone union occurred in 30 out of 38 patients (79%) in an average of 7 months (range 3 to 12) and all healed patients had full weight bearing after 9 months on average (range 6 to 12) from injection. In 19 cases the osteosynthesis was removed 12 months on average (range 3 to 36) from surgery. One patient developed infection at the non-union site after treatment. Percutaneous injection of DBM, BMC, and PRF is an effective treatment for long-bone non-unions. This technique allows the bone to heal with a minimally invasive approach and with a hospitalization of 2 days. Key elements of bone regeneration consist of a combination of biological and biomechanical therapeutic approach.

Platelet activation and antimicrobial activity of L-PRF: a preliminary study.

Leukocyte and platelet rich fibrin (L-PRF) is one of the platelet concentrates used to support regeneration and healing process. Many studies showed possible immunological and antibacterial properties of L-PRF. We perform an in vitro study to analyze the effect of L-PRF on platelet activation, platelet-leukocytes interactions and antimicrobial activity, important components in the healing process. Molecular biomarkers related with platelet activation and platelet-leukocyte interactions were analyzed by means of flow cytometry when L-PRF exudate was added to whole blood platelets. L-PRF membrane was used to evaluate antimicrobial activity using Enterococcus faecalis (ATCC 29212), Pseudomonas aeruginosa (ATCC 27853) and Candida albicans (ATCC 90028). Our experimental design allows to evaluate platelet activation and analyze molecular biomarkers of other immune cells and platelet-leukocyte interactions. From the results obtained we can conclude that L-PRF can be a valuable tool in healing process, efficient in activating platelets of whole blood and inhibiting microbial growth. In our opinion, the use of L-PRF exudate, in addition to L-PRF membrane, presents some advantages that have to be considered in clinical trials. Additional research on the characterization and quantification of cells and its products present in the L-PRF exudate, as well as on the temporal factor released. Also, further studies using strains isolated from clinical cases are needed.

Impact of g force and timing on the characteristics of platelet-rich fibrin matrices.

Recently, new centrifugation protocols for the preparation of platelet-rich fibrin (PRF) have been introduced in an attempt to further improve the beneficial impact of these 2nd generation platelet concentrate membranes. This in-vitro study aimed to compare the biological and physical characteristics of three types of PRF membranes using two different centrifuges with adapted relative centrifugal forces (RCF): leucocyte- and platelet-rich fibrin, advanced platelet-rich fibrin, and advanced platelet-rich fibrin+. Release of growth factors, macroscopic dimensions, cellular content and mechanical properties of the respective membranes, prepared from blood of the same individual were explored. Furthermore, the impact of timing (blood draw-centrifugation and centrifugation-membrane preparation) was assessed morphologically as well as by electron microscopy scanning. No statistically significant differences amongst the three PRF modifications could be observed, neither in their release of growth factors or the cellular content, nor in clot/membrane dimensions. The difference between both centrifuges were negligible when the same g-force was used. A lower g-force, however, reduced membrane tensile strength. Timing in the preparation process had a significant impact. Adaptation of RCF only had a minimal impact on the final characteristics of PRF membranes.

Advanced-platelet-rich fibrin extract promotes adipogenic and osteogenic differentiation of human adipose-derived stem cells in a dose-dependent manner in vitro.

Advanced platelet-rich fibrin (A-PRF) is an autogenous biological material obtained from peripheral blood. A-PRF extract (A-PRFe) contains a high concentration of various cytokines that are increasingly appreciated for their roles in improving stem cell repairing function during tissue regeneration. However, the optimal A-PRFe concentration to stimulate stem cells is unknown. This study aimed to identify the optimal concentrations of A-PRFe to promote adipogenic and osteogenic differentiation of human adipose-derived stem cells (ASCs). We produced A-PRFe from A-PRF clots by centrifuging fresh peripheral blood samples and isolated and identified ASCs using surface CD markers and multilineage differentiation potential. Enzyme-linked immunosorbent assay (ELISA) showed the concentrations of several cytokines, including b-FGF, PDGF-BB, and others, increased gradually, peaked on day 7 and then decreased. Cell proliferation assays showed A-PRFe significantly stimulated ASC proliferation, and proliferation significantly increased at higher A-PRFe doses. The degree of adipogenic and osteogenic differentiation increased at higher A-PRFe concentrations in the culture medium, as determined by oil red O and alizarin red staining. Reverse transcription polymerase chain reaction (RT-PCR) showed that expression levels of genes related to adipogenic/osteogenic differentiation (PPARγ2, C/EBPα, FABP4, Adiponectin, and ALP, OPN, OCN, RUNX2), paracrine (HIF-1α, VEGF, IGF-2) and immunoregulation (HSP70, IL-8) function were higher in groups with a higher concentration of A-PRFe than in lower concentration groups. This study demonstrates that A-PRFe is ideal for use in ASC applications in regenerative medicine because it improves biological functions, including proliferation, adipogenic/osteogenic differentiation, and paracrine function in a dose-dependent manner.

Effects of rotor angle and time after centrifugation on the biological in vitro properties of platelet rich fibrin membranes.

This study evaluated the impact of rotor angle and time of storage after centrifugation on the in vitro biological properties of platelet-rich fibrin (PRF) membranes. Blood samples (n = 9) were processed with a vertical fixed-angle (V) or a swing-out horizontal (H) centrifuge, with 20-60 min of sample storage after centrifugation. Leukocytes, platelets, and red blood cells were counted, and fibrin architecture was observed by scanning electron microscopy (SEM). The release of FGF2, PDGFbb, VEGF, IL-6, and IL-1ß was measured after incubation on culture media for 7-21 days. Cell content was equivalent in all experimental groups (p > .05). The fibrin matrix was similar for fixed-angle and horizontal centrifugation. Horizontal centrifugation induced a twofold increase in PDGF and 1.7× increase on FGF release as compared to V samples, while IL-1ß was significantly reduced (p < .05). No significant difference was observed on the release of growth factors and cytokines at different times after centrifugation (p < .05). These data suggest that both angles of centrifugation produce PRF membranes with similar structure and cellularity, but horizontal centrifugation induces a higher release of growth factors. Higher times of storage after centrifugation did not impact on cell content and the release of growth factors.

Relative Centrifugal Force (RCF; G-Force) Affects the Distribution of TGF-ß in PRF Membranes Produced Using Horizontal Centrifugation.

Solid platelet-rich fibrin (PRF) is produced with centrifugation tubes designed to accelerate clotting. Thus, activated platelets may accumulate within the fibrin-rich extracellular matrix even before centrifugation is initiated. It can thus be assumed that platelets and their growth factors such as transforming growth factor-ß (TGF-ß) are trapped within PRF independent of their relative centrifugal force (RCF), the gravitation or g-force. To test this assumption, we prepared PRF membranes with tubes where clotting is activated by a silicone-coated interior. Tubes underwent 210 g, 650 g and 1500 g for 12 min in a horizontal centrifuge. The respective PRF membranes, either in total or separated into a platelet-poor plasma and buffy coat fraction, were subjected to repeated freeze-thawing to prepare lysates. Gingival fibroblasts were exposed to the PRF lysates to provoke the expression of TGF-ß target genes. We show here that the expression of interleukin 11 (IL11) and NADPH oxidase 4 (NOX4), and Smad2/3 signaling were similarly activated by all lysates when normalized to the size of the PRF membranes. Notably, platelet-poor plasma had significantly less TGF-ß activity than the buffy coat fraction at both high-speed protocols. In contrast to our original assumption, the TGF-ß activity in PRF lysates produced using horizontal centrifugation follows a gradient with increasing concentration from the platelet-poor plasma towards the buffy coat layer.

A new insight of Platelet-Rich Fibrin clots morphology and their elemental composition.

This study analyzed the architecture of Platelet-Rich Fibrin (PRF) clots and assessed their elemental composition in order to provide new insight into this biomaterial. Five surplus PRF clots (2,700 RPM, 12 min.) donated by patients (63.6 ±â€¯12.3 years old) were prepared for use in dental clinical procedures. The internal three-dimensional morphology of the red zones and the thirds of the yellow zones of the clots were analyzed by Variable Pressure Scanning Electron Microscope (VPSEM) after sample preparation by two methods: 1. Fixation (2.5% gluataraldehyde); and 2. Fixation with subsequent partial removal of extracellular elements (8 N, HCl). Semi-quantitative elemental analysis was performed by energy-dispersive X-ray spectrometry (EDX). VPSEM analysis showed erythrocytes in both the red zone and the yellow zone, which consisted mainly of fibrin. Removal of extracellular elements enriched the morphology of both zones; the organization of the fibrin was observed to differ in the thirds of the yellow zone, with increasing density and organization to distal. The elements that compose organic substances (C-Carbon, N-Nitrogen, O-Oxygen, Na-Sodium and P-Phosphorus) and halogens (Cl-Chloride and S-Sulfur) were detected; the highest concentrations were of C, followed by O (p < 0.05), in the proximal region of the fibrin. The results of the present study suggest organization of fibrin in the PRF clot, and also reveal the distribution of the elements present in the different regions of the clot. Improved understanding of these characteristics may favor the use of this biomaterial by increasing its efficiency and functionality.

Platelet-Released Growth Factors and Platelet-Rich Fibrin Induce Expression of Factors Involved in Extracellular Matrix Organization in Human Keratinocytes

Platelet-released growth factor (PRGF) is a thrombocyte concentrate lysate which, like its clinically equivalent variations (e.g., Vivostat PRF® (platelet-rich fibrin)), is known to support the healing of chronic and hard-to-heal wounds. However, studies on the effect of PRGF on keratinocytes remain scarce. This study aims to identify genes in keratinocytes that are significantly influenced by PRGF. Therefore, we performed a whole transcriptome and gene ontology (GO) enrichment analysis of PRGF-stimulated human primary keratinocytes. This revealed an increased expression of genes involved in extracellular matrix (ECM) organization. Real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) analysis confirmed the PRGF-mediated induction of selected ECM-related factors such as transforming growth factor beta-induced protein, fibronectin 1, matrix metalloproteinase-9, transglutaminase 2, fermitin family member 1, collagen type I alpha 1 and collagen type XXII alpha 1. PRGF-induced expression of the above factors was influenced by blockade of the epidermal growth factor receptor (EGFR), a receptor playing a crucial role in wound healing. A differential induction of the investigated factors was also detected in skin explants exposed to PRGF and in experimentally generated in vivo wounds treated with Vivostat PRF®. Together, our study indicates that the induction of ECM-related factors may contribute to the beneficial wound-healing effects of PRGF-based formulations.

Evaluation of PRF Efficiency in the Treatment of Infrabony Defects.

AIM: The present study aimed to investigate the effectiveness of PRF in the treatment of infrabony defects in patients with chronic periodontitis by evaluating the clinical outcome through periodontal depth, clinical attachment level at the baseline, 6 and 9 months post operatively. MATERIAL AND METHODS: Sixty infrabony defects with probing depth ≥ 5 mm were treated. The inclusion criterion was the necessity for surgical bilateral maxillary treatment. By using split-mouth study design, each patient had one side treated with conventional flap surgery and the other side with conventional flap surgery and PRF. Clinical parameters, such as probing depth (PD) and clinical attachment lost (CAL), were recorded in both groups at baseline, 6 and 9 months post operatively. RESULTS: Positive effects for all clinical and radiographic parameters were evident in the group with PRF. Mean PD reduction demonstrated statistically significant greater results in the test group (4.00±1.07 mm) compared to the control one (4.83±0.99 mm), p = 0.003 after 9 months postoperatively. After 9 months, there were better results in the test group compared to the control group for CAL (5.60±1.61 mm, 6.20±1.58 mm), but statistically not significant. CONCLUSION: Additional use of PRF in the conventional surgical treatment of infrabony defects demonstrated better parameters than the open flap debridement alone.

Combination of an allogenic and a xenogenic bone substitute material with injectable platelet-rich fibrin - A comparative in vitro study.

The aim of the in vitro study was a comparison of an allogenic (ABSM) and a xenogenic bone substitute material (XBSM) with and without injectable platelet-rich fibrin (ABSM-i-PRF & XBSM-i-PRF) on cell characteristics of human osteoblasts (HOB). Here, ABSM and XBSM (+ i-PRF = test; - i-PRF = control) were incubated with HOB for 3, 7 and 10 days. HOB viability, migration, proliferation and differentiation (RT-PCR on alkaline phosphatase (AP), bone morphogenetic protein 2 (BMP-2) and osteonectin (OCN)) were measured and compared between groups. At day 3, an increased viability, migration and proliferation was seen for ABSM-i-PRF. For viability and proliferation (days 7 and 10) and for migration (day 10), ABSM-i-PRF/XBSM-i-PRF showed higher values compared to ABSM/XBSM with maximum values for ABSM-i-PRF and minimum values for XBSM. At days 3 and 7, the highest expression of AP was detected in ABSM-i-PRF/XBSM-i-PRF when compared to ABSM/XBSM, whereas at day 10, AP expression levels were elevated in ABSM-i-PRF/ABSM. The highest BMP-2 expression was seen in ABSM-i-PRF whereas OCN expression showed higher levels in ABSM-i-PRF/XBSM-i-PRF at days 3 and 7 with lowest expression for ABSM. Later on, elevated OC levels were detected for ABSM-i-PRF only. In conclusion, i-PRF in combination with ABSM enhances HOB activity when compared to XBSM-i-PRF or untreated BSM in vitro. Therefore, addition of i-PRF to ABSM and - to a lower extent - to XBSM may influence osteoblast activity in vivo.

Critical-sized bone defects regeneration using a bone-inspired 3D bilayer collagen membrane in combination with leukocyte and platelet-rich fibrin membrane (L-PRF): An in vivo study.

OBJECTIVES: We aim to develop a 3D-bilayer collagen (COL) membrane reinforced with nano beta-tricalcium-phosphate (nß-TCP) particles and to evaluate its bone regeneration in combination with leukocyte-platelet-rich fibrin (L-PRF) in vivo. BACKGROUND DATA: L-PRF has exhibited promising results as a cell carrier in bone regeneration in a number of clinical studies, however there are some studies that did not confirm the positive results of L-PRF application. METHODS: Mechanical & physiochemical characteristics of the COL/nß-TCP membrane (1/2 & 1/4) were tested. Proliferation and osteogenic differentiation of seeded cells on bilayer collagen/nß-TCP thick membrane was examined. Then, critical-sized calvarial defects in 8 white New Zealand rabbits were filled with either Col, Col/nß-TCP, Col/nß-TCP combined with L-PRF membrane, or left empty. New bone formation (NBF) was measured histomorphometrically 4 & 8 weeks postoperatively. RESULTS: Compressive modulus increases while porosity decreases with higher ß-TCP concentrations. Mechanical properties improve, with 89 % porosity (pore size ∼100 µm) in the bilayer-collagen/nß-TCP membrane. The bilayer design also enhances the proliferation and ALP activity. In vivo study shows no significant difference among test groups at 4 weeks, but Col/nß-TCP + L-PRF demonstrates more NBF compared to others (P < 0.05) after 8 weeks. CONCLUSION: The bilayer-collagen/nß-TCP thick membrane shows promising physiochemical in vitro results and significant NBF, as ¾ of the defect is filled with lamellar bone when combined with L-PRF membrane.

The Effects of Leukocyte-Platelet Rich Fibrin (L-PRF) on Suppression of the Expressions of the Pro-Inflammatory Cytokines, and Proliferation of Schwann Cell, and Neurotrophic Factors.

This study evaluates the use of L-PRF as an autologous scaffold in nerve regeneration, and Schwann cells (SCs) proliferation and secretion of neurotrophic factors and its anti-inflammatory effect on SC Porphyromonas Gingivalis-Lipopolysaccharide (PG-LPS)-induced inflammatory responses in vitro. SEM was done to investigate various features of L-PRF. L-PRF-extracts was used to investigate the release of growth factors and treatment of SCs line. ELISA was applied to examine the release of IGF-1. The proliferative effect of L-PRF on SCs was assessed with CCK-8 assay. The effect of L-PRF on the mRNA and protein expression of SC neurotrophic factors were analyzed by RT-qPCR and ELISA. CCK-8 assay and RT-qPCR were used to determine the required concentration and the action time of PG-LPS before the anti-inflammatory effect of L-PRF was determined by measuring the changes in IL-1ß, IL-6, and TNF-a with RT-qPCR and ELISA. There are different features in L-PRF. Fourteen days was sufficient to release adequate GF. The mRNA expressions of the pro-inflammatory cytokines were notably raised by PG-LPS in 3-hours treatment. L-PRF can increase SC proliferation, neurotrophic factors secretion, and suppress SC PG-LPS-induced inflammatory responses in vitro. L-PRF has the potential as an autologous biological additive for peripheral nerve regeneration in the event of nerve inflammation and injuries.

Platelet-Rich Fibrin Facilitates One-Stage Cartilage Repair by Promoting Chondrocytes Viability, Migration, and Matrix Synthesis.

The main aim of this study is to develop a one-stage method to combine platelet-rich fibrin (PRF) and autologous cartilage autografts for porcine articular cartilage repair. The porcine chondrocytes were treated with different concentrations of PRF-conditioned media and were evaluated for their cell viability and extracellular glycosaminoglycan (GAG) synthesis during six day cultivation. The chemotactic effects of PRF on chondrocytes on undigested cartilage autografts were revealed in explant cultures. For the in vivo part, porcine chondral defects were created at the medial femoral condyles of which were (1) left untreated, (2) implanted with PRF combined with hand-diced cartilage grafts, or (3) implanted with PRF combined with device-diced cartilage grafts. After six months, gross grades, histological, and immunohistochemical analyses were compared. The results showed that PRF promotes the viability and GAG expression of the cultured chondrocytes. Additionally, the PRF-conditioned media induce significant cellular migration and outgrowth of chondrocytes from undigested cartilage grafts. In the in vivo study, gross grading and histological scores showed significantly better outcomes in the treatment groups as compared with controls. Moreover, both treatment groups showed significantly more type II collagen staining and minimal type I collagen staining as compared with controls, indicating more hyaline-like cartilage and less fibrous tissue. In conclusion, PRF enhances the viability, differentiation, and migration of chondrocytes, thus, showing an appealing capacity for cartilage repair. The data altogether provide evidences to confirm the feasibility of a one-stage, culture-free method of combining PRF and cartilage autografts for repairing articular cartilage defects. From translational standpoints, these advantages benefit clinical applications by simplifying and potentiating the efficacy of cartilage autograft transplants.