Antibacterial effects of platelet-rich fibrin produced by horizontal centrifugation

Abstract

Platelet-rich fibrin (PRF) has been widely used owing to its ability to stimulate tissue regeneration. To date, few studies have described the antibacterial properties of PRF. Previously, PRF prepared by horizontal centrifugation (H-PRF) was shown to contain more immune cells than leukocyte- and platelet-rich fibrin (L-PRF). This study aimed to compare the antimicrobial effects of PRFs against Staphylococcus aureus and Escherichia coli in vitro and to determine whether the antibacterial effects correlated with the number of immune cells. Blood samples were obtained from eight healthy donors to prepare L-PRF and H-PRF. The sizes and weights of L-PRF and H-PRF were first evaluated, and their antibacterial effects against S. aureus and E. coli were then tested in vitro using the inhibition ring and plate-counting test methods. Flow-cytometric analysis of the cell components of L-PRF and H-PRF was also performed. No significant differences in size or weight were observed between the L-PRF and H-PRF groups. The H-PRF group contained more leukocytes than the L-PRF group. While both PRFs had notable antimicrobial activity against S. aureus and E. coli, H-PRF demonstrated a significantly better antibacterial effect than L-PRF. Furthermore, the antimicrobial ability of the PRF solid was less efficient than that of wet PRF. In conclusion, H-PRF exhibited better antibacterial activity than L-PRF, which might have been attributed to having more immune cells.

References

[1] Pjetursson BE, Asgeirsson AG, Zwahlen M, Sailer I. Improvements in implant dentistry over the last decade: comparison of survival and complication rates in older and newer publications. Int. J. Oral. Maxillofac. Implants, 2014, 29: 308-324.

[2] Castro AB, . Regenerative potential of leucocyte- and platelet-rich fibrin. Part B: sinus floor elevation, alveolar ridge preservation and implant therapy. A systematic review. J. Clin. Periodontol., 2017, 44: 225-234.

[3] Strauss FJ, Stähli A, Gruber R. The use of platelet-rich fibrin to enhance the outcomes of implant therapy: a systematic review. Clin. Oral. Implants Res., 2018, 29: 6-19.

[4] Cortellini S, . Leucocyte- and platelet-rich fibrin block for bone augmentation procedure: a proof-of-concept study. J. Clin. Periodontol., 2018, 45: 624-634.

[5] Miron RJ, . A novel method for evaluating and quantifying cell types in platelet rich fibrin and an introduction to horizontal centrifugation. J. Biomed. Mater. Res. A, 2019, 107: 2257-2271.

[6] Dohan Ehrenfest DM, Del Corso M, Diss A, Mouhyi J, Charrier JB. Three-dimensional architecture and cell composition of a Choukroun’s platelet-rich fibrin clot and membrane. J. Periodontol., 2010, 81: 546-555.

[7] Kobayashi E, . Comparative release of growth factors from PRP, PRF, and advanced-PRF. Clin. Oral. Investig., 2016, 20: 2353-2360.

[8] Alsousou J, Ali A, Willett K, Harrison P. The role of platelet-rich plasma in tissue regeneration. Platelets, 2013, 24: 173-182.

[9] Miron RJ, . Use of platelet-rich fibrin in regenerative dentistry: a systematic review. Clin. Oral. Investig., 2017, 21: 1913-1927.

[10] Sampaio-Maia B, Caldas IM, Pereira ML, Pérez-Mongiovi D, Araujo R. The oral microbiome in health and its implication in oral and systemic diseases. Adv. Appl. Microbiol., 2016, 97: 171-210.

[11] Dewhirst FE, . The human oral microbiome. J. Bacteriol., 2010, 192: 5002-5017.

[12] Subbiahdoss G, . In vitro interactions between bacteria, osteoblast-like cells and macrophages in the pathogenesis of biomaterial-associated infections. PLoS ONE, 2011, 6

[13] Mack D, . Mechanisms of biofilm formation in Staphylococcus epidermidis and Staphylococcus aureus: functional molecules, regulatory circuits, and adaptive responses. Int. J. Med. Microbiol., 2004, 294: 203-212.

[14] Alam F, Balani K. Adhesion force of staphylococcus aureus on various biomaterial surfaces. J. Mech. Behav. Biomed. Mater., 2017, 65: 872-880.

[15] Sanz M, Chapple IL. & Working Group 4 of the VIII European Workshop on Periodontology. Clinical research on peri-implant diseases: consensus report of Working Group 4. J. Clin. Periodontol., 2012, 39: 202-206.

[16] Burnouf T, Chou ML, Wu YW, Su CY, Lee LW. Antimicrobial activity of platelet (PLT)-poor plasma, PLT-rich plasma, PLT gel, and solvent/detergent-treated PLT lysate biomaterials against wound bacteria. Transfusion, 2013, 53: 138-146.

[17] Kour P, Pudakalkatti PS, Vas AM, Das S, Padmanabhan S. Comparative evaluation of antimicrobial efficacy of platelet-rich plasma, platelet-rich fibrin, and injectable platelet-rich fibrin on the standard strains of Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans. Contemp. Clin. Dent., 2018, 9: S325-S330.

[18] Kubesch A, . A low-speed centrifugation concept leads to cell accumulation and vascularization of solid platelet-rich fibrin: an experimental study in vivo. Platelets, 2019, 30: 329-340.

[19] Miron RJ, Pinto NR, Quirynen M, Ghanaati S. Standardization of relative centrifugal forces in studies related to platelet-rich fibrin. J. Periodontol., 2019, 90: 817-820.

[20] Tsujino T, . Striking differences in platelet distribution between advanced-platelet-rich fibrin and concentrated growth factors: effects of silica-containing plastic tubes. J. Funct. Biomater., 2019, 10: 43.

[21] Miron RJ, . Platelet-rich fibrin and soft tissue wound healing: a systematic review. Tissue Eng. Part B Rev., 2017, 23: 83-99.

[22] Krijgsveld J, . Thrombocidins, microbicidal proteins from human blood platelets, are C-terminal deletion products of CXC chemokines. J. Biol. Chem., 2000, 275: 20374-20381.

[23] Tang YQ, Yeaman MR, Selsted ME. Antimicrobial peptides from human platelets. Infect. Immun., 2002, 70: 6524-6533.

[24] Yeaman MR, Bayer AS. Antimicrobial peptides from platelets. Drug Resist. Updat., 1999, 2: 116-126.

[25] Edelblute CM, Donate AL, Hargrave BY, Heller LC. Human platelet gel supernatant inactivates opportunistic wound pathogens on skin. Platelets, 2015, 26: 13-16.

[26] Bielecki T, Dohan Ehrenfest DM, Everts PA, Wiczkowski A. The role of leukocytes from L-PRP/L-PRF in wound healing and immune defense: new perspectives. Curr. Pharm. Biotechnol., 2012, 13: 1153-1162.

[27] Ghebrehiwet B, Kaplan AP, Joseph K, Peerschke EI. The complement and contact activation systems: partnership in pathogenesis beyond angioedema. Immunol. Rev., 2016, 274: 281-289.

[28] Castro A, . Antimicrobial capacity of leucocyte-and platelet rich fibrin against periodontal pathogens. Sci. Rep., 2019, 9

[29] Bielecki TM, . Antibacterial effect of autologous platelet gel enriched with growth factors and other active substances: an in vitro study. J. Bone Jt. Surg. Br., 2007, 89: 417-420.

[30] Wang Y, Wan J, Miron RJ, Zhao Y, Zhang Y. Antibacterial properties and mechanisms of gold-silver nanocages. Nanoscale, 2016, 8: 11143-11152.

[31] Castro AB, . Characterization of the leukocyte- and platelet-rich fibrin block: release of growth factors, cellular content, and structure. Int. J. Oral. Maxillofac. Implants, 2019, 34: 855-864.

[32] Cieślik-Bielecka A, Reichert P, Skowroński R, Królikowska A, Bielecki T. A new aspect of in vitro antimicrobial leukocyte- and platelet-rich plasma activity based on flow cytometry assessment. Platelets, 2019, 30: 728-736.

[33] Dudal S, . Release of LL-37 by activated human Vgamma9Vdelta2 T cells: a microbicidal weapon against Brucella suis. J. Immunol., 2006, 177: 5533-5539.

[34] Shiromizu CM, Jancic CC. γδ T Lymphocytes: an effector cell in autoimmunity and infection. Front. Immunol., 2018, 9: 2389.

[35] Conti L, . Reciprocal activating interaction between dendritic cells and pamidronate-stimulated gammadelta T cells: role of CD86 and inflammatory cytokines. J. Immunol., 2005, 174: 252-260.

[36] Garib FY, Rizopulu AP. T-regulatory cells as part of strategy of immune evasion by pathogens. Biochem. (Mosc.), 2015, 80: 957-971.

[37] Hamada S, . IL-17A produced by gammadelta T cells plays a critical role in innate immunity against listeria monocytogenes infection in the liver. J. Immunol., 2008, 181: 3456-3463.

[38] Miron RJ, . Comparison of platelet-rich fibrin (PRF) produced using 3 commercially available centrifuges at both high (~ 700 g) and low (~ 200 g) relative centrifugation forces. Clin. Oral. Investig., 2020, 24: 1171-1182.

[39] Yajamanya SR, Chatterjee A, Babu CN, Karunanithi D. Fibrin network pattern changes of platelet-rich fibrin in young versus old age group of individuals: a cell block cytology study. J. Indian Soc. Periodontol., 2016, 20: 151-156.

[40] Miron RJ, . The effect of age, gender, and time between blood draw and start of centrifugation on the size outcomes of platelet-rich fibrin (PRF) membranes. Clin. Oral. Investig., 2019, 23: 2179-2185.

[41] Li, H. & Li, B. PRP as a new approach to prevent infection: preparation and in vitro antimicrobial properties of PRP. J. Vis. Exp. 50351. https://doi.org/10.3791/50351 (2013).

[42] Shao W, . Preparation, characterization, and antibacterial activity of silver nanoparticle-decorated graphene oxide nanocomposite. ACS Appl. Mater. Interfaces, 2015, 7: 6966-6973.

Funding

National Natural Science Foundation of China (National Science Foundation of China)(81771050, 81771050, 81771050, 81771050, 81771050, 81771050, 81771050, 81771050)