|Year : 2021 | Volume
| Issue : 6 | Page : 1696-1699
Assessment of growth factors with three different platelet preparations, namely platelet-rich fibrin, platelet-rich plasma, and lyophilized platelet: An In vitro study
R Hariprasad1, TL Suganya2, Serin Sarah John2, P Rajesh Raj3, Feby kuriakose4, VN Anoop5
1 Full Time Specialist in Endodontics, Department of Dentistry, Employees State Insurance Corporation Hospital, Ezhukone, Kollam, Kerala, India
2 Senior Lecturer, Department of Conservative Dentistry and Endodontics, Rajas Dental College, Dr MGR Medical University, Kavalkinaru, Tamilnadu, India
3 Consultant in Oral Medicine Diagnosis and Radiology, Karunya Dental Clinics, Pulincunnu, Kavalam, Thuruthy. Kottayam District, Kerala, India
4 Assistant Professor, Department of Conservative Dentistry and Endodontics, Mar Baselios Dental College, Kothamangalam. Affiliated to KUHS (Kerala university of health sciences), Kerala, India
5 Senior lecturer, Department of Conservative Eentistry and Endodontics, Noorul Islam College of Dental Sciences, Aralumoodu, Kerala, India
|Date of Submission||30-Apr-2021|
|Date of Acceptance||09-May-2021|
|Date of Web Publication||10-Nov-2021|
Department of Dentistry, Employees State Insurance Corporation Hospital, Ezhukone, Kollam, kerala
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Assessment of growth factors with three different platelet preparations namely platelet rich fibrin, platelet rich plasma and lyophilized platelet-An invitro study Abstract Enhancing wound healing with patients own blood, particularly the platelet preparations have gained more popularity recently. These platelets excrete large amount of growth factors and cytokines that are associated with the healing process and regeneration. Even though these platelet concentrates are widely used in various forms, the mechanism behind that remains unclear. Therefore, this study aims at investigating the levels of growth factors in three different platelet preparations namely platelet rich plasma (PRP), platelet rich fibrin (PRF) and lyophilized platelets. Materials and Methods Autologous blood for preparing the platelet preparations was obtained from healthy donors aged between 25 to 35 years. The samples were then divided into three experimental groups. The preparation of PRP was done with the addition of anticoagulant and the PRF is prepared without adding it. The platelet counts in the blood were analyzed and the growth factors were quantitatively measured using ELISA reader. The statistical analysis was performed by using the Chi square test. Results In the quantitative analysis of growth factors LPL showed significant increase of the liberation of growth factors compared to PRP and PRF. Conclusions With the various recent advances in technologies for preparing these platelet concentrates this can be widely used in clinical practice more accurate in the future.
Keywords: Elisa, growth factors, lyophilized platelet, platelet preparations, surgical additives
|How to cite this article:|
Hariprasad R, Suganya T L, John SS, Raj P R, kuriakose F, Anoop V N. Assessment of growth factors with three different platelet preparations, namely platelet-rich fibrin, platelet-rich plasma, and lyophilized platelet: An In vitro study. J Pharm Bioall Sci 2021;13, Suppl S2:1696-9
|How to cite this URL:|
Hariprasad R, Suganya T L, John SS, Raj P R, kuriakose F, Anoop V N. Assessment of growth factors with three different platelet preparations, namely platelet-rich fibrin, platelet-rich plasma, and lyophilized platelet: An In vitro study. J Pharm Bioall Sci [serial online] 2021 [cited 2022 Aug 20];13, Suppl S2:1696-9. Available from: https://www.jpbsonline.org/text.asp?2021/13/6/1696/330107
| Introduction|| |
Healing is a process of becoming or making healthy again. Enhancement of this healing is a constant issue in all the medical disciplines, and recently, it was achieved by means of newer developments of biomaterials and pharmaceutical preparations. Wound healing completely depends on the process of hemostasis and it triggers a cascade of reactions which seals the vascular breach with the platelets aggregates. These platelets not only stop the hemorrhage but also plug the damaged tissue, releases growth factors and thus help in regeneration.
In spite of its excellent properties, the clinical application of this as a surgical additive is limited because of its very short lifespan of about 8–10 days. Within these days, about 15–40 × 109 platelets have to be produced daily from megakaryocytes to maintain the normal blood count (1.5–4.5 × 105/μL). So for effective clinical application, these platelets have to be concentrated in various forms.
The scientific rationale behind the use of these platelet preparations lies in the fact that these platelet concentrates acts as a reservoir of many growth factors which plays a crucial role in hard- and soft-tissue repair mechanism. The aim of this study is to assess the amount of growth factors with three different platelet preparations, namely platelet-rich plasma, platelet-rich fibrin (PRF), and lyophilized platelets.
| Materials and Methodology|| |
Preparation of platelet-rich plasma
Platelet-rich plasma was prepared from the 10 ml of venous blood, collected from the patient with the addition of an anticoagulant, namely citrate dextrose. Then, the sample was centrifuged at 1500 rpm for 10 min, from the obtained product the platelet part was separated and the leukocytes and erythrocytes were discarded. Then, the remaining sample was subjected to second centrifugation at 2500 rpm for 8 min to concentrate the platelets. Then, the samples were activated by the addition of 1000 units of thrombin and 10% CaCl2.
Preparation of platelet-rich fibrin
For the preparation of PRF, 10 ml of the patient's blood sample is collected by using a butterfly needle without adding an anticoagulant and it was transferred in a vaccutainer. After the collection of the blood, it is immediately centrifuged using a centrifuge machine at a rate of 2500 rpm for 10–15 min. After centrifugation, three layers were obtained. The top most layer is the acellular platelet poor plasma, the middle layer is the PRF clot, and the bottom layer of the test tube contains the red blood cells (RBCs). From the three layers, the PRF clot layer is removed with a sterile tweezer and was separated from the underlying RBC layer using a scissor, and then, it was transferred to a sterile dish and stored in a refrigerator.
Growth factor quantification
Each sample clotted was used to determine the levels of platelet-derived growth factor (PDGF) and transforming growth factor-beta 1 by ELISA (R and D system, Minneapolis Minn.) according to the manufacturer's instructions. At least, five determinations were carried out for each sample.
Statistical analysis was performed using the SPSS software. The Chi-square test is used. Elisa reader is used to quantify the growth factor released [Figure 1], [Figure 2], [Figure 3]. The amount of growth factor released from different platelet preparation is depicted in the descriptive statistical analysis [Table 1].
|Figure 1: Optical density value of Elisa – vascular endothelial growth factor|
Click here to view
|Figure 2: Optical density value of Elisa – platelet-derived growth factor|
Click here to view
|Table 1: Comparison of growth factor release, greatest with lipoprotein lipase|
Click here to view
| Results|| |
Optical density values of growth factors in different platelet concentrations were assessed by means of Elisa reader. Platelet is a rich storehouse of growth factors among that PDGF, vascular endothelial growth factor (VEGF) and FRF plays a critical role in healing of dental defects. Among the various platelet preparations used, PRF showed greater concentration of VEGF, PRP showed greater concentration of PDGF, but LPL showed greater concentration of VEGF, fibroblast growth factor (FGF), and PDGF [Figure 4].
|Figure 4: Comparison of growth factor release by different platelet preparations|
Click here to view
| Discussion|| |
PRP is an autologous plasma preparation containing high level of concentrated platelets along with plasma which contains large amount of growth factors which enables healing and regeneration of tissues. PRF is a matrix in which the cytokines, cells, and the growth factors are trapped within a resorbable membrane. In spite of the various advantages of these preparations, the major disadvantage is its short shelf life (5 days). Lyophilized platelets are dehydrated form of platelets which is produced by a technique called lyophilization which increases the shelf life of platelets.
The tissues of the human body, when damaged, are regenerated and healed by cell growth and its redistribution and rebuilding, and the following three factors play important roles therein. The first factor is a scaffold that forms a framework; the second factor is undifferentiated cells; and the third is a growth factor that is a signal protein secreted from the platelets, plasma, and white blood cells. Among the three factors, growth factors play the most important role. It is reported that growth factors are contained in the highest concentrations in the platelets, and that they play a pivotal role during the wound healing process.,,,,,
PDGF is reported to have chemotactic and mitogenic effects on mesenchymal cells and possesses angiogenic ability. VEGF is a potent angiogenic factor and was first described as an essential growth factor for vascular endothelial cells (EC). FGFs are a family of cytokines that possess a strong mitogenic and chemoattractant activity for EC and participate in their recruitment and proliferation.
| Conclusion|| |
Platelet concentrates are autologous products which can be prepared chair side and exhibit excellent osteoinductive antimicrobial property. Platelets are considered to be a store house of growth factors. Traditional preparation of platelet concentrates usually requires sophisticated instruments and also invasive to patients. With the invention of LPL granules, invasive methods can be avoided. This study observed that LPL granules exhibit excellent healing property due to the presence of greater amount of growth factors in comparison with PRP and PRF preparations.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Dohan Ehrenfest DM, Coelho PG, Kang BS, Sul YT, Albrektsson T. Classification of osseointegrated implant surfaces: Materials, chemistry and topography. Trends Biotechnol 2010;28:198-206.
Gresele PP, Fuster V, Vermylen J. Platelets in Thrombotic and Non-thrombotic Disorders. United Kingdom Cambridge University Press; 2002. p. 25.
Su CY, Kuo YP, Tseng YH, Su CH, Burnouf T. In vitro
release of growth factors from platelet-rich fibrin (PRF): A proposal to optimize the clinical applications of PRF. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2009;108:56-61.
Malathi K, Muthukumaraswamy A, Beri S. Periodontal regeneration of an intrabony osseous defect with combination of platelet rich fibrin and bovine derived demineralized bone matrix: A case report. IOSR-JDMS 2013;4:20-6.
Lee JW, Kwon OH, Kim TK, Cho YK, Choi KY, Chung HY, et al.
Platelet-rich plasma: Quantitative assessment of growth factor levels and comparative analysis of activated and inactivated groups. Arch Plast Surg 2013;40:530-5.
Gupta V, Bains VK, Singh GP, Mathur A, Bains R. Regenerative potential of platelet rich fibrin in dentistry: Literature review. Asian J Oral Health Allied Sci 2011;1:22-8.
Ramachandran N, Hiles MC. Use of sterilized, lyophilized platelets for multiple applications. Cytotherapy 2014;16(Supp);16:S60.
Kim ES, Park EJ, Choung PH. Platelet concentration and its effect on bone formation in calvarial defects: An experimental study in rabbits. J Prosthet Dent 2001;86:428-33.
Weibrich G, Kleis WK, Hafner G. Growth factor levels in the platelet-rich plasma produced by 2 different methods: Curasan-type PRP kit versus PCCS PRP system. Int J Oral Maxillofac Implants 2002;17:184-90.
Marx RE. Platelet-rich plasma: Evidence to support its use. J Oral Maxillofac Surg 2004;62:489-96.
Min SK, Kim HJ, Cha SR. Study on platelet rich plasma concentration according to processing method. J Korean Assoc Maxillofac Plast Reconstr Surg 2005;27:24-31.
Everts PA, Brown Mahoney C, Hoffmann JJ, Schönberger JP, Box HA, van Zundert A, et al.
Platelet-rich plasma preparation using three devices: Implications for platelet activation and platelet growth factor release. Growth Factors 2006;24:165-71.
Pietrzak WS, Eppley BL. Platelet rich plasma: Biology and new technology. J Craniofac Surg 2005;16:1043-54.
Kim SG, Zhou J, Solomon C, Zheng Y, Suzuki T, Chen M, et al.
Effects of growth factors on dental stem/progenitor cells. Dent Clin North Am 2012;56:563-75.
Duffy AM, Bouchier-Hayes DJ, Harmey JH. Vascular Endothelial Growth Factor (VEGF) and Its Role in Non-Endothelial Cells: Autocrine Signalling by VEGF. Austin (TX): Landes Bioscience; 2000-2013.
Brunner G, Nguyen H, Gabrilove J, Rifkin DB, Wilson EL. Basic fibroblast growth factor expression in human bone marrow and peripheral blood cells. Blood 1993;81:63-8.
[Figure 1], [Figure 2], [Figure 3], [Figure 4]