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      Growth factor and pro-inflammatory cytokine contents in platelet-rich plasma (PRP), plasma rich in growth factors (PRGF), advanced platelet-rich fibrin (A-PRF), and concentrated growth factors (CGF)

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          Abstract

          Background

          The development of platelet-rich fibrin (PRF) drastically simplified the preparation procedure of platelet-concentrated biomaterials, such as platelet-rich plasma (PRP), and facilitated their clinical application. PRF’s clinical effectiveness has often been demonstrated in pre-clinical and clinical studies; however, it is still controversial whether growth factors are significantly concentrated in PRF preparations to facilitate wound healing and tissue regeneration. To address this matter, we performed a comparative study of growth factor contents in PRP and its derivatives, such as advanced PRF (A-PRF) and concentrated growth factors (CGF).

          Methods

          PRP and its derivatives were prepared from the same peripheral blood samples collected from healthy donors. A-PRF and CGF preparations were homogenized and centrifuged to produce extracts. Platelet and white blood cell counts in A-PRF and CGF preparations were determined by subtracting those counts in red blood cell fractions, supernatant acellular serum fractions, and A-PRF/CGF exudate fractions from those counts of whole blood samples. Concentrations of growth factors (TGF-β1, PDGF-BB, VEGF) and pro-inflammatory cytokines (IL-1β, IL-6) were determined using ELISA kits.

          Results

          Compared to PRP preparations, both A-PRF and CGF extracts contained compatible or higher levels of platelets and platelet-derived growth factors. In a cell proliferation assay, both A-PRF and CGF extracts significantly stimulated the proliferation of human periosteal cells without significant reduction at higher doses.

          Conclusions

          These data clearly demonstrate that both A-PRF and CGF preparations contain significant amounts of growth factors capable of stimulating periosteal cell proliferation, suggesting that A-PRF and CGF preparations function not only as a scaffolding material but also as a reservoir to deliver certain growth factors at the site of application.

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          Most cited references17

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          A comparative study of platelet-rich fibrin (PRF) and platelet-rich plasma (PRP) on the effect of proliferation and differentiation of rat osteoblasts in vitro.

          The purpose of this study was to evaluate the effect of biologic characteristics of platelet-rich plasma (PRP) and platelet-rich fibrin (PRF) on proliferation and differentiation of rat osteoblasts. Blood samples were collected from 14 healthy volunteers (7 male) with a mean age of 23.2 +/- 2.24 years. PRP and PRF were prepared with standard protocols. The exudates of PRP and PRF were collected at the time points of 1, 7, 14, 21, and 28 days. The levels of platelet-derived growth factor AB (PDGF-AB) and transforming growth factor beta1 (TGF-beta1) were quantified in PRP and PRF. Then the exudates of PRP and PRF were used to culture rat calvaria osteoblasts. The biologic characteristics of osteoblasts were analyzed in vitro for 14 days. PRP released the highest amounts of TGF-beta1 and PDGF-AB at the first day, followed by significantly decreased release at later time points. PRF released the highest amount of TGF-beta1 at day 14 and the highest amount of PDGF-AB at day 7. Exudates of PRP collected at day 1 and exudates of PRF collected at day 14 expressed maximum alkaline phosphatase (ALP) activity, though no significance was shown. Cells treated with exudates of PRF collected at day 14 reached peak mineralization significantly more than both negative control and positive control groups. PRF is superior to PRP, from the aspects of expression of ALP and induction of mineralization. PRF released autologous growth factors gradually and expressed stronger and more durable effect on proliferation and differentiation of rat osteoblasts than PRP in vitro.
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            Platelet-rich plasma contains high levels of platelet-derived growth factor and transforming growth factor-beta and modulates the proliferation of periodontally related cells in vitro.

            Platelet-rich plasma (PRP) is a fraction of plasma, in which platelet-derived growth factor (PDGF) and transforming growth factor-beta (TGF-beta) are thought to be concentrated. It is plausible that topically-applied PRP up-regulates cellular activity and subsequently promotes periodontal regeneration in vivo. However, the concentrations of these growth factors in PRP have not been specifically determined and the biological effects of PRP at the cellular and molecular levels have not been determined. PRP obtained from 20 healthy subjects was prepared from plasma by centrifugation. These PRP preparations were immediately subjected to an evaluation for PDGF-AB and TGF-beta1 using enzyme-linked immunosorbent assay (ELISA) kits. The biological effects of the PRP preparations were evaluated on osteoblastic, epithelial, fibroblastic, and periodontal ligament cells. Cellular mitogenic activity was evaluated by counting cell numbers or evaluating 5-bromodeoxyuridine (BrdU) incorporation. Expression of alkaline phosphatase (ALP) was immunocytochemically evaluated. In the PRP preparations, platelets were concentrated up to 70.9 x 10(4) cells/microl (283.4% of the unconcentrated plasma). The levels of PDGF-AB and TGF-beta1 were also concentrated up to 182.0 ng/ml (440.6%) and 140.9 ng/ml (346.6%), respectively. Scatter plots revealed significant correlations between platelet counts and levels of these growth factors. PRP stimulated osteoblastic DNA synthesis and cell division (138% of control), with simultaneous down-regulation of ALP, but suppressed epithelial cell division (80% of control). PRP also stimulated DNA synthesis in gingival fibroblasts and periodontal ligament cells. These data demonstrated that both PDGF-AB and TGF-beta1 were highly concentrated in the PRP preparations. It is suggested PRP modulates cell proliferation in a cell type-specific manner similar to what has been observed with TGF-beta1. Since synchronized behavior of related cell types is thought to be required for successful periodontal regeneration, it is further suggested these cell type-specific actions may be beneficial for periodontal regenerative therapy.
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              Platelet-rich plasma and platelet-rich fibrin in human cell culture.

              The clinical use of platelet-rich plasma (PRP) for preprosthetic surgery has been a matter of controversy until now. Only recently, a new blood preparation has been developed which results in platelet-rich fibrin (PRF). The objective of the present investigation was to examine the growth factor release from PRP and PRF in vitro. Whole blood samples from healthy participants (n = 10) were drawn to generate PRP and PRF. Human osteoblasts (O), human fibroblasts (F), and human osteoblast-derived osteosarcoma cells (Saos-2) were used for the cell culture. Cells of each cell line were cultivated, and PRP- or PRF-preparations added for ten days. The drawn medium was pooled and the quantities of growth factors (platelet-derived growth factor isomers AB and BB, insulin-like growth factor I, and transforming growth factor (TGF) isomers beta1 and beta2) analyzed by enzyme-linked immunosorbent assay. In osteoblast and Saos-2 cultures, cytokine concentrations were significantly higher for PRP than for PRF (P < .05). In fibroblast cultures, results were the same with the exception of TGF-beta2 (P < .05). This study demonstrates that PRP application in cell cultures leads to higher levels of growth factors than PRF application.
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                Author and article information

                Contributors
                kawase@dent.niigata-u.ac.jp
                Journal
                Int J Implant Dent
                Int J Implant Dent
                International Journal of Implant Dentistry
                Springer Berlin Heidelberg (Berlin/Heidelberg )
                2198-4034
                22 August 2016
                22 August 2016
                December 2016
                : 2
                : 1
                : 19
                Affiliations
                [1 ]Tokyo Plastic Dental Society, Kita-ku, Tokyo, Japan
                [2 ]Bioscience Medical Research Center, Niigata University Medical and Dental Hospital, Niigata, Japan
                [3 ]Department of Dentistry, National Yang-Ming University, Taipei, Taiwan
                [4 ]Division of Oral Bioengineering, Institute of Medicine and Dentistry, Niigata University, Niigata, Japan
                Author information
                http://orcid.org/0000-0003-4160-396X
                Article
                52
                10.1186/s40729-016-0052-4
                5005757
                50acc701-f959-4c66-a4b4-b9c4be4cf178
                © The Author(s). 2016

                Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License ( http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.

                History
                : 21 June 2016
                : 18 August 2016
                Categories
                Research
                Custom metadata
                © The Author(s) 2016

                growth factor,platelet-rich plasma,platelet-rich fibrin,plasma rich in growth factors,concentrated growth factors

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