scholarly journals Study of a Two-Step Centrifugation Protocol for Concentrating Cells and Growth Factors in Bovine Platelet-Rich Plasma

2017 ◽  
Vol 2017 ◽  
pp. 1-8
Author(s):  
Claudia M. Gutiérrez ◽  
Catalina López ◽  
Carlos E. Giraldo ◽  
Jorge U. Carmona

There is a lack of information about the methods used for bovine platelet-rich plasma (PRP)/platelet-rich gel (PRG) procurement, including information on platelet (PLT), white blood cell (WBC) in PRP, and growth factor release from PRG supernatants. The aims of this study were to compare and to correlate the PLT, WBC, transforming growth factor beta-1 (TGF-β1), and platelet-derived growth factor BB (PDGF-BB) concentrations in bovine whole blood, plasma, and four PRP layers and their respective PRG supernatants: A and B (obtained by a single centrifugation tube method at 720g/5 min) and C and D (obtained by a double centrifugation tube method, by using two centrifugation episodes at 720g/5 min). PLT and WBC counts were significantly higher in PRP-C, followed by whole blood, PRP-A, PRP-B, and PRP-D. TGF-β1 concentrations were significantly higher in PRG-B supernatants and its correspondent PRP-B lysate when compared to the other PRG supernatants and plasma. Supernatants from PRG-A, PRG-B, and PRG-D had equivalent TGF-β1 concentrations. PDGF-BB concentrations were not statistically different between the hemoderivatives. Significant Pearson correlations were noted between PLT counts and WBC counts (0.8) and between PLT counts and PLT distribution width (0.6). Further studies should be performed to assess the potential clinical applications of these PRPs.

2016 ◽  
Vol 8 (3) ◽  
Author(s):  
Lucia Nirmalasari ◽  
Maximillian Ch. Oley ◽  
Eko Prasetyo ◽  
Mendy Hatibie ◽  
Lily L. Loho

Abstract: Recently, platelet rich plasma has been popular and its use has begin on human in developed countries. Platelet rich plasma is defined as autologus blood with concentration of platelets three to five times above baseline level, which contains at least seven growth factors like Platelet Derived Growth Factor (PDGF), Platelet Derived Angiogenesis Factor (PDAF), Platelet Derived Endothelial Growth Factor (PDEGF), Transforming Growth Factor Beta (TGF- β), Insulin like Growth Factor (IGF), Fibroblast Growth Factor (FGF), and Vascular Endothelial Growth Factor (VEGF). The golden standard for reconstruction of cranial bone defects demonstrates osteoconduction scaffold, osteoinduction like growth factors, and osteogenesis. Alloplastic biomaterials have revolutionalized craniofacial reconstruction. Carbonated hydroxyapatite (CHA) has been studied for years as implant material due to its similarity with the mineral component of bone. In this study we investigated and compare the effects of PRP and CHA on bone regeneration in rat cranial defects. This was an experimental study with a true experimental design on white male rats (Rattus norvegicus). Cranial deffects of 3 mm diameter were created in rat cranium and grafted with CHA and PRP combination, CHA alone, and control. The relationships among them were analyzed by using Mann Whitney and SPSS Statistics Program Package Version 22.0. The results showed that the experimental group of 2 weeks had no different between inflammatory reaction (P = 0.119), woven bone (P = 0.094) and lamellar bone (P = 0.130). At 4 weeks,a combination of PRP and CHA showed a superior growth of lamellar bone compared to CHA (P = 0.009). Conclusion: A combination of PRP and CHA in bone regeneration showed a histological tendency toward increased bone formation. However, future investigations should be conducted in different period times.Keywords: platelet rich plasma, carbonated hydroxyapatite, cranial defectAbstrak: Plasma kaya trombosit makin banyak digunakan dalam dunia kedokteran. Di negara maju pengunaannya sudah mulai diteliti pada manusia. Plasma kaya trombosit adalah fraksi plasma darah dengan konsentrasi platelet 3-5 kali diatas nilai normal yang mengandung sekurang-kurangnya 7 faktor pertumbuhan, diantaranya Platelet Derived Growth Factor (PDGF), Platelet Derived Angiogenesis Factor (PDAF), Platelet Derived Endothelial Growth Factor (PDEGF), Transforming Growth Factor Beta (TGF- β), Insulin like Growth Factor (IGF), Fibroblast Growth Factor (FGF), dan Vascular Endothelial Growth Factor (VEGF) yang dapat meningkatkan proses osteogenesis. Karbonat hidroksiapatit adalah material pengganti tulang yang dapat mempercepat regenerasi jaringan tulang serta memiliki kandungan kalsium,fosfat dan karbonat yang mirip dengan tulang manusia. Tulang yang tumbuh pada awal berupa tulang muda yang memiliki serat kolagen yang tidak teratur dan banyak osteosit disebut tulang imatur. Tulang imatur kemudian akan diganti oleh tulang matur yang memiliki serabut kolagen yang teratur. Jenis penelitian ini ialah eksperimental pada 36 hewan coba tikus putih wistar (Rattus norvegicus). Defek kalvaria pada tikus dengan diameter 3 mm diisi sesuai perlakuan: plasma kaya trombosit dengan karbonat hidroksiapatit, karbonat apatit tunggal, dan kontrol. Plasma kaya trombosit dibuat dari autologus darah tikus yang diberi perlakuan plasma kaya trombosit serta karbonat hidroksiapatit dan karbonat apatit tunggal. Data dianalisis dengan uji Mann Whitney dan diolah dengan SPSS. Hasil penelitian memperlihatkan pada minggu ke-2, tidak terdapat perbedaan bermakna reaksi inflamasi (P = 0,119), tulang imatur (P = 0,094), dan tulang matur (P = 0,130) diantara ketiga perlakuan. Pada minggu ke-4, tulang matur yang terbentuk lebih banyak pada perlakuan plasma kaya trombosit dan karbonat hidroksiapatit (P = 0,009). Simpulan: Pemberian plasma kaya trombosit dan karbonat hidroksiapatit dapat meningkatkan proses penutupan defek tulang kepala hewan percobaan tikus.Kata kunci : plasma kaya trombosit, karbonat hidroksiapatit, defek tulang kepala.


2016 ◽  
Vol 45 (4) ◽  
pp. 954-960 ◽  
Author(s):  
Matthias Kieb ◽  
Frank Sander ◽  
Cornelia Prinz ◽  
Stefanie Adam ◽  
Anett Mau-Möller ◽  
...  

Background: Platelet-rich plasma (PRP) is widely used in sports medicine. Available PRP preparations differ in white blood cell, platelet, and growth factor concentrations, making standardized research and clinical application challenging. Purpose: To characterize a newly standardized procedure for pooled PRP that provides defined growth factor concentrations. Study Design: Controlled laboratory study. Methods: A standardized growth factor preparation (lyophilized PRP powder) was prepared using 12 pooled platelet concentrates (PCs) derived from different donors via apheresis. Blood samples and commercially available PRP (SmartPrep-2) served as controls (n = 5). Baseline blood counts were analyzed. Additionally, single PCs (n = 5) were produced by standard platelet apheresis. The concentrations of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), platelet-derived growth factor AB (PDGF-AB), transforming growth factor β1 (TGF-β1), insulin-like growth factor 1 (IGF-1), interleukin (IL)–1α, IL-1β, and IL-1 receptor agonist (IL-1RA) were analyzed by enzyme-linked immunosorbent assay, and statistical analyses were performed using descriptive statistics, mean differences, 95% CIs, and P values (analysis of variance). Results: All growth factor preparation methods showed elevated concentrations of the growth factors VEGF, bFGF, PDGF-AB, and TGF-β1 compared with those of whole blood. Large interindividual differences were found in VEGF and bFGF concentrations. Respective values (mean ± SD in pg/mL) for whole blood, SmartPrep-2, PC, and PRP powder were as follows: VEGF (574 ± 147, 528 ± 233, 1087 ± 535, and 1722), bFGF (198 ± 164, 410 ± 259, 151 ± 99, and 542), PDGF-AB (2394 ± 451, 17,846 ± 3087, 18,461 ± 4455, and 23,023), and TGF-β1 (14,356 ± 4527, 77,533 ± 13,918, 68,582 ± 7388, and 87,495). IGF-1 was found in SmartPrep-2 (1539 ± 348 pg/mL). For PC (2266 ± 485 pg/mL), IGF-1 was measured at the same levels of whole blood (2317 ± 711 pg/mL) but was not detectable in PRP powder. IL-1α was detectable in whole blood (111 ± 35 pg/mL) and SmartPrep-2 (119 ± 44 pg/mL). Conclusion: Problems with PRP such as absent standardization, lack of consistency among studies, and black box dosage could be solved by using characterized PRP powder made by pooling and lyophilizing multiple PCs. The new PRP powder opens up new possibilities for PRP research as well as for the treatment of patients. Clinical Relevance: The preparation of pooled PRP by means of lyophilization may allow physicians to apply a defined amount of growth factors by using a defined amount of PRP powder. Moreover, PRP powder as a dry substance with no need for centrifugation could become ubiquitously available, thus saving time and staff resources in clinical practice. However, before transferring the results of this basic science study to clinical application, regulatory issues have to be cleared.


2019 ◽  
Vol 47 (5) ◽  
pp. 1223-1229 ◽  
Author(s):  
Prathap Jayaram ◽  
Peter Yeh ◽  
Shiv J. Patel ◽  
Racel Cela ◽  
Theodore B. Shybut ◽  
...  

Background: The benefits of platelet-rich plasma (PRP) are believed to be in part dependent on growth factor release after platelet activation. Platelet activation is complex and involves multiple mechanisms. One important mechanism is driven by cyclooxygenase 1 (COX-1)–mediated conversion of arachidonic acid (AA) to precursor prostaglandins that then mediate proinflammatory responses that trigger growth factor release. Acetylsalicylic acid (ASA; also known as aspirin) is known to irreversibly inhibit COX-1, thereby blocking AA-mediated signaling; however, it is unclear whether ASA use alters growth factor release from freshly isolated PRP. Purpose: To assess the effects of low-dose ASA use on activation of growth factor release from freshly isolated human PRP via AA and thrombin (TBN). Study Design: Controlled laboratory study. Methods: Twelve healthy men underwent blood collection and leukocyte-rich PRP (LR-PRP) preparation through a double-spin protocol to obtain baseline whole blood and PRP counts the same day. PRP was aliquoted into 3 groups: nonactivated, AA activated, and TBN activated. Immediately after activation, the concentrations of transforming growth factor β1 (TGF-β1), vascular endothelial growth factor (VEGF), and platelet-derived growth factor AB (PDGF-AB) were measured using enzyme-linked immunosorbent assays (ELISAs). The same 12 participants were then placed on an 81-mg daily dose of oral ASA for 14 days. Repeat characterization of whole blood and PRP analyses was done on day 14, followed by repeat ELISAs of growth factors under the same nonactivated and activated settings as previously stated. Results: Fourteen days of daily ASA had no effect on the number of platelets and leukocytes measured in whole blood and LR-PRP. Compared with nonactivated LR-PRP, AA- and TBN-mediated activation led to significant release of VEGF and PDGF-AB. In contrast, release of TGF-β1 from LR-PRP was observed only with activation by AA, not with TBN. Consistent with its inhibitory role in AA signaling, ASA significantly inhibited AA-mediated release of all 3 growth factors measured in this study. Although ASA had no effect on TBN-mediated release of VEGF and TGF-β1 from LR-PRP, ASA did partially block TBN-mediated release of PDGF-AB, although the mechanism remains unclear. Conclusion: Daily use of low-dose ASA reduces VEGF, PDGF-AB, and TGF-β1 expression in freshly isolated human LR-PRP when activated with AA. Clinical Relevance: Reduction in growth factor release attributed to daily use of low-dose ASA or other COX inhibitors can be mitigated when PRP samples are activated with TBN. Clinical studies are needed to determine whether activation before PRP injection is needed in all applications where ASA is in use and to what extent ASA may inhibit growth factor release in vivo at the site of injury.


2020 ◽  
Vol 9 (4) ◽  
pp. 1099
Author(s):  
Giorgio Serafini ◽  
Mariangela Lopreiato ◽  
Marco Lollobrigida ◽  
Luca Lamazza ◽  
Giulia Mazzucchi ◽  
...  

Liquid fibrinogen is an injectable platelet concentrate rich in platelets, leukocytes, and fibrinogen obtained by blood centrifugation. The aim of this study was to analyze the release of different growth factors in the liquid fibrinogen at different times and to assess possible correlations between growth factors and cell counts. The concentration of transforming growth factor beta 1 (TGF-β1), platelet-derived growth factor-AB (PDGF-AB), platelet-derived growth factor-BB (PDGF-BB), bone morphogenetic protein 2 (BMP-2), fibroblast growth factor 2 (FGF-2) and vascular endothelial growth factor (VEGF) released by liquid fibrinogen were examined with ELISA at three time points (T0, time of collection; T7, 7 days; T14, 14 days). The cellular content of the liquid fibrinogen and whole blood was also calculated for each volunteer. A mean accumulation of platelets of almost 1.5-fold in liquid fibrinogen compared to whole blood samples was found. An increase of TGF-β1, PDGF-AB, FGF-2, and VEGF levels was detected at T7. At T14, the level of TGF-β1 returned to T0 level; PDGF-AB amount remained high; the levels of FGF-2 and VEGF decreased with respect to T7, but remained higher than the T0 levels; PDGF-BB was high at all time points; BMP-2 level was low and remained constant at all time points. TGF-β1, PDGF-AB, and PDGF-BB showed a correlation with platelet amount, whereas BMP-2, FGF-2, and VEGF showed a mild correlation with platelet amount. Due to the high concentration of platelets, liquid fibrinogen does contain important growth factors for the regeneration of both soft and hard tissue. The centrifugation protocol tested in this study provides a valid solution to stimulate wound healing in oral and periodontal surgery.


2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
Catalina López ◽  
María E. Álvarez ◽  
Jorge U. Carmona

The aims were (1) to evaluate the bacteriostatic effect of platelet-rich plasma (PRP), platelet-rich gel (PRG), leukocyte-poor plasma (LPP), leukocyte-poor gel (LPG), plasma, and heat-inactivated plasma (IP) on both methicillin-sensitive Staphylococcus aureus (MSSA) and methicillin-resistant Staphylococcus aureus (MRSA) over a period of 24 h; (2) to determine and to compare the concentrations and degradation over time of platelet factor 4 (PF-4), transforming growth factor beta 1 (TGF-β1), and platelet-derived growth factor isoform BB (PDGF-BB); and (3) to identify any correlations between MSSA and MRSA growth and either the cellular, PF-4, TGF-β1, or PDGF-BB concentrations in the blood components. PRP and its byproducts from 18 horses were obtained by the tube method. All blood components were cultured with either MSSA or MRSA. Bacterial growth, PF-4, TGF-β1, and PDGF-BB were determined at 6 h and 24 h. At six hours, bacterial growth was significantly inhibited by all blood components, with the exception of IP. MSSA was more sensitive to the treatments than MRSA. At 24 hours, bacterial growth was significantly higher in IP. MRSA bacterial growth was significantly higher in PRP, LPP, and plasma when compared to MSSA. Growth factor concentrations were not significantly affected by bacteria.


2020 ◽  
Vol 2 (1) ◽  
pp. e31-e47
Author(s):  
Chris Cherian ◽  
Gerard Malanga ◽  
Ken Mautner

Platelet-rich plasma (PRP) is an orthobiologic treatment that has gained popularity as a potential alternative treatment for various musculoskeletal conditions. The physiologic role of platelets in the healing cascade provides clarity regarding its potential as it releases various growth factors such as platelet-derived growth factor (PDGF), transforming growth factor beta-1 (TGF-β1), and vascular endothelial growth factor (VEGF). However, there are various characteristics of PRP treatments including platelet count, presence or absence of leukocytes and red blood cells, as well as the use of an activating agent that introduces heterogeneity among preparations. This aim of this article is to provide clarity, where available, regarding the optimal characteristics for PRP treatments regarding tendon and ligament injuries as well as articular and muscular pathology.


2018 ◽  
Vol 2018 ◽  
pp. 1-10 ◽  
Author(s):  
Neveen Salem ◽  
Nawal Helmi ◽  
Naglaa Assaf

Platelet-rich plasma (PRP) has grown as an attractive biologic instrument in regenerative medicine for its powerful healing properties. It is considered as a source of growth factors that may induce tissue repairing and improve fibrosis. This product has proven its efficacy in multiple studies, but its effect on cisplatin-induced nephrotoxicity has not yet been elucidated. The present investigation was performed to estimate the protective impact of platelet-rich plasma against cisplatin- (CP-) evoked nephrotoxicity in male rats. Nephrotoxicity was induced in male Wistar rats by right uninephrectomy followed by CP administration. Uninephrectomized rats were assigned into four groups: (1) control group, (2) PRP group, (3) CP group, and (4) CP + PRP group. PRP was administered by subcapsular renal injection. Renal function, inflammatory cytokines, and growth factor level as well as histopathological investigation were carried out. Treatment with PRP attenuated the severity of CP-induced nephrotoxicity as evidenced by suppressed creatinine, blood urea nitrogen (BUN), and N-acetyl glucosaminidase (NAG) levels. Moreover, PRP depressed intercellular adhesion molecule-1 (ICAM-1), kidney injury molecule-1 (KIM-1), caspase-3, and transforming growth factor-beta 1 (TGF-β1) levels, while enhanced the epidermal growth factor (EGF) level. These biochemical results were reinforced by the histopathological investigation, which revealed restoration of normal renal tissue architectures. These findings highlight evidence for the possible protective effects of PRP in a rat model of CP-induced nephrotoxicity, suggesting a new avenue for using PRP to improve the therapeutic index of cisplatin.


2020 ◽  
Vol 2020 ◽  
pp. 1-9
Author(s):  
Rayra Pereira Santiago ◽  
Camylla Vilas Boas Figueiredo ◽  
Luciana Magalhães Fiuza ◽  
Sétondji Cocou Modeste Alexandre Yahouédéhou ◽  
Rodrigo Mota Oliveira ◽  
...  

Individuals with sickle cell anemia (SCA) present chronic anemia, hemolysis, an exacerbated inflammatory response, and heterogeneous clinical complications, which may be modulated by the transforming growth factor beta (TGF-β) pathway. Thus, we aimed to investigate polymorphisms (rs1805110 and rs7526590) of the transforming growth factor beta receptor III gene (TGFBR3) with regard to laboratory biomarkers and clinical manifestations in individuals with SCA. Hematological, biochemical, immunological, and genetic analyses were carried out, as well as serum endothelin-1 measurements. The minor allele (A) of the TGFBR3 rs1805110 polymorphism was associated with increased hemoglobin, hematocrit, reticulocyte counts, total cholesterol, low-density lipoprotein, uric acid, and endothelin levels, as well as decreased platelet distribution width (PDW) and the occurrence of bone alterations. The minor allele (T) of TGFBR3 rs7526590 was associated with increased red cell distribution width, PDW, alkaline phosphatase, aspartate aminotransferase, total and indirect bilirubin, and lactate dehydrogenase levels, as well as lower ferritin levels and the occurrence of leg ulcers. Our data suggest that the minor allele (A) of TGFBR3 rs1805110 is associated with inflammation and bone alterations, while the minor allele (T) of TGFBR3 rs7526590 is related to hemolysis and the occurrence of leg ulcers.


2020 ◽  
Vol 8 (2) ◽  
pp. 67-77
Author(s):  
V. L. Medvedev ◽  
M. I. Kogan ◽  
I. V. Mihailov ◽  
S. N. Lepetunov

Platelet-rich autologous plasma (PRP) is often used in various branches of medicine. The scope of PRP therapy has expanded from stimulating bone regeneration, healing wounds and ulcers, and musculoskeletal injuries to improving the ability to engrave various types of grafts. Due to the natural properties of platelet-rich plasma, its introduction into the human body is one of the most promising procedures for tissue restoration. After the destruction of platelets, PRP contains α-granules, from which many factors are released after activation, such as transforming growth factor-beta (TGF-β), vascular endothelial growth factor (VGFF) and epidermal growth factor (EGF). The current state of the problem of using APOT has a huge perspective on the development of the methodology, which is due to many aspects that make this procedure simple. PRP can improve the course of many urological diseases, such as erectile dysfunction, Peyronie’s disease, urethral stricture, vesicovaginal fistulas, interstitial cystitis, and stress urinary incontinence. There are many protocols for preparing PRP, each of which has its standardized parameters and stated results. The article presents a review of the literature on the use of platelet-rich plasma in urology, focuses on the definition of PRP, various methods of preparation and activation, as well as the concentration of growth factors.


2020 ◽  
Vol 8 (4) ◽  
pp. 93
Author(s):  
Inmaculada Peris ◽  
Blanca Esquembre ◽  
Jose Ignacio Redondo ◽  
Vicente Jose Sifre ◽  
Carme Soler ◽  
...  

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