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Bone & Joint Research
Vol. 5, Issue 12 | Pages 602 - 609
1 Dec 2016
Muto T Kokubu T Mifune Y Inui A Sakata R Harada Y Takase F Kurosaka M

Objectives. Triamcinolone acetonide (TA) is widely used for the treatment of rotator cuff injury because of its anti-inflammatory properties. However, TA can also produce deleterious effects such as tendon degeneration or rupture. These harmful effects could be prevented by the addition of platelet-rich plasma (PRP), however, the anti-inflammatory and anti-degenerative effects of the combined use of TA and PRP have not yet been made clear. The objective of this study was to determine how the combination of TA and PRP might influence the inflammation and degeneration of the rotator cuff by examining rotator cuff-derived cells induced by interleukin (IL)-1ß. Methods. Rotator cuff-derived cells were seeded under inflammatory stimulation conditions (with serum-free medium with 1 ng/ml IL-1ß for three hours), and then cultured in different media: serum-free (control group), serum-free + TA (0.1mg/ml) (TA group), serum-free + 10% PRP (PRP group), and serum-free + TA (0.1mg/ml) + 10% PRP (TA+PRP group). Cell morphology, cell viability, and expression of inflammatory and degenerative mediators were assessed. Results. Exposure to TA significantly decreased cell viability and changed the cell morphology; these effects were prevented by the simultaneous administration of PRP. Compared with the control group, expression levels of inflammatory genes and reactive oxygen species production were reduced in the TA, PRP, and TA+PRP groups. PRP significantly decreased the expression levels of degenerative marker genes. Conclusions. The combination of TA plus PRP exerts anti-inflammatory and anti-degenerative effects on rotator cuff-derived cells stimulated by IL-1ß. This combination has the potential to relieve the symptoms of rotator cuff injury. Cite this article: T. Muto, T. Kokubu, Y. Mifune, A. Inui, R. Sakata, Y. Harada, F. Takase, M. Kurosaka. Effects of platelet-rich plasma and triamcinolone acetonide on interleukin-1ß-stimulated human rotator cuff-derived cells. Bone Joint Res 2016;5:602–609. DOI: 10.1302/2046-3758.512.2000582


The Journal of Bone & Joint Surgery British Volume
Vol. 90-B, Issue 7 | Pages 966 - 972
1 Jul 2008
Kawasumi M Kitoh H Siwicka KA Ishiguro N

The aim of our study was to investigate the effect of platelet-rich plasma on the proliferation and differentiation of rat bone-marrow cells and to determine an optimal platelet concentration in plasma for osseous tissue engineering. Rat bone-marrow cells embedded in different concentrations of platelet-rich plasma gel were cultured for six days. Their potential for proliferation and osteogenic differentiation was analysed. Using a rat limb-lengthening model, the cultured rat bone-marrow cells with platelet-rich plasma of variable concentrations were transplanted into the distraction gap and the quality of the regenerate bone was evaluated radiologically. Cellular proliferation was enhanced in all the platelet-rich plasma groups in a dose-dependent manner. Although no significant differences in the production and mRNA expression of alkaline phosphatase were detected among these groups, mature bone regenerates were more prevalent in the group with the highest concentration of platelets. Our results indicate that a high platelet concentration in the platelet-rich plasma in combination with osteoblastic cells could accelerate the formation of new bone during limb-lengthening procedures


Orthopaedic Proceedings
Vol. 105-B, Issue SUPP_9 | Pages 30 - 30
17 Apr 2023
Muthu S
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Platelet-rich plasma (PRP) has been demonstrated to benefit a variety of disciplines. But there exists heterogeneity in results obtained due to lack of standardization of the preparation protocols employed in them. We aim to identify and standardize a preparation protocol for PRP with maximum recovery of platelets to obtain reproducible results across studies. Blood samples were collected from 20 healthy volunteers. The double spin protocol of PRP preparation was analyzed for variables such as centrifugal acceleration, time, and volume of blood processed and final product utilized. The final PRP prepared was investigated for platelet recovery, concentration, integrity, and viability. We noted maximum platelet recovery (86-99%) with a mean concentration factor of 6-times baseline, with double centrifugation protocol at 100xg and 1600xg for 20 minutes each. We also noted that 10 ml of blood in a 15 ml tube was the ideal volume of blood to be processed to maximize platelet recovery. We demonstrated that the lower 1/3rd is the ideal volume to be utilized for clinical application. We did not note a loss of integrity or viability of the platelets in the final product from the above-said protocol. Preparation of PRP by the double spin protocol of 10 ml of blood at 100xg and 1600xg for 20 minutes each in a 15ml tube and using the lower 1/3rd of the final product demonstrated consistent high platelet recovery (86-99%) and concentration (6x) without disturbing the platelet integrity or viability


Bone & Joint Research
Vol. 6, Issue 4 | Pages 231 - 244
1 Apr 2017
Zhang J Yuan T Zheng N Zhou Y Hogan MV Wang JH

Objectives. After an injury, the biological reattachment of tendon to bone is a challenge because healing takes place between a soft (tendon) and a hard (bone) tissue. Even after healing, the transition zone in the enthesis is not completely regenerated, making it susceptible to re-injury. In this study, we aimed to regenerate Achilles tendon entheses (ATEs) in wounded rats using a combination of kartogenin (KGN) and platelet-rich plasma (PRP). Methods. Wounds created in rat ATEs were given three different treatments: kartogenin platelet-rich plasma (KGN-PRP); PRP; or saline (control), followed by histological and immunochemical analyses, and mechanical testing of the rat ATEs after three months of healing. Results. Histological analysis showed well organised arrangement of collagen fibres and proteoglycan formation in the wounded ATEs in the KGN-PRP group. Furthermore, immunohistochemical analysis revealed fibrocartilage formation in the KGN-PRP-treated ATEs, evidenced by the presence of both collagen I and II in the healed ATE. Larger positively stained collagen III areas were found in both PRP and saline groups than those in the KGN-PRP group. Chondrocyte-related genes, SOX9 and collagen II, and tenocyte-related genes, collagen I and scleraxis (SCX), were also upregulated by KGN-PRP. Moreover, mechanical testing results showed higher ultimate tensile strength in the KGN-PRP group than in the saline control group. In contrast, PRP treatment appeared to have healed the injured ATE but induced no apparent formation of fibrocartilage. The saline-treated group showed poor healing without fibrocartilage tissue formation in the ATEs. Conclusions. Our results show that injection of KGN-PRP induces fibrocartilage formation in the wounded rat ATEs. Hence, KGN-PRP may be a clinically relevant, biological approach to regenerate injured enthesis effectively. Cite this article: J. Zhang, T. Yuan, N. Zheng, Y. Zhou, M. V. Hogan, J. H-C. Wang. The combined use of kartogenin and platelet-rich plasma promotes fibrocartilage formation in the wounded rat Achilles tendon entheses. Bone Joint Res 2017;6:231–244. DOI: 10.1302/2046-3758.64.BJR-2017-0268.R1


Bone & Joint Research
Vol. 5, Issue 2 | Pages 37 - 45
1 Feb 2016
Roh YH Kim W Park KU Oh JH

Objectives. This study was conducted to evaluate the cytokine-release kinetics of platelet-rich plasma (PRP) according to different activation protocols. Methods. Two manual preparation procedures (single-spin (SS) at 900 g for five minutes; double-spin (DS) at 900 g for five minutes and then 1500 g for 15 minutes) were performed for each of 14 healthy subjects. Both preparations were tested for platelet activation by one of three activation protocols: no activation, activation with calcium (Ca) only, or calcium with a low dose (50 IU per 1 ml PRP) of thrombin. Each preparation was divided into four aliquots and incubated for one hour, 24 hours, 72 hours, and seven days. The cytokine-release kinetics were evaluated by assessing PDGF, TGF, VEGF, FGF, IL-1, and MMP-9 concentrations with bead-based sandwich immunoassay. Results. The concentration of cytokine released from PRP varied over time and was influenced by various activation protocols. Ca-only activation had a significant effect on the DS PRPs (where the VEGF, FGF, and IL-1 concentrations were sustained) while Ca/thrombin activation had effects on both SS and DS PRPs (where the PDGF and VEGF concentrations were sustained and the TGF and FGF concentrations were short). The IL-1 content showed a significant increase with Ca-only or Ca/thrombin activation while these activations did not increase the MMP-9 concentration. Conclusion. The SS and DS methods differed in their effect on cytokine release, and this effect varied among the cytokines analysed. In addition, low dose of thrombin/calcium activation increased the overall cytokine release of the PRP preparations over seven days, relative to that with a calcium-only supplement or non-activation. Cite this article: Professor J. H. Oh. Cytokine-release kinetics of platelet-rich plasma according to various activation protocols. Bone Joint Res 2016;5:37–45. doi: 10.1302/2046-3758.52.2000540


We performed this systematic overview on the overlapping meta-analyses that analyzed autologous platelet-rich plasma (PRP) as an adjuvant in the repair of rotator cuff tears and identify the studies which provide the current best evidence on this subject and generate recommendations for the same. We conducted independent and duplicate electronic database searches in PubMed, Web of Science, Scopus, Embase, Cochrane Database of Systematic Reviews, and the Database of Abstracts of Reviews of Effects on September 8, 2021, to identify meta-analyses that analyzed the efficacy of PRP as an adjuvant in the repair of rotator cuff tears. Methodological quality assessment was made using Oxford Levels of Evidence, AMSTAR scoring, and AMSTAR 2 grades and used the Jadad decision algorithm to generate recommendations. 20 meta-analyses fulfilling the eligibility criteria were included. The AMSTAR scores of the included studies varied from 6–10 (mean:7.9). All the included studies had critically low reliability in their summary of results due to their methodological flaws according to AMSTAR 2 grades. The initial size of the tear and type of repair performed do not seem to affect the benefit of PRPs. Among the different preparations used, leucocyte poor (LP)-PRP possibly offers the greatest benefit as a biological augment in these situations. Based on this systematic overview, we give a Level II recommendation that intra-operative use of PRPs at the bone-tendon interface can augment the healing rate, reduce re-tears, enhance the functional outcomes and mitigate pain in patients undergoing arthroscopic rotator cuff repair


Orthopaedic Proceedings
Vol. 96-B, Issue SUPP_11 | Pages 171 - 171
1 Jul 2014
Papalia R Vadalà G Franceschi F Franceschetti E Zampogna B Maffulli N Denaro V
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Introduction. Rotator cuff healing after an arthroscopic repair is discussible because of the high incidence of failures. Among biologic augmentations currently used, platelet-rich plasma (PRP) is one of the most applied, supposed to enhance and accelerate the healing process in different musculoskeletal disorders. However, the evidence supporting its successful administration is still lacking, especially in the field of the rotator cuff repair. Our purpose is to clarify if the recovery is accelerated and the integrity of repaired construct is increased in patients undergoing PRP injections after arthroscopic repair of the rotator cuff. Patients & Methods. Thirty-eight patients with full-thickness rotator cuff tears have been enrolled after they had been informed about the use of PRP and the timing of its application postoperatively. Seventeen patients underwent arthroscopic rotator cuff repair and PRP injections (3 injections at 10 days each other), 21 underwent arthroscopic rotator cuff repair without PRP injections. Outcomes were assessed preoperatively, at 3, 6, 12, and minimum 16 months after surgery (average 17.7 +/− 1.7 months). Constant system, the University of California at Los Angeles (UCLA) system and a Visual Analogue Scale (VAS) scale were used; range of motion and strength in all planes were also assessed. The healing of the repair was assessed at magnetic resonance imaging at a minimum follow up of 6 months from surgery. All patients had the same rehabilitation protocol. Results. Platelet-rich plasma gel application after to arthroscopic rotator cuff tear repairs did not accelerate recovery with respect to pain, range of motion, strength, functional scores, or overall satisfaction as compared with conventional repair at any time point. There was no difference between the 2 groups after 3, 6, 12, months and at final follow up. The follow-up MRI showed no significant difference in the healing rate of the rotator cuff tear. In addition, magnetic resonance imaging, at a minimum of 6 months after surgery, demonstrated a retear rate of 23.5 % in the PRP group and 19% in the conventional group, there was no statistical significance between the groups (P = .658). Discussion/Conclusion. Although PRP application after arthroscopic repair of the rotator cuff has no effects on clinical recovery and structural integrity, it reduces the postoperative occurrence of shoulder stiffness. Further studies should support these findings


Orthopaedic Proceedings
Vol. 102-B, Issue SUPP_11 | Pages 95 - 95
1 Dec 2020
Russo F Ambrosio L Peroglio M Wangler S Guo W Grad S Alini M Vadalà G Papalia R Denaro V
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The use of stem cells transplanted into the intervertebral disc (IVD) is a promising regenerative approach to treat intervertebral disc degeneration (IDD). The aim of this study was to assess the effect of a hydrogel composed of hyaluronic acid (HA) and platelet-rich plasma (PRP) loaded with human mesenchymal stem cells (hMSCs), on IVD extracellular matrix synthesis and nucleus pulposus (NP) marker expression in a whole IVD culture model. HA was blended with batroxobin (BTX), a gelling agent activated in presence of PRP to construct a hydrogel. Bovine IVDs (n=25) were nucleotomised and filled with 1×10. 6. or 2×10. 6. hMSCs suspended in ∼150 mL of the PRP/HA/BTX hydrogel. IVDs harvested at day 0 and nucleotomised IVDs with no hMSCs and/or hydrogel were used as controls. hMSCs alone or encapsulated in the hydrogel were also cultured in well plates to examine the effect of the IVD microenvironment on hMSCs. After 1 week, tissue structure, scaffold integration and gene expression of anabolic (collagen type I, collagen type II and aggrecan), catabolic (matrix metalloproteinase 3 – MMP-3 –, MMP-13 and a disintegrin and metalloproteinase with thrombospondin motifs 4) and NP cell (cytokeratin 19, carbonic anhydrase 12, cluster of differentiation 24) markers were assessed. Histological analysis showed a good integration of the scaffold within the NP area with cell repopulation. At the gene expression level, the hMSC-loaded hydrogels demonstrated to increase disc cell anabolic and catabolic marker expression and promoted hMSC differentiation towards a NP cell phenotype. This study demonstrated that the HA/PRP/BTX may represent a valid carrier for hMSCs being capable of stimulating cell activity and NP marker expression as well as achieving a good integration with the surrounding tissues


Orthopaedic Proceedings
Vol. 100-B, Issue SUPP_14 | Pages 26 - 26
1 Nov 2018
Bastos R do Amaral RJFC Mathias M Andrade R Bastos R Balduino A Schott V Rodeo S Mendes JE
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Intra-articular injections of human mesenchymal stromal cells (MSCs) and platelet-rich plasma (PRP) have been intensively investigated as therapies for knee osteoarthritis (OA) with positive outcomes. In this work we evaluated weather a combination of the treatments (MSCs + PRP) would be beneficial compared to MSCs alone (MSCs) and standard corticosteroid injection (Control group). Forty seven patients (24 males and 23 females; 53.3 ± 10.7 years old) with radiographic symptomatic knee OA (Dejour grades II–IV) were randomized to receive intra-articular injections of MSCs (n = 16), MSCs + PRP (n = 14) or corticosteroid (n=17). MSCs were obtained after mononuclear cells separation from bone marrow aspiration collected from both posterior iliac crests using Sepax automated closed system and expanded in culture until reaching the number of 4 × 10. 7. PRP was obtained by double-centrifugation of whole blood according to a protocol developed in house. After 12 months follow-up, the MSCs and MSCs+PRP groups achieved higher percentages of expected improvement when comparing to the corticosteroid group for the KOOS-symptoms, pain, function and daily living, domains and global score. For the population older or equal to 60 years old the MSCs+PRP group showed significant superiority for the KOOS-ADL domain at 12 months. Cytokines quantification evidenced anti-inflammatory aspects of the treatments. This work evidences the safety and efficacy of intra-articular injection of MSCs for the treatment of early knee OA, with greater improvement with PRP addition particularly to the older population


Orthopaedic Proceedings
Vol. 99-B, Issue SUPP_2 | Pages 101 - 101
1 Jan 2017
Bottegoni C Gigante A
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The objective of this study was to evaluate the safety and the effect of platelet-rich plasma (PRP) intra-articular injections obtained from blood donors (homologous PRP) on elderly patients with early or moderate knee osteoarthritis (OA) who are not candidates for autologous PRP treatment. A total of 60 symptomatic patients, aged 65–86 years, affected by hematologic disorders and early or moderate knee OA, were treated with 5 ml of homologous PRP intraarticular injections every 14 days for a total of three injections. Clinical evaluations before the treatment, and after 2 and 6 months were performed by International Knee Documentation Committee (IKDC), Knee injury and Osteoarthritis Outcome Score (KOOS) and Equal Visual Analogue Scale (EQ VAS) scores. Adverse events and patient satisfaction were recorded. No severe complications were noted during the treatment and the follow-up period. A statistically significant improvement from basal evaluation to the 2-month follow-up visit was observed, whereas a statistically significant worsening from the 2-month to the 6-month follow-up visit was showed. The overall worst results were observed in patients aged 80 years or over and in those affected by minor bone attrition. It was found that 90% of patients were satisfied at the 6-month evaluation. Homologous PRP has an excellent safety profile but offers only a short-term clinical improvement in selected elderly patients with knee OA who are not candidates for autologous PRP treatment. Increasing age and developing degeneration result in a decreased potential for homologous PRP injection therapy. Further studies are needed to confirm these findings


Orthopaedic Proceedings
Vol. 106-B, Issue SUPP_2 | Pages 92 - 92
2 Jan 2024
Fidan B Demirdis I Çiftçi E Aydinli H Kaplan O Çelebier M Boyacioglu Ö Korkusuz P Karanfil Y Korkusuz F
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Platelet Rich Plasma (PRP), either rich (L-PRP) or poor (P-PRP) of leukocytes, is frequently used as an anti-inflammatory and regenerative tool in osteoarthritis (OA). PRP contains proteins but not genes as it is derived from megakaryocytes. Proteomics but not metabolomics of PRP was recently studied. Metabolomics is a field of ‘omics’ research involved in comprehensive portrayal of the small molecules, metabolites, in the metabolome. These small molecules can be endogenous metabolites or exogenous compounds found in an organism (1). Our aim was to determine the difference between L-PRP and P-PRP.

A cross-sectional clinical study was designed in six recreational male athletes between the ages of 18 and 35 years. 3 mL P-PRP and 3 mL -LPRP was prepared from 60 mL of venous blood after treating with 9 mL of sodium citrate and centrifugation at 2.700 rpm for 10 min. Half of the prepared PRP's were frozen at −20°C for a week. Fresh and frozen samples were analyzed at the Q-TOF LC/MS device after thawing to room temperature.

Untargeted metabolomic results revealed that the metabolomic profile of the L-PRP and P-PRP were significantly different from each other. A total of 33.438 peaks were found. Statistically significant (p<0.05) peaks were uploaded to the MetaboAnalyst 5.0 platform. Exogenous out of 2.308 metabolites were eliminated and metabolites found significant for our study were subjected to pathway analysis. Steroid biosynthesis, sphingolipid metabolism and metabolism of lipid pathways were affected. In the L-PRP samples, Nicotinamide riboside (FC: 2.2), MHPG (FC: 3.0), estrone sulfate (FC: 7.5), thiamine diphosphate (FC: 2.0), leukotriene E4 (FC: 7.5), PC(18:1 (9Z)e/2:0) (FC: 9.8) and Ap4A (FC: 2.1) were higher compared to P-PRP. C24 sulfatide (FC: −11.8), 3-hexaprenyl-4,5-dihydroxybenzoic acid (FC: −2.8) metabolites were furthermore lower in P-PRP. Clinical outcomes of PRP application should consider these metabolic pathways in future studies (2).


Orthopaedic Proceedings
Vol. 103-B, Issue SUPP_13 | Pages 135 - 135
1 Nov 2021
Calafiore F Giannetti A Mazzoleni MG Ronca A Taurino F Mandoliti G Calvisi V
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Introduction and Objective

Platelet-Rich-plasma (PRP) has been used in combination with stem cells, from different sources, with encouraging results both in vitro and in vivo in osteochondral defects management. Adipose-derived Stem Cells (ADSCs) represents an ideal resource for their ease of isolation, abundance, proliferation and differentiation properties into different cell lineages. Furthermore, Stem Cells in the adipose tissue are more numerous than from other sources. Aim of this study was to evaluate the potential of ADSCs in enhancing the effect of arthroscopic mesenchymal stimulation combined with infiltration of PRP.

Materials and Methods

The study includes 82 patients. 41 patients were treated with knee arthroscopy, Steadman microfractures technique and intraoperative PRP infiltration, Group A. In the Group B, 41 patients were treated knee arthroscopy, Steadman microfractures and intraoperative infiltration of PRP and ADSCs (Group B). Group A was used as a control group. Inclusion criteria were: Age between 40 and 65 years, Outerbridge grade III-IV chondral lesions, Kellegren-Lawrence Grade I-II. Patient-reported outcome measures (PROMs) evaluated with KOOS, IKDC, VAS, SF-12 were assessed pre-operatively and at 3 weeks, 6 months, 1-year post-operative. 2 patients of Group A and 3 patients of Group B, with indication of Puddu plate removal after high tibial osteotomy (HTO), underwent an arthroscopic second look, after specific informed consent obtained. On this occasion, a bioptic sample was taken from the repair tissue of the chondral lesion previously treated with Steadman microfractures.


Orthopaedic Proceedings
Vol. 100-B, Issue SUPP_3 | Pages 82 - 82
1 Apr 2018
Soufi M Hastie G Wilson J Roy B
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Introduction

Lateral epicondylitis, also known as “tennis elbow,” is a degenerative disorder of the common extensor origin of the lateral humeral epicondyle. The mainstay of treatment is non-operative and includes physiotherapy, activity modification, bracing, nonsteroidal anti-inflammatory drugs, and injections. There is a subgroup of patients however who do not respond to non-operative measures and require operative intervention.

Methods

We conducted a retrospective review of prospectively collected data to assess whether the introduction of PRP injections for lateral epicondylitis led to a reduction in patients subsequently undergoing surgical release.


Orthopaedic Proceedings
Vol. 100-B, Issue SUPP_14 | Pages 129 - 129
1 Nov 2018
Sá MJC Segundo FAS Freitas VML Azevedo AS Silva ACF de Lima GG Devine DM
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The aim of this study was to evaluate the trochlear bone and cartilaginous regeneration of rabbits using a composite based on platelet rich plasma (PRP), chitosan and hydroxyapatite. The study was approved by the ethics committee of the Federal University of Campina Grande under number 72/2017. Surgical holes measuring four millimetres in diameter were performed in rabbit trochleae, one surgical hole in each animal remained empty and another one was filled with the composite. Clinical-orthopaedic and radiographic evaluations were carried out for 60 days, after which the animals were euthanized for histomorphometric evaluations. Clinical-evaluations exhibited lameness of two members of the treatment (T) group and one member of control (C) group. The radiographic evaluation of T group exhibited absence of subchondral bone reaction (33%); nonetheless, presence of moderate subchondral bone reaction was more frequently reported in group C with 67%. Microscopic evaluation revealed the presence of tissue neoformation, composed of dense connective tissue. Microscopic findings were similar in both groups, with a difference in the amount of neoformed tissue, which was confirmed after the morphometric analysis, revealing a significant difference in the quantity of newly formed tissue at the bone / cartilage / implant interface in the T group. The results indicate that the composite based on chitosan, hydroxyapatite and PRP enhanced bone and cartilage healing.


Orthopaedic Proceedings
Vol. 99-B, Issue SUPP_2 | Pages 87 - 87
1 Jan 2017
Zahn J Herrmann M Loibl M Alini M Verrier S
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Angiogenesis is a key factor in early stages of wound healing and is crucial for tissue regeneration. Gold standard for large bone defect treatment is the transplantation of autologous bone grafts, but is not entirely satisfying (e.g. limited amount). Cell therapies and tissue engineering approaches may overcome these problems by using cells and autologous blood components obtainable by less invasive procedures. Pre-clinical studies previously showed promising results combining endothelial progenitor cells (EPCs) and mesenchymal stem cell (MSCs) in polyurethane scaffolds in presence of PRP (1). A systemic investigation of the chemical and mechanical characteristics of different PRP gels formulations suggested their potential use as sustained autologous growth factor delivery system (2). Here we investigate PRP hydrogels as autologous injectable cell delivery systems for EPCs and MSCs and their efficacy in promoting fast neo-vascularization for bone repair applications.

PRP hydrogel and corresponding platelet lysate (PL) were produced from platelet concentrates as described before (3). MSCs were isolated by Ficoll-Paque centrifugation from human bone marrow (EK_regensburg12-101-0127), and cultured in alpha MEM containing 10% FCS and 5 ng/mL basic-FGF (GIBCO). EPCs (CD133+/CD34+) were isolated from MSC fractions using magnetic-activated cell sorting (MACS®) and further cultured in IMDM (GIBCO) containing 5% FCS and 5% PL. GFP positive HUVECs are from Angio-Proteomie, (Boston, USA). Prior to gel encapsulation, MSC and EPCs were pre-stained using PKH26-red® and PKH67-green® respectively. Cells in different proportions were encapsulated in 3D PRP gels, in FDA approved Fibrin gels and in Matrigel®. The gels were cultured in Ibidi microwells placed in an onstage incubator linked to an EVOS Auto Cell Imaging System. The cellular network formation capacity of HUVEC or EPCs and MSC in different proportions was analyzed for the 3 types of hydrogels using time lapse movies recorded over a period of 14 days. Parallel cultures were performed in a classical cell culture CO2 incubator and sample gels were taken at different time points for additional immunostaining and gene expression analysis.

Preliminary results indicate high cell viability in all of the three tested gels. PRP hydrogels present a favorable environment for the formation of a 3 dimensional cellular network in cell co-culture. The formation of these networks was apparent as early as 4 days after seeding. Networks increase in complexity and branching over time. The same was observed when cells were embedded in Matrigel®, which is known for its pro-angiogenic properties. Further experiments are currently in process looking at the involvement of MSCs in this process and the effect of PRP 3D co-culture on their differentiation.

PRP was previously shown as a potent growth factor delivery system for tissue engineering. In the present work, the high cell viability together with the 3 dimensional capillary-like networks observed at early time points suggest that PRP can also be used as an autologous cell delivery and pro-angiogenic system for bone tissue repair.


Orthopaedic Proceedings
Vol. 96-B, Issue SUPP_11 | Pages 195 - 195
1 Jul 2014
Malhotra A Pelletier M Yu Y Christou C Walsh W
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Summary Statement

An autologous thrombin activated 3-fold PRP, mixed with a biphasic calcium phosphate at a 1mL:1cc ratio, is beneficial for early bone healing in older age sheep.

Introduction

The management of bone defects continues to present challenges. Upon activation, platelets secrete an array of growth factors that contribute to bone regeneration. Therefore, combining platelet rich plasma (PRP) with bone graft substitutes has the potential to reduce or replace the reliance on autograft. The simple, autologous nature of PRP has encouraged its use. However, this enthusiasm has failed to consistently translate to clinical expediency. Lack of standardisation and improper use may contribute to the conflicting outcomes reported within both pre-clinical and clinical investigations. This study investigates the potential of PRP for bone augmentation in an older age sheep model. Specifically, PRP dose is controlled to provide clearer indications for its clinical use.


Orthopaedic Proceedings
Vol. 106-B, Issue SUPP_2 | Pages 84 - 84
2 Jan 2024
Tashmetov E Saginova D Kamyshanskiy Y Saginov A Koshanova A
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Various approaches have been implemented to enhance bone regeneration, including the utilization of autologous platelet-rich plasma and bone morphogenetic protein-2. The objective of this study was to evaluate the impact of Marburg Bone Bank-derived bone grafts in conjunction with platelet-rich plasma (PRP), recombinant human bone morphogenetic protein-2 (rhBMP-2), and zoledronic acid (ZA) on osteogenesis within rabbit bone defects. Methodology. Bone defects (5mm in diameter) were created in the femurs of 96 male rabbits. The animals were allocated into five groups: (1) bone graft + PRP (BG + PRP), (2) bone graft + 5μg rhBMP-2 (BG + rhBMP-2), (3) bone graft + 5μg ZA (BG + ZA), (4) bone graft + 10μg rhBMP-2 + 5μg ZA (BG + rhBMP-2 + ZA), and (5) bone graft (BG). Marburg Bone Bank-processed human femoral head allografts were utilized for bone grafting. The rabbits were euthanized at 14-, 30-, and 60-days post-surgery, and their femurs underwent histopathological and histomorphometric assessments. Results. Histomorphometric analysis revealed significantly enhanced de novo osteogenesis within the bone allografts in the BG + PRP and BG + rhBMP-2 groups compared to the BG, BG + ZA, and BG + rhBMP-2 + ZA groups at 14 and 30 days (p < 0.05). However, on day 60, the BG + rhBMP-2 group exhibited elevated osteoclastic activity (early resorption). The local co-administration of ZA with thermally treated grafts impeded both bone graft resorption and new bone formation within the bone defect across all time points. The addition of ZA to BG + rhBMP-2 resulted in diminished osteogenic activity compared to the BG + rhBMP-2 group (p < 0.000). Conclusion. The study findings indicated that the combination of PRP and rhBMP-2 with Marburg bone grafts facilitates early-stage osteogenesis in bone defect healing. Incorporating ZA into the thermally treated bone graft hinders both graft resorption and de novo bone formation


The Journal of Bone & Joint Surgery British Volume
Vol. 89-B, Issue 3 | Pages 417 - 420
1 Mar 2007
Bielecki TM Gazdzik TS Arendt J Szczepanski T Kròl W Wielkoszynski T

Platelet-rich plasma is a new inductive therapy which is being increasingly used for the treatment of the complications of bone healing, such as infection and nonunion. The activator for platelet-rich plasma is a mixture of thrombin and calcium chloride which produces a platelet-rich gel. We analysed the antibacterial effect of platelet-rich gel in vitro by using the platelet-rich plasma samples of 20 volunteers. In vitro laboratory susceptibility to platelet-rich gel was determined by the Kirby-Bauer disc-diffusion method. Baseline antimicrobial activity was assessed by measuring the zones of inhibition on agar plates coated with selected bacterial strains. Zones of inhibition produced by platelet-rich gel ranged between 6 mm and 24 mm (mean 9.83 mm) in diameter. Platelet-rich gel inhibited the growth of Staphylococcus aureus and was also active against Escherichia coli. There was no activity against Klebsiella pneumoniae, Enterococcus faecalis, and Pseudomonas aeruginosa. Moreover, platelet-rich gel seemed to induce the in vitro growth of Ps. aeruginosa, suggesting that it may cause an exacerbation of infections with this organism. We believe that a combination of the inductive and antimicrobial properties of platelet-rich gel can improve the treatment of infected delayed healing and nonunion


Orthopaedic Proceedings
Vol. 106-B, Issue SUPP_1 | Pages 62 - 62
2 Jan 2024
Platania V Tavernaraki N Gontika I Fragiadaki E Triantopoulou N Papadaki H Alpantaki K Vidaki M Chatzinikolaidou M
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Biofabrication is a popular technique to produce personalized constructs for tissue engineering. In this study we combined laponite (Lap), gellan gum (GG) with platelet-rich plasma (PRP) aiming to enhance the endothelial regeneration through the synergistic effects of their individual properties. Laponite has the ability to form porous three-dimensional networks mimicking the extracellular matrix structure, and PRP delivery of growth factors stimulates the endothelial cell proliferation and migration, offering a composite bioink for cell growth and support. The sustained release of these growth factors from the GG-laponite-PRP composite material over time provides a continuous source of stimulation for the cells, leading to more effective tissue engineering strategies for endothelial tissue regeneration. Four blend compositions comprising 1% w/v GG and 0.5 or 1% w/v Lap and 25% v/v PRP were combined with Wharton jelly mesenchymal stem cells (WJ-MSCs) and bioprinted into vessel-like structures with an inner diameter of 3 mm and a wall thickness of 1 mm. Stress/strain analysis revealed the elastomeric properties of the hydrogels with Young modulus values of 10 MPa. Increasing the Lap concentration led to a non-significant decrease of swelling ratio from 93 to 91%. Live/dead assay revealed cell viability of at least 76%, with the 0.5%Lap-GG viability exceeding 99% on day 21. Gradual increase of glycosaminoglycans accumulation and collagen production indicate promotion of ECM formation. The expression and membranous localization of PECAM-1 from day 7 and the granular intracellular localization of vWF after 2 weeks demonstrate in vitro endothelial functionality. In vivo subcutaneous implantation indicated the absence of any adverse immunological reactions. The results reveal the expression of both vWF and PECAM-1 by WJ-MSCs entrapped in all four construct compositions with significantly higher expression of vWF in the presence of PRP


Orthopaedic Proceedings
Vol. 106-B, Issue SUPP_1 | Pages 92 - 92
2 Jan 2024
Forteza-Genestra MA Antich-Rosselló M Ráez-Meseguer C Ramis-Munar G Sangenís AT Calvo J Gayà A Monjo M Ramis JM
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Osteoarthritis (OA) is a degenerative disease that lacks regenerative treatment options. Current research focuses on mesenchymal stem cells (MSCs) and Platelet-Rich Plasma (PRP) as regenerative therapies, but extracellular vesicles (EVs) have shown to be more advantageous. This study compares the regenerative potential of human umbilical cord MSC-derived EVs (cEVs) and platelet-derived EVs (pEVs) in ex vivo and in vivo OA models. In the ex vivo study, OA conditions were induced in human cartilage explants, which were then treated either with pEVs or cEVs. Results showed a higher content of DNA and collagen in the pEVs group compared to control and cEVs groups, suggesting that pEVs could be a potential alternative to cEVs. In the in vivo study, an OA model was established in the knee joints of rats through MIA (monoiodoacetate) injection and then treated either with pEVs or cEVs. Results showed that pEVs-treated knee joints had better subchondral bone integrity and greater OA reversion, particularly in female rats, indicating that pEVs are a viable regeneration treatment for OA and outperform cEVs in terms of efficacy. Overall, the study demonstrates the potential of EVs as a regenerative treatment for OA, with pEVs showing promising results in both ex vivo and in vivo models. The use of pEVs in clinical practice could provide a faster path to translation due to the established use of platelet concentrates in therapeutics. However, further studies are needed to fully evaluate the potential of pEVs for OA treatment and to elucidate the mechanisms behind their regenerative effects. Acknowledgments: The authors thank Dr Fernando Hierro (UIB) for their technical contribution with TEM, Mª Trinidad García (UIB) for the access to radioactivity facilities, Aina Arbós (IUNICS) for her contribution in the histology staining, María Tortosa (IdISBa) for her assistance with the animal care and ADEMA School of Dentistry for the access to the cone beam computed tomography (CBCT). Funding: This research was funded by Instituto de Salud Carlos III, Ministerio de Economía y Competitividad, co-funded by the ESF European Social Fund and the ERDF European Regional Development Fund (MS16/00124; CP16/00124), PROGRAMA JUNIOR del proyecto TALENT PLUS, construyendo SALUD, generando VALOR (JUNIOR01/18), financed by the sustainable tourism tax of the Balearic Islands; the Direcció General d'Investigació and Conselleria d'Investigació, Govern Balear (FPI/2046/2017); the Mecanisme de Recuperació i Resiliència, intended to execute research projects of «Noves polítiques públiques per a un mercat de treball dinàmic, resilient i inclusiu», collected in Pla de Recuperació, Transformació i Resiliència, financed by European Union-Next Generation EU and driven by SOIB and Conselleria de Fons Europeus, Universitat i Cultura i la Conselleria de Model Econòmic, Turisme i Treball (NG0421) and the grant SYN20/03 from IdISBa