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 Table of Contents  
ORIGINAL ARTICLE
Year : 2017  |  Volume : 42  |  Issue : 1  |  Page : 31-35

Assessment of autologous platelet gel injection in nonunited long bones


1 Department of Orthopaedic Surgery, Faculty of Medicine, Suez Canal University Hospitals, Suez Canal University, Ismailia, Egypt
2 Department of Clinical Pathology, Faculty of Medicine, Suez Canal University Hospitals, Suez Canal University, Ismailia, Egypt

Date of Submission09-May-2016
Date of Acceptance10-May-2016
Date of Web Publication18-May-2017

Correspondence Address:
Noha Kamel
Department of Clinical Pathology, Faculty of Medicine, Suez Canal University Hospitals, Suez Canal University, 4.5 Km Ring Road, Ismailia, 41111
Egypt
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/1110-1067.206430

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  Abstract 

Aim Growing interest in nontransfusional hemoconcentrate use is evident. The aim of this study was to assess the effectiveness of autologous platelet gel injection in nonunited long bones.
Settings and design This single-center prospective study was conducted on 20 patients with long-bone nonunions.
Patients and methods Patients with clinical and radiological signs of long-bone nonunions and eligible for autologous donation were included. Autologous platelets were prepared in blood bank and activated with calcium gluconate and autologous thrombin shortly before injection. Fracture healing was evaluated clinically and with radiographic views in two planes at serial follow-ups of 6 weeks after injection and then every 4 weeks until healing was confirmed.
Results The median time between injury and platelet gel injection was 8 months. Evidence of callus formation was seen in 17 (85%) patients by the end of 17th week; the remaining three patients failed to achieve union at 24 weeks’ follow-up.
Conclusion Platelet gel injection is a safe nonoperative modality for nonunited long bones.

Keywords: femur, fracture, nonunited, platelet gel


How to cite this article:
Tawfik A, Kamel N. Assessment of autologous platelet gel injection in nonunited long bones. Egypt J Haematol 2017;42:31-5

How to cite this URL:
Tawfik A, Kamel N. Assessment of autologous platelet gel injection in nonunited long bones. Egypt J Haematol [serial online] 2017 [cited 2017 Aug 22];42:31-5. Available from: http://www.ehj.eg.net/text.asp?2017/42/1/31/206430


  Introduction Top


The bony skeleton is the most susceptible part of the locomotor system to trauma and high energy shuts. The result of trauma to bones is fractures or deformities. The regeneration of large bone defects without intervention is uncommon [1].

The prevalence of nonunion fractures is reported to be 2–6%, 2–8%, and 3–6.4% in the humerus, femur, and tibia, respectively [2].

PRP is increasingly being used regularly in the musculoskeletal system. Platelet gel has a high platelet concentration that releases bioactive proteins and growth factors necessary to initiate and accelerate tissue repair and regeneration. Platelet gel is effective in bony reconstruction; it provides adhesion for the consolidation of cancellous bone and comminuted fracture segments [3]. A study by Daif [4] concluded that direct application of the Platelet Rich Plasma (PRP) along the fracture lines might enhance bone regeneration.

Adjuvant use of autologous platelet concentrate results in an acceptable time to union and may be a useful adjunct to promote osseous healing in high-risk patients undergoing elective foot and ankle surgery [5].

PRP is commonly used as an autologous product, and hence it is intrinsically free of concerns over transmissible diseases. Therefore, it is generally well accepted by patients and surgeons [6]. Therefore, PRP is gaining increasing clinical interest, directly applied either alone or mixed with grafting materials, for orthopedic applications [7].

PRP injection in nonunited fractures can save an extra surgery and shorten the union time [8]. There are reports on the use of PRP for enhancing fracture healing, treatment of existing nonunion, enhancing bone repair in spinal and ankle fusion, high tibial osteotomy, and distraction osteogenesis [9].

Therefore, the present study assessed the role, feasibility, and effectiveness of a simple, inexpensive autologous platelet gel preparation in the management of long-bone nonunion.


  Patients and methods Top


This prospective case series was carried out in our institute during the period between January 2014 and January 2015. The Institutional Research Review Board of our institute approved the study protocol and the consent forms.

The studied group included 20 consecutive patients with nonunions treated in our hospital. History taking including the date of fracture and risk factors and examination of risk factors that could affect bone healing (systemic and local) were carried out ([Table 1]).
Table 1 All risk factors relevant to bone healing and presence of any risk factor in the studied patients

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Patients were included according to the following criteria:
  1. Nonunion was defined as a fracture that had not shown progressive evidence of healing more than 6 months after injury or more than 4 months from last fracture site operation.
  2. Clinical and radiological signs of nonunion of the long bones, stable internal fixation, or stable reduction with plaster immobilization.
  3. More than 90% contact between the fracture fragments.
  4. Platelet count of more than 150 000/μl for all patients (to ensure better yield of platelet concentrates).


Exclusion criteria

The following patients were excluded from the study: patients with gap nonunions, skin infections, or pseudarthrosis; those with platelet count less than 150 000/μl; patients younger than 18 years; patients with hypofibrogenemia; and patients taking medicines known to influence platelet function (such as aspirin). However, we have to mention that, in a recent study [10], the authors suggested that treatment with PRP could be effective in patients taking chronic antiplatelet therapy as well.

Presurgical evaluation

Nonunions were classified according to the Weber–Cech classification [11] by the orthopedist. A nonhypertrophic nonunion was considered if there was no radiographic bone healing or periosteal bone formation and if the bone ends were atrophic or oligotrophic in appearance (with little or no callus formation), or if a gap between the fracture ends and tapering of the bone ends were present. Preoperative standard radiographs of anteroposterior and lateral projections were obtained.

Platelet gel preparation

Platelet gel preparation was carried out in Suez Canal University Hospital blood bank.

Autologous blood donations were made on the previous or the same day of intervention. The platelet concentrates were prepared in blood bank within the hospital and transferred to the operating room for injection.

Approximately 450 ml of whole blood was collected from every patient in triple blood bags containing 63 ml of CPDA-1 (JMS, Ang Mo Kio, Singapore). The PRP amount was ∼10% of the volume of whole blood that was collected. The whole blood was centrifuged at 750g and 22°C for 7 min; only the upper layer (composed of plasma, platelets, and some white blood cells) was transferred to a satellite bag, which was centrifuged at 5300g and 22°C for 10 min to prepare a platelet concentrate (centrifuge model: RC 12BP Plus; Thermo Scientific Inc., Waltham, Massachusetts, USA). Most of the platelet-poor plasma was transferred to another satellite bag. An average of 50±5 ml of plasma was left with the platelet concentrates.

The platelet concentrate was then used to prepare the platelet gel. The packed red cells were transfused back to the patients.

Determination of platelet and leukocyte count

Platelet and white blood cell count were assessed in whole blood and platelet concentrates with a fully automated hematology analyzer (ABX Micros ES 60; HORIBA ABX Diagnostics, Montpellier, France), on 2 ml of sample taken under sterile conditions from all whole blood units and platelet concentrate units.

Platelet gel preparation

Shortly before the intervention, 10 ml of whole blood was collected in plain tubes from patients and left for 15 min at room temperature for clotting, and then centrifuged at 3000 rpm for 10 min. The supernatant was aspirated with a syringe and was used as autologous thrombin.

About 20 ml of platelet concentrate was activated with 5 ml of serum containing thrombin and 2 ml of calcium gluconate (0.05 g/ml) just 5 min before injecting at the operating room.

Calcium acts as a cofactor of thrombin, which modulates the elongation of fibers during polymerization by promoting lateral branching, and functions in clot stability. Calcium and thrombin activate platelets, allowing the release of growth factors and cytokines. Thus, the fibrin matrix provides an optimized medium for healing [12].

Injection of platelet gel

Activated platelet gel was percutaneously injected without exposing the fracture site. The platelet gel was injected shortly after addition of calcium gluconate and thrombin. The volume injected was usually in the range of 25–30 ml, as the volume tolerated at different sites of injection might be slightly variable. Percutaneous injections of autologous platelet gel were performed under mild anesthesia. An image intensifier was used to precisely locate the area of nonunion.

Outcome measurements

Fracture healing was evaluated clinically and radiologically at serial follow-ups of 6 weeks after injection and then every 4 weeks until healing was confirmed. At each follow-up, standard radiographic views were taken in two planes. Healing was defined radiologically by the presence of a bridging callus, even if the fracture line remained visible, or by bone trabeculae crossing the original nonunion.

Clinical diagnosis of union was determined by the absence of pain and absence of motion at the fracture site upon manual stress testing in the sagittal and coronal planes and with functional recovery of range of motion of the involved extremity.

In cases of suboptimal outcomes by 10–16 weeks after treatment, a computed tomography scan was obtained to consider either repeated platelet gel injections or surgery. All patients were followed up for a minimum of 6 months


  Results Top


Patients’ demographic data including sex, age, mechanism of injury, affected bone, fracture type, and previous interventions are shown in [Table 2]. The study included 20 patients (11 male and 9 female). The mean age was 37.8±12 years. All patients had pain. Nonhypertrophic nonunion on radiographs was confirmed in all patients. The median time between injury and platelet injection was 8 months (range: 6–11 months). Evidence of callus formation was seen in two patients by the end of 8 weeks. By the end of 17th week, 17 of the 20 patients had bridging trabeculae; the remaining three patients (patients 1, 11, and 19) failed to achieve union at 24 weeks’ follow-up.
Table 2 Demographic data and clinical status before intervention

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The method of preparation of PRP and the centrifuge used in the current study gave us PRP with very low leukocytic count (0.15±0.02×103/μl), thus reducing neutrophil secretions that may destroy injured or healthy surrounding cells. The mean initial platelet count in whole blood was 251±39×103/µl. It was significantly increased in PRP after double spin to reach 989±265×103/µl, in which the mean platelet yield in fresh PRP was 3.9.

[Table 1] shows patients with risk factors for impaired or delayed healing, which include type 2 diabetes, nicotine abuse, drugs, neurologic injury, surrounding soft-tissue injuries, comminuted fractures, displaced fractures, and presence of necrotic tissue.

The longest time to union was 17 weeks in one patient with a complex humeral shaft fracture (case 15). Five patients received another injection (cases 1, 5, 11, 12, and 19) after 8 weeks from the first injection of PRP; two of them (cases 5 and 12) succeeded to complete bone union, and the other three patients failed to achieve union. These three patients were treated with a revision of fixation and autologous cancellous iliac bone grafting; all evidenced union at an average 3 months postoperatively ([Table 3]).
Table 3 Time to bone union and weight-bearing

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Weight-bearing or full function of the united bone completed by the end of 24 weeks after injection of PRP, ranging from 16 weeks in case 5 to 26 weeks in case 12, with a mean time of 21 weeks.

No consequent complications such as osteomyelitis, infection, neurovascular injury, or thromboembolic conditions were reported, and, in the 17 patients, there was a good correlation between the radiologic signs of healing and clinical ones. Serial radiographs (case 14, [Table 1]) are illustrated in [Figure 1].
Figure 1 (a–e) A 22-year-old male with road traffic accident (reported as case 14 in [Table 2]). Radiographs showing progression of healing in case 14.

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  Discussion Top


Several researchers have investigated the role of PRP application in the treatment of nonunion fractures with conflicting results [13],[14],[15],[16].

Our study results showed that the use of autologous PRP is a safe management modality that showed a high rate of success for the treatment of long-bone nonunions. Clinical and radiographic signs of healing at 8–17 weeks were achieved in 85% of studied patients.

It is difficult to determine how many injections are sufficient. This minimally invasive treatment option produced fair results after one injection in 15 of 20 (75%) cases, and there were good results after two injections in two cases. It failed in three patients and no trials for further injections were carried out because no signs of bone healing appeared after 12 weeks from second injections, and the three patients refused to undergo further trials.

Recently, Malhotra et al. [17] investigated the efficacy of PRP in the management of 94 patients with established long-bone nonunions. They found, without reported complications, that 82 patients achieved union after 4 months, whereas 12 patients did not show any sign of union after the same duration.

Growth factors are released during different phases of tissue healing and have a crucial role in promoting tissue regeneration. The healing effects of platelet gel are attributed to the numerous growth factors released by activated platelets, such as platelet-derived growth factor, transforming growth factor α and β, epidermal growth factor, fibroblast growth factor, insulin growth factor, platelet-derived epidermal growth factor, platelet-derived angiogenesis factor, IL-8, and angiopoetin [18],[19].

There is growing interest in the role of PRP in integration of bone graft, graft substitutes, or implants in surgically treated patients. The graft may act as a release carrier for growth factors from PRP, favoring a more sustained release of proteins and continuous cellular activation [20].

The pitfall of our study was the lack of a comparison group. Hence, further randomized prospective controlled clinical trials with high statistical power will help to elucidate the outcomes of PRP usage in long-bone nonunions. The choice of hemoconcentrate and its preparation method and whether it must be accompanied with bone graft or not, and the timing and rate of intervention will be the challenges that must be solved for every situation according to injury type and associated risk factors. Further studies are required to tailor this promising treatment option for every patient.

Finally, platelet gel injection is a safe management modality for nonunited long bones.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

 
  References Top

1.
Ghaffarpasand F, Dehghankhalili M, Shahrezaei M. Platelet rich plasma for traumatic non-union fractures: a novel but controversial bone regeneration strategy. Bull Emerg Trauma 2013; 1:99–101.  Back to cited text no. 1
    
2.
Tzioupis C, Giannoudis PV. Prevalence of long-bone non-unions. Injury 2007; 38(Suppl 2):S3–S9.  Back to cited text no. 2
    
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Bhanot S, Alex JC. Current applications of platelet gels in facial plastic surgery. Facial Plast Surg 2002; 18:27–33.  Back to cited text no. 3
    
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Daif ET. Effect of autologous platelet-rich plasma on bone regeneration in mandibular fractures. Dent Traumatol 2013; 29:399–403.  Back to cited text no. 4
    
5.
Bibbo C, Bono CM, Lin SS. Union rates using autologous platelet concentrate alone and with bone graft at high-risk foot and ankle surgery patients. J Surg Orthop Adv 2005; 14:17–22.  Back to cited text no. 5
    
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Mei-Dan O, Lippi G, Sánchez M, Andia I, Maffulli N. Autologous platelet-rich plasma: a revolution in soft tissue sports injury management? Phys Sports Med 2010; 38:127–135.  Back to cited text no. 6
    
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Sheth U, Simunovic N, Klein G, Fu F, Einhorn TA, Schemitsch E et al. Efficacy of autologous platelet-rich plasma use for orthopaedic indications: a meta-analysis. J Bone Joint Surg Am 2012; 94:298–307.  Back to cited text no. 7
    
8.
Malhotra R, Kumar V, Trikha A. Platelet concentrate in treatment of nonunion of long bones. Paper #27. Presented at the American Academy of Orthopaedic Surgeons 2012 Annual Meeting, San Francisco, Feb. 7–11.  Back to cited text no. 8
    
9.
Roberto C, Armando M, Lorenzo N, Birgit R, Massimo I. The use of autologous blood-derived growth factors in bone regeneration. Clin Cases Miner Bone Metab 2011; 8:25–31.  Back to cited text no. 9
    
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Di Matteo B, Filardo G, Lo Presti M, Kon E, Marcacci M. Chronic anti-platelet therapy: a contraindication for platelet-rich plasma intra-articular injections? Eur Rev Med Pharmacol Sci 2014; 18(Suppl):55–59.  Back to cited text no. 10
    
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Weber BG, Cech O. Pseudoarthrosis: pathophysiology, biomechanics, therapy, results. Bern, Switzerland: Hans Huber Publishers; 1976.  Back to cited text no. 11
    
12.
Shimojo AA, Perez AG, Galdames SE, Brissac IC, Santana MH. Performance of PRP associated with porous chitosan as a composite scaffold for regenerative medicine. ScientificWorldJournal 2015; 2015:396131.  Back to cited text no. 12
    
13.
Sun Y, Feng Y, Zhang CQ, Chen SB, Cheng XG. The regenerative effect of platelet-rich plasma on healing in large osteochondral defects. Int Orthop 2010; 34:589–597.  Back to cited text no. 13
    
14.
Galasso O, Mariconda M, Romano G, Capuano N, Romano L, Iannò B et al. Expandable intramedullary nailing and platelet rich plasma to treat long bone non-unions. J Orthop Traumatol 2008; 9:129–134.  Back to cited text no. 14
    
15.
Hakimi M, Jungbluth P, Thelen S, Betsch M, Linhart W, Flohé S et al. Platelet-rich plasma combined with autologous cancellous bone: an alternative therapy for persistent non-union? Unfallchirurg 2011; 114:998–1006.  Back to cited text no. 15
    
16.
Calori GM, Tagliabue L, Gala L, d’Imporzano M, Peretti G, Albisetti W. Application of rhBMP-7 and platelet-rich plasma in the treatment of long bone non-unions: a prospective randomized clinical study on 120 patients. Injury 2008; 39:1391–1402.  Back to cited text no. 16
    
17.
Malhotra R, Kumar V, Garg B, Singh R, Jain V, Coshic P et al. Role of autologous platelet-rich plasma in treatment of long-bone nonunions: a prospective study. Musculoskelet Surg 2015; 99:243–248.  Back to cited text no. 17
    
18.
Frechette JP, Martineau I, Gagnon G. Platelet-rich plasmas: growth factor content and roles in wound healing. J Dent Res 2005; 84:434–439.  Back to cited text no. 18
    
19.
Lieberman JR, Daluiski A, Einhorn TA. The role of growth factors in the repair of bone. Biology and clinical applications. J Bone Joint Surg Am 2002; 84-A:1032–1044.  Back to cited text no. 19
    
20.
Sanchez M, Anitua E, Cugat R, Azofra J, Guadilla J, Seijas R et al. Nonunions treated with autologous preparation rich in growth factors. J Orthop Trauma 2009; 23:52–59.  Back to cited text no. 20
    


    Figures

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    Tables

  [Table 1], [Table 2], [Table 3]



 

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