It is widely accepted that growth factors play a central role in the healing process and tissue regeneration [4
]. This conclusion has lead to significant research efforts examining varying growth factors and their role in repair of tissues [4
]. However, there are conflicting reports in the literature regarding potential benefits. Although some authors have reported improved bone formation and tissue healing with PRP, others have had less success [4
]. These varying results are likely attributed to the need for additional standardized PRP protocols, preparations, and techniques. There are a variety of commercially FDA approved kits available with variable platelet concentrations, clot activators, and leukocyte counts which could theoretically affect the data.
Alpha granules are storage units within platelets, which contain pre-packaged growth factors in an inactive form (Fig. ). The main growth factors contained in these granules are transforming growth factor beta (TGFbeta), vascular endothelial growth factor (VEGF) platelet-derived growth factor (PDGF), and epithelial growth factor (EGF) (Table ). The granules also contain vitronectin, a cell adhesion molecule which helps with osseointegration and osseoconduction.
Table 1 Growth factor chart [Printed with permission from: Eppley BL, Woodell JE, Higgins J. Platelet quantification and growth factor analysis from platelet-rich plasma: implications for wound healing. Plast Reconstr Surg. 2004 November;114(6):1502–8] (more ...)
TGFbeta is active during inflammation, and influences the regulation of cellular migration and proliferation; stimulate cell replication, and fibronectin binding interactions [23
] (Fig. ). VEGF is produced at its highest levels only after the inflammatory phase, and is a potent stimulator of angiogenesis. Anitua et al. showed that in vitro VEGF and Hepatocyte Growth Factor (HGF) considerably increased following exposure to the pool of released growth factors; suggesting they accelerate tendon cell proliferation and stimulate type I collagen synthesis [11
]. PDGF is produced following tendon damage and helps stimulate the production of other growth factors and has roles in tissue remodeling. PDGF promotes mesenchymal stem cell replication, osteoid production, endothelial cell replication, and collagen synthesis. It is likely the first growth factor present in a wound and starts connective tissue healing by promoting collagen and protein synthesis [7
]. However, a recent animal study by Ranly et al. suggests that PDGF may actually inhibit bone growth [24
In vitro and in vivo studies have shown that bFGF is both a powerful stimulator of angiogenesis and a regulator of cellular migration and proliferation [23
]. IGF-I is highly expressed during the early inflammatory phase in a number of animal tendon healing models, and likely assists in the proliferation and migration of fibroblasts and to increase collagen production [23
]. However, a laboratory analysis of human PRP samples demonstrated increased concentrations of PDGF, TGFbeta, VEGF, and EGF, while not showing an increase in IGF-1 [25
]. EGF effects are limited to basal cells of skin and mucous membrane while inducing cell migration and replication.
Various blood separation devices have differing preparation steps essentially accomplishing similar goals. The Biomet Biologics GPS III system is described here for simplicity. About 30–60 ml of venous blood is drawn with aseptic technique from the anticubital vein. An 18 or 19 g butterfly needle is advised, in efforts of avoiding irritation and trauma to the platelets which are in a resting state. The blood is then placed in an FDA approved device and centrifuged for 15 min at 3,200 rpm (Fig. ). Afterward, the blood is separated into platelet poor plasma (PPP), RBC, and PRP. Next the PPP is extracted through a special port and discarded from the device (Fig. ). While the PRP is in a vacuumed space, the device is shaken for 30 s to re-suspend the platelets. Afterwards the PRP is withdrawn (Fig. ). Depending on the initial blood draw, there is approximately 3 or 6 cc of PRP available.
GPS III system and centrifuge
GPS III system, withdrawing of platelet poor plasma to be discarded
GPS III withdrawing of platelet rich plasma for injection/graft
The area of injury is marked while taking into account the clinical exam, and data from imaging studies such as MRI and radiographs. It is recommended to use dynamic musculoskeletal ultrasound with a transducer of 6–13 Hz in an effort to more accurately localize the PRP injection. Under sterile conditions, the patient receives a PRP injection with or without approximately 1 cc of 1% lidocaine and 1 cc of 0.25 Marcaine directly into the area of injury. Calcium chloride and thrombin may be added to provide a gel matrix for the PRP to adhere to, potentially maximizing the benefit in the case of a joint space. We recommend using a peppering technique spreading in a clock-like manner to achieve a more expansive zone of delivery. The patient is observed in a supine position for 15–20 min afterwards, and is then discharged home. Patients typically experience minimal to moderate discomfort following the injection which may last for up to 1 week. They are instructed to ice the injected area if needed for pain control in addition to elevation of the limb and modification of activity as tolerated. We recommend acetaminophen as the optimal analgesic, or Vicodin for break through pain, and dissuade the use of NSAID’s in the early post-injection period (Fig. ).
Musculoskeletal ultrasound, common extensor tendinosis
Any concerns of immunogenic reactions or disease transfer are eliminated because PRP is prepared from autologous blood. No studies have documented that PRP promotes hyperplasia, carcinogenesis, or tumor growth. Growth factors act on cell membranes rather than on the cell nucleus and activate normal gene expression [7
]. Growth Factors are not mutagenic and naturally act through gene regulation and normal wound healing feed-back control mechanisms [6
]. Relative contraindications include the presence of a tumor, metastatic disease, active infections, or platelet count < 10 5/ul Hgb < 10 g/dl. Pregnancy or active breastfeeding are contraindications. Patients with an allergy to Bupivicaine (Marcaine) should not receive a local anesthetic with these substances.
The patients should be informed of the possibility of temporary worsening symptoms after the injection. This is likely due to the stimulation of the body’s natural response to inflammatory mediators. Although adverse effects are uncommon, as with any injection there is a possibility of infection, no relief of symptoms, and neurovascular injury. Scar tissue formation and calcification at the injection site are also remote risks.
An allergic reaction or local toxicity to Bupivacaine HCL or Lidocaine, although uncommon could trigger an adverse reaction. Additionally, when used in surgical applications for grafting or with intra-articular injections, PRP may be combined with calcium chloride and bovine thrombin to form a gel matrix. This bovine thrombin which is used to activate PRP, in the past has been associated with life threatening coagulopathies as a result of antibodies to clotting factors V, XI, and thrombin [7
]. However, since 1997 production has eliminated contamination of bovine thrombin with bovine factor Va. Prior to 1997, Va levels were 50 mg/ml and now are <0.2 mg/ml with no further reports of complications [6