Platelet Rich Plasma (PRP

IoCP Guidelines

What is PRP?

PRP is derived from autologous blood, with concentrations of platelets above normal baseline levels; it also contains at least 7 growth factors. Normal blood contains approximately 6% platelets, but in PRP there is a concentration of around 94%. This translates to a powerful cocktail containing cytokines and growth factors. These stimulate cellular proliferation and tissue regeneration to dramatically accelerate healing.

Platelets are derived from the patients own blood (autologous), which reduces the chances of unpleasant side effects or infection, as long as sterile techniques are maintained throughout the procedure. It is advised that antiinflammatory medications are stopped prior to treatment to enable the PRP to work to maximum effect. Also, following injection, if pain relief is required, paracetamol is recommended if it is safe for the patient to take. If there is heat or swelling, an ice pack can be used to reduce inflammation.

Although blood is mainly plasma, it also contains a number of solid components ie. Red blood cells, white blood cells and platelets. Platelets are known for their importance in blood clotting. However, platelets are also rich in growth factors, which are extremely important in the healing of injuries. PRP is plasma containing many more platelets than are typically found in blood. The concentrations of platelets, and therefore the concentration of growth factors, can be 5-10 times greater than usual.

Growth Factors in Platelets

Platelet derived growth factor (PDGF -aa, PDGF -bb, PDGF -ab)

Stimulates cell replication Promotes angiogenesis Promotes epithelialisation Promotes granulation tissue formation

Transforming Growth Factor (TGF -B1, TGF B2)

Promotes formation of extracellular matrix Promotes bone cell metabolism

Vascular Endothelial Growth Factor (VEGF)

Promotes angiogenesis

Epidermal Growth Factors (EGF)

Promotes cell differentiation and stimulates re-epithelialisation, angiogenesis and collagenase activity

Fibroblast Growth Factor (FGF)

Promotes proliferation of endothelial cells and fibroblasts

Stimulation of angiogenesis

PRP in Podiatry Practice

Pain Management

The goal of PRP in pain management is to reduce or eliminate pain through healing. The platelets in PRP release growth factors that play a vital role in bone and soft tissue trauma.

Healing Soft Tissue Injuries

PRP injection is exceptionally effective in treating acute soft tissue injuries and chronic tendinopathies, such as Achilles’ tendon repair, acute ligament injury, muscle injury, meniscal repair and intra articular tissue repair.

Orthopaedic Healing

The platelets in PRP accelerate repair and strengthen damaged tissues naturally. By invoking the patients’ inflammatory response, PRP allows healing without significant risks of surgery, such as joint replacement or other invasive procedures. • Plantar fasciitis, sports ligament injuries • Achilles’ tendonitis, tendinopathies, chondromalacia atellae, ankle ligament injury, cartilage damage, degenerative arthritis etc

Whole blood Constituents

93%

red blood cells

6%

platelets

1%

White blood cells

1. Gel Separator System

These tubes use osmosis to insure platelet concentrate gets captured in a gel layer, separated from WBCs and RBCs. The platelet count is lower than other systems but it is the most effective way to collect plasma that has a lower leucocyte count.

2. Buffy Coat System

Kits using buffy coat trap the WBC and platelets in a layer above a separated RBC layer. It produces a high platelet count, but the buffy coat traps the WBCs and a considerable leucocyte count as well.

3. Double Spin Buffy Coat

This means of collection is better able to filter other cell types away from the final product, increasing platelet recovery. It allows for the PRP sample to consist of almost pure, concentrated platelets remaining in a plasma layer, which is easily removed from the gathered RBCs below.

Single Spin

Single spin, such as Eclipse and Regenkit, rely on a soluble polymer to stratify blood elements. The polymer has a specific density between platelets and red blood cells. Upon centrifugation, the gel polymer creates a physical barrier which isolates red blood cells, creating a serum with low haematocrit levels (0%). The complete removal of red blood cells eliminates the potential for an inflammatory response which occurs when red blood cells are introduced outside of vascular pathways. Though RBCs are successfully eliminated, these single spin kits produce a serum with a platelet concentration less than whole blood and are also known as platelet poor plasma.

Dual Spin Kits

Dual spin centrifugation kits apply radial force to stratify blood elements and concentrate platelets up to 6.7 times that of whole blood. Blood is drawn in the presence of an anticoagulant, which prevents platelet activation during the mechanical stress of centrifugation.The sample is spun according to the manufacturer’s recommendations, and the resulting supernatant is collected for a second spin. The supernatant contains plasma, platelets, WBCs and a slight collection of RBCs. The second round of centrifugation is another opportunity for platelets still suspended in plasma to fall out of solution and result in a highly concentrated PRP.

Platelet Concentrations and Patient Outcomes

Single spin kits may have the appeal of a more streamlined process. Unfortunately, the polymer at the centre of the single spin process only targets platelets of a certain density. The exact polymer utilised is unknown, but the substance possibly has a specific density just above that of red blood cells, given the very low haemaocrit levels achieved. This process, therefore, will only capture platelets which correspond to the density of the polymer, whereas platelet density falls along a whole spectrum. Successful PRP therapy harnesses the healing capacity of platelets by recirculating the cell fragments into damage tissue. Platelets are activated in the presence of calcium. Once activated, platelets re-granulate and start to release tiny, bioactive molecules known as growth factors. Growth factors signal surrounding cells to increase migration and proliferation.

Anticoagulants Silva, P et al. 2016. Platelet rich plasma obtained with different anti-coagulants and their effect of platelet numbers and mesenchymal stromal cells’ behaviour in vitro. Stem Cells International Vol 2016

Silva looked at sodium citrate (SC), ethylenecliaminetetracetic acid (EDTA) and anticoagulant citrate dextrose (ACD) during the process of obtaining PRP. Their results indicated that SC produced the highest platelet recovery and minimal change in mesenchymal stromal cell gene expression.

Sodium citrate

1 part SC to 9 parts while blood eg. 1ml SC : 9mls blood

ACD A

ACD A can be more uncomfortable injecting back into the tissues due to its acidity. Again, the ratio is 1:9 Both SC and ACD A are citrate based and utilise the ability to chelate ionised calcium present in blood to prevent coagulation, which forms non-ionised calcium citrate complex calcium ions.

Damaged joints would appear to activate the PRP due to the exposed collagen. Clinical outcomes appear to be very similar in ACD A and SC.

Turn down – Turn up Collection vs Single Spin vs Double Spin

Machado, Z.S. et al. 2019. Turn - down, turn – up: a simple and low cost protocol for preparing PRP.Clinics 74

A very interesting piece of research comparing single spin, double spin and a new, double spin technique using ACD A vacutainers.

Results

The results were very favourable: single spin increased platelet levels to 1.17 x greater than basal concentration. Double spin produced 3.09 x greater than basal concentration

Turn down – turn up increased platelet levels 4.07 x greater than basal concentration

Turn down – Turn up Protocol

1. Collect blood – 8.5mls blood with 1.5mls ACD

2. Centrifuge 200 x g for 15mins, cap of vacutainer facing down

3. Carefully remove tube from centrifuge and maintain in cap down position without turning

4. Carefully aspirate 3.5mls of haematic layer through rubber cap

5. Turn tube to upright (cap up)

6. Centrifuge at 1600 x g for 10 minutes

7. Aspirate 3.5mls of upper portion (PPP)

8. Aspirate 1-2 mls of PRP from lower portion of tube