Table 5.

Pre-clinical studies on PRF for tendons repair.

End Use Destination	Hemocomponent/Experimental Groups	PRF Preparation Protocol	Characterization Parameters	Major Findings	Reference
Sheeps Achilles tendon injected at 2.5 cm proximal to the bone insertion	- Injection of autologous sheep calcified PR-plasma - Injection of autologous sheep PP-plasma - Injection of saline	- Blood collection into 3.8% (wt/vol) sodium citrate - Centrifugation at 4 °C: (a) PR-plasma → 460× g, 8 min (b) PP-plasma → 4500× g, 12 min - Platelet counts before clotting - Addition of calcium chloride at a final concentration of 22.8 mM	- Cell density, morphology and distribution - Vascularization - Inflammation	- Higher increase in cell density in the fascicles treated with PR- and PP-plasma - Ovoid but aligned cells in PR- and PP- treated tendons - Neovascularization is promoted with both PR-and PP-plasma - No inflammatory cells in both PR-and PP-plasma treatment	Anitua et al., 2006 [101]
Sheeps acute model of Achilles tendon rupture	- Re-approximation of the tendon ends with suture only - Re-approximation with suture augmented with ADP wrapped around the repair and sutured to the tendon - ADP wrapped around the proximal and distal margins of the tendon, bridging a 1.5 cm gap, with autologous PRPFM sutured in place within the gap	- PRPFM—Cascade Autologous Platelet System-4, Musculoskeletal Transplant Foundation	- Mechanical tests - Cell and tissue morphology - Vascularization - Scaffold incorporation- Inflammation	- Significant difference in elongation between the operated limb vs unoperated limb in suture only group and ADP + PRPFM group but not in suture + ADP group - No apparent fibrosis in all groups - Increased tendon thickness in suture only group - New tendon fibers without increasing tendon thickness (2/6 animals) in suture + ADP group - Complete bridging of the gap, with no change in tendon thickness in ADP + PRPFM (2/6 animals) - Peripheral integration of the APD to tendon fibers - APD +/− PRPFM augments Achilles tendon repair	Sarrafian et al., 2010 [114]
Dogs patellar tendon; sharp incision of the central third	- Autologous dog PRF membrane to fill the injury site - Surgical closure following resection of the central third of the patellar tendon	- Blood collection in tubes with trisodium citrate and a separator gel. - 1st centrifugation (1100× g, 6 min) - Transfer of PRP supernatant in a vial containing 1.0 M calcium chloride. - 2nd centrifugation (4500× g, 25 min) while fibrin polymerization ensued	- Gross healing assessment and cross-sectional area - Cell density - Vascularization - Collagen and GAG	- Repair tissue in both groups - No histological significant difference (i.e., cellularity, vascularity, collagen organization, or GAG content) - Hypercellular fibrovascular repair tissue in defect site of both groups - Significantly greater cross-sectional area of PRF membrane–treated tendons vs the control group - PRF membrane did not enhance the rate/quality of tendon healing but it increases repair tissue surrounding the defect.	Visser et al., 2011 [117]
Rabbits Toe flexor tendon; sharp transection between the A1 and A2 pulley and immediate surgical repair	- Allogenic PRP - Allogenic PRP-F matrix - Commercial fibrin (Beriplast P Combi Set; CSL Behing K.K., Tokyo, Japan)   Control: Natural healing of the repair site	- Blood collection in syringe with acid citrate dextrose-A - 1st centrifugation (2400 rpm, 10 min at 4 °C) - 2nd centrifugation of plasma (3600 rpm, 10 min at 4 °C) - Platelets count - Addition of fibrin matrix (Beriplast P Combi-Set; CSL Behring K.K., Tokyo, Japan): liquid A (0.25 µL) + liquid B (0.25 µL)	- Edema of the toes - Adhesions extent - Mechanical tests - Histological analysis	- No significant difference in edema/adhesion scores - Significantly increased healing strength by PRP-F matrix	Sato et al., 2012 [115]
Rabbits Experiment 1 Bone-patellar tendon-bone. Removal of the central half of each patellar tendon   Experiment 2 Removal of medial collateral ligament	Experiment 1 - Allogenic rabbit CPFS; Control: untreated defect of the controlateral patella   Experiment 2 Allogenic CPFS sheet Control: Insertion of rivets without reconstruction of the controlatelar medial collateral ligament	- Blood collection in tubes with a sodium citrate solution (5% wt/vol) - 1st centrifugation (3000 rpm, 15 min at 4 °C) - 2nd centrifugation of platelet poor plasma (3000 rpm, 15 min at 4 °C) - Freezing of buffy coat layer and platelet poor plasma (−80 °C) - Defrosting and enriching by ultrafiltration twice of platelet poor plasma; defrosting of buffy coat. - Blending of the two fractions and addition of calcium gluconate (final concentration 23 mM) - Incubation at 37 °C for 3 h - Pressure treatment in aqueous solution of 10 mM calcium chloride at 4 °C	- Repair tissue thickness - Mechanical tests - Inflammation	Experiment 1 - the ultimate failure load and stiffness were higher for the CPFS-treated group than untreated knee - Presence of dense and longitudinally aligned collagen bundles - No signs of immunological rejection of allogenic scaffold Experiment 2 - CPFS promoted ligament repair tissue vs the untreated side - The ultimate failure load of the CPFS repair tissue at 20 weeks was 78% of that in healthy controls of the same age   CPFS enhanced/accelerated healing of tendons and ligaments	Matsunaga et al., 2013 [118]
Rats Tendon-bone insertion site, rotator cuff. Transection and transosseous suture repair of the supraspinatus tendon	- Surgical repair + allogenic PRFM   Control: - Controlateral shoulder, only surgical repair	- Blood collection in syringe with 0.5 cc of acid citrate dextrose anticoagulant solution and thixotropic polyester separator gel. - 1st centrifugation (1500 rpm, 15 min) - 2nd centrifugation of the platelet-rich layer (3000 rpm, 6 min)	- Mechanical tests - Histological analysis (i.e., collagen tissue organization/maturation; cartilage formation	- Higher ultimate load to failure, stress, and stiffness values for experimental group repairs - No differences in biomechanical testing between the groups - Less collagen organization and cartilage formation at the insertion site in the experimental group - PRF-membrane does not recapitulate the native enthesis with exuberant/disordered healing response with fibrovascular scar tissue	Hasan et al., 2016 [108]
Rabbits Flexor digitorum profundus tendon	Part I - Autologous rabbit PRF, wrapped around the repair site, tagged with suture Part II - Autologous rabbit PRF interposed between the tendon repair ends by a 2-strand repair   Control: Control tendons	- Blood collection without anticoagulant - Centrifugation (2700 rpm, 12 min at room temperature) - Compression of the PRF clot	- Range of motions analysis - Cross-sectional area - Mechanical tests	- No significant increase in range of motion - Significant increase in cross-sectional area of the tendons in the PRF group - The control had a higher load and stress to failure but similar stiffness and modulus to the PRF groups - The PRF did not have a major influence on cellular organization - Undesirable effect on the biomechanical properties of repaired flexor tendons	Liao et al., 2017 [116]

ADP, Acellular Porcine Dermal patch; CPFS, Compact Platelet-rich Fibrin Scaffold; GAGs, Glycosaminoglycan; PLTs, Platelets concentration; PP, Platelet-Poor; PR, Platelet-rich; PRFM, Platelet Rich Fibrin Matrix; PRP-F matrix, Platelet-Rich Plasma and Fibrin matrix; vs, versus; WB, Whole Blood; +/−, with or without.