Effects of repetitive platelet-rich plasma application on human tenocyte proliferation.

Current clinical application of platelet-rich plasma is showing a trend toward multiple treatments. The goal of this study was to show the benefit of interval platelet-rich plasma application in the healing and recovery of human tenocytes using an in vitro cell model. Eight volunteers (6 men and 2 women) were included in this study (mean±SD age, 31.6±10.9 years). Venous blood was collected from new blood draws at 3 different times. Two blood products were prepared on each day of treatment: platelet-rich plasma derived from a single-spin process (PRPSS) and platelet-rich plasma derived from a double-spin process (PRPDS). The study had 2 limbs: 2-day and 4-day intervals. Cell proliferation, measured as disintegrations per minute, was then examined via a radioactive thymidine assay. In the 2-day-interval group, the difference in disintegrations per minute between days 0 and 2 in the PRPSS group reached statistical significance (P =.006). In the PRPDS group, statistical difference was seen between days 0 and 4 (P=.001) and between days 2 and 4 (P=.030). In the 4-day-interval group, the difference in disintegrations per minute between days 4 and 8 in the PRPSS group reached statistical significance, showing a decrease in cell proliferation (P =.013). In the PRPDS group, a statistical difference was seen between days 0 and 8 (P=.021), also showing a decrease in cell proliferation. The greatest effect of platelet-rich plasma, which has a positive effect on tenocyte proliferation and recovery, is seen on initial application. Its effect is diminished with repetitive application, and this finding leads to questioning of the efficacy of interval platelet-rich plasma dosing.

Histomorphologic changes of the long head of the biceps tendon in common shoulder pathologies.

PURPOSE: To assess molecular and histologic differences between the proximal (intra-articular) and distal (extra-articular) portions of the long head of the biceps (LHB) tendon in 3 different disease states (biceps instability, tendinosis, and degenerative joint disease [DJD]) compared with a healthy tendon (fresh frozen). METHODS: We used 32 LHB tendons of patients undergoing tenodesis (mean age, 54.7 ± 10.1 years) and 9 harvested tissue donors. Tendons were divided according to 4 diagnostic groups: (1) biceps instability, (2) tendinosis, (3) DJD, and (4) normal control. After sectioning, tendons were fixed in formalin and stained with H&E and alcian blue for histologic analysis. Measurements of collagen organization by use of polarized light microscopy was then performed, and protein expression for type I and type III collagen, tenascin C, and decorin was determined. RESULTS: There were no statistical differences found for protein expression of type I or type III collagen, tenascin C, or decorin. The proximal and distal regions of the tendons had statistically significant differences in alcian blue staining, with the proximal portion containing a higher amount of proteoglycan (instability, P = .001; tendinosis, P = .005; DJD, P = .008; control, P = .011). When compared with the nonpathologic control tendons, a significant increase in alcian blue staining for the proximal region was seen in all 3 groups. Total polarized light analysis showed that the distal tendon had a significantly higher intensity (organization) compared with the proximal tendon (P < .001); this was also seen in all of the diagnostic groups (instability, P = .010; tendinosis, P = .013; DJD, P = .07; control, P = .028). CONCLUSIONS: This study showed a greater degree of degeneration of the proximal (intra-articular) regions of the LHB tendon when compared with the distal regions in all pathologic groups. However, no major differences at the cellular level were found among groups. CLINICAL RELEVANCE: The pathomechanisms of the various forms of known LHB diagnoses are not yet fully understood and basic science studies may help in understanding their etiology and therefore optimizing treatment options.

Graft fixation is highest with anatomic tunnel positioning in acromioclavicular reconstruction.

PURPOSE: The purposes were to determine the bone density at specific bone tunnel locations in the clavicle and to determine ultimate load to failure of a graft fixed with an interference screw at specific areas. METHODS: Bone mass densitometry was tested at 5-mm intervals from the lateral to the medial end of 11 clavicles (mean age, 70.0 ± 17.7 years). Tunnels were drilled in 10-mm increments from the lateral edge, and tenodesis screws were used to fix semitendinosus grafts in the tunnel. Grafts were cyclically loaded, followed by load to failure. RESULTS: The bone mineral density (BMD) of the cadaveric clavicles increased from lateral (0.304 ± 0.078 g/cm(2) at 10 mm) to medial (0.760 ± 0.103 g/cm(2) at 50 mm). Load to failure increased from lateral to medial, and most specimens failed by tendon pullout. The load was 125.3 ± 42.5 N at the most lateral tunnel and 349.3 ± 120.3 N at the most medial tunnel. The Pearson correlation coefficient was 0.653 between tunnel position and load to failure, 0.659 between bone density and load to failure, and 0.803 between tunnel position and bone density. These all showed strong correlation. CONCLUSIONS: BMD shows that optimal bone density is found in the anatomic insertion area of the coracoclavicular ligaments between 20 mm and 50 mm from the lateral end of the clavicle. Low BMD correlated with decreased load to failure. CLINICAL RELEVANCE: Failure at the lateral bone tunnel in coracoclavicular ligament reconstruction may be a result of poor bone quality. When one is drilling bone tunnels for this surgery, consideration should be given to both anatomic position and bone quality.

Comparison of mesenchymal stem cells (osteoprogenitors) harvested from proximal humerus and distal femur during arthroscopic surgery.

PURPOSE: The aim of this study was to examine the relations between age, gender, and number of viable mesenchymal stem cells (MSCs) in concentrated bone marrow (BM) obtained from the proximal humerus and distal femur during arthroscopic surgery. METHODS: BM was aspirated from either the proximal humerus (n = 55) or distal femur (n = 29) during arthroscopic surgery in 84 patients (51.3 ± 11.6 years). MSCs were obtained from fractionated bone marrow after a 5-minute spin at 1,500 rpm. Volume of BM and number of nucleated cells (NCs) were calculated, and samples were cultured for 6 days, after which point colony-forming units (CFUs) were quantified and fluorescence-activated cell sorting (FACS) analysis was performed. Simple linear regression was used to explore relations between age, gender, volume of aspirated BM, and MSCs per milliliter. RESULTS: BM aspirations yielded a mean quantity of 22.6 ± 12.3 mL. After centrifugation, 30.0 ± 16.7 × 10(6) nucleated cells/mL of concentrated BM were harvested. The proximal humerus provided 38.7 ± 52.6 × 10(6), and the distal femur, 25.9 ± 14.3 × 10(6), for an overall 766.3 ± 545.3 MSCs/mL of concentrated BM (proximal humerus: 883.9 ± 577.6, distal femur: 551.3 ± 408.1). Values did not significantly differ by age, gender, or donor site. CONCLUSIONS: Arthroscopic aspiration of bone marrow from the proximal humerus and distal femur is a reproducible technique and yields reliable concentrations of MSCs. The use of an intraoperative concentration method resulted in consistent amounts of MSCs in all clinically relevant age groups without a significant drop of the number of isolated MSCs. CLINICAL RELEVANCE: Human MSCs derived from concentrated bone marrow aspirate are a promising biological addition that may have practical use in the future of soft tissue augmentation. Arthroscopic techniques for bone marrow aspiration that do not require an additional surgical site for aspiration (e.g., iliac crest) or a second operative procedure may facilitate future use of MSCs in arthroscopic surgery.

Biomechanical performance of subpectoral biceps tenodesis: a comparison of interference screw fixation, cortical button fixation, and interference screw diameter.

BACKGROUND: Subpectoral biceps tenodesis with interference screw fixation allows reproducible positioning of the tendon to help maintain the length-tension relationship. The aim of our study was to evaluate the role of cortical button fixation in isolation or as an augment to interference screw fixation and to determine if the diameter of the interference screw affected fixation strength. MATERIALS AND METHODS: Thirty-two cadaveric shoulders were dissected and randomized to 1 of 4 groups: (1) 7-mm interference screw and cortical button, (2) cortical button alone, (3) 7-mm interference screw, or (4) 8-mm interference screw. Testing was performed on a materials testing system with a 100-N load cycled at 1 Hz for 5000 cycles, followed by an axial load to failure test. Cyclic displacement, ultimate load to failure, and site of failure were recorded for each specimen. RESULTS: The mean ultimate failure loads were 7-mm interference screw with cortical button augmentation, 237.8 ± 120.4 N; cortical button alone, 99.4 ± 16.9 N; 7-mm interference screw, 275.5 ± 56 N; 8-mm interference screw, 277.1 ± 42.1 N. All specimens failed through tendon failure at the screw-tendon-bone interface. CONCLUSIONS: The biomechanical performance of subpectoral biceps tenodesis with interference screw fixation was not improved with cortical button augmentation. In addition, cortical button fixation alone yielded a significantly lower ultimate load to failure compared with interference screws. Finally, the biomechanical performance of smaller-diameter interference screws with matching bone tunnels was not affected by interference screw diameter.

The positive effects of different platelet-rich plasma methods on human muscle, bone, and tendon cells.

BACKGROUND: Clinical application of platelet-rich plasma (PRP) in the realm of orthopaedic sports medicine has yielded variable results. Differences in separation methods and variability of the individual may contribute to these variable results. PURPOSE: To compare the effects of different PRP separation methods on human bone, muscle, and tendon cells in an in vitro model. STUDY DESIGN: Controlled laboratory study. METHODS: Blood collected from 8 participants (mean ± SD age 31.6 ± 10.9 years) was used to obtain PRP preparations. Three different PRP separation methods were used: a single-spin process yielding a lower platelet concentration (PRP(LP)), a single-spin process yielding high platelet and white blood cell concentrations (PRP(HP)), and a double-spin that produces a higher platelet concentration and lower white blood cell concentration (PRP(DS)). Human bone, muscle, and tendon cells obtained from discarded tissue samples during shoulder surgery were placed into culture and treated with the 3 PRP preparations, control media (2% fetal bovine serum [FBS] and 10% FBS), and native blood. Radioactive thymidine assays were obtained to examine cell proliferation, and testing with enzyme-linked immunosorbent assay was used to determine growth factor concentrations. RESULTS: Addition of PRP(LP) to osteocytes, myocytes, and tenocytes significantly increased cell proliferation (P ≤ .05) compared with the controls. Adding PRP(DS) to osteoblasts and tenocytes increased cell proliferation significantly (P ≤ .05), but no significance was shown for its addition to myocytes. The addition of PRP(HP) significantly increased cell proliferation compared with the controls only when added to tenocytes (P ≤ .05). Osteoblasts: Proliferation was significantly increased by addition of PRP(LP) compared with all controls (2% FBS, 10% FBS, native blood) (P ≤ .05). Addition of PRP(DS) led to significantly increased proliferation compared with all controls, native blood, and PRP(HP) (P ≤ .05). Proliferation was significantly less when PRP(HP) was added compared with PRP(DS) (P ≤ .05). Myocytes: Proliferation was significantly increased by addition of PRP(LP) compared with native blood (P ≤ .05). Adding PRP(HP) or PRP(DS) to myocytes showed no significant increase in proliferation compared with the controls or the other separations. Tenocytes: Proliferation was significantly increased by addition of PRP(LP) compared with all controls (2% FBS, 10% FBS, native blood) (P ≤ .05). Addition of PRP(DS) showed a significant increase compared with the controls and native blood. For tenocytes, there was a significant increase (P ≤ .05) seen when PRP(HP) was added compared with the controls and native blood but not compared with the other separations. CONCLUSION: The primary findings of this study suggest the application of different PRP separations may result in a potential beneficial effect on the clinically relevant target cells in vitro. However, it is unclear which platelet concentration or PRP preparation may be optimal for the treatment of various cell types. In addition, a "more is better" theory for the use of higher platelet concentrations cannot be supported. This study was not intended to prove efficacy but to provide a platform for future research to be built upon. CLINICAL RELEVANCE: The utilization of different PRP separations may result in a potentially beneficial effect on the clinically relevant target cells in vitro, but it is unclear which platelet concentration or PRP preparation may be optimal for the treatment of various cell types.

Biomechanical evaluation of effect of coracoid tunnel placement on load to failure of fixation during repair of acromioclavicular joint dislocations.

PURPOSE: To evaluate the effect of entry and exit points of the coracoid tunnel on load to failure and mode of failure, to reduce the incidence of coracoid fractures and acromioclavicular joint repair failures. METHODS: This study investigates 5 tunnel placements based on different entry and exit points in the coracoid process: center-center orientation represents perfect placement of the bone tunnel and served as perfect tunnel placement in our study. Four common errors in drilling were then tested and acted as the experimental groups in our study (medial-center, center-medial, lateral-center, and center-lateral). Using 35 cadaveric shoulders (mean age, 68.0 ± 13.0 years), we tested these 5 tunnel orientations using a single repair technique (cortical button) loaded to failure on an MTS 858 Servohydraulic test system (MTS Systems, Eden Prairie, MN). A control group of 7 cadaveric shoulders without the presence of a coracoid tunnel was also tested to determine the type of fracture pattern that occurred. RESULTS: The coracoids without tunnel drilling fractured in patterns similar to traumatic coracoid injuries. With regard to the 5 tunnel groups, it was found that the loads to failure with center-center and medial-center tunnel placement were significantly higher than those with center-medial, center-lateral, and lateral-center tunnel placement. The failure modes of the former were primarily within the repair constructs, whereas those of the latter were primarily due to bony failure. CONCLUSIONS: Our biomechanical results showed a higher peak load to failure with a center-center or medial-center tunnel orientation, which may lessen the risk of coracoid fracture during drilling with a 6-mm cannulated drill bit. CLINICAL RELEVANCE: Proper trajectory of the drill during formation of a coracoid bone tunnel can help reduce the risk of coracoid process fracture and repair failure.

Stability of double-row rotator cuff repair is not adversely affected by scaffold interposition between tendon and bone.

BACKGROUND: Rotator cuff reconstructions may be improved by adding growth factors, cells, or other biologic factors into the repair zone. This usually requires a biological carrier (scaffold) to be integrated into the construct and placed in the area of tendon-to-bone healing. This needs to be done without affecting the constructs mechanics. Hypothesis/ PURPOSE: The hypothesis was that scaffold placement, as an interposition, has no adverse effects on biomechanical properties of double-row rotator cuff repair. The purpose of this study was to examine the effect of scaffold interposition on the initial strength of rotator cuff repairs. STUDY DESIGN: Controlled laboratory study. METHODS: Twenty-five fresh-frozen shoulders (mean age: 65.5 ± 8.9 years) were randomly assigned to 5 groups. Groups were chosen to represent a broad spectrum of commonly used scaffold types: (1) double-row repair without augmentation, (2) double-row repair with interposition of a fibrin clot (Viscogel), (3) double-row repair with interposition of a collagen scaffold (Mucograft) between tendon and bone, (4) double-row repair with interposition of human dermis patch (ArthroFlex) between tendon and bone, and (5) double-row repair with human dermis patch (ArthroFlex) placed on top of the repair. Cyclic loading to measure displacement was performed to 3000 cycles at 1 Hz with an applied 10- to 100-N load. The ultimate load to failure was determined at a rate of 31 mm/min. RESULTS: There were no significant differences in mean displacement under cyclic loading, slope, or energy absorbed to failure between all groups (P = .128, P = .981, P = .105). Ultimate load to failure of repairs that used the collagen patch as an interposition (573.3 ± 75.6 N) and a dermis patch on top of the reconstruction (575.8 ± 22.6 N) was higher compared with the repair without a scaffold (348.9 ± 98.8 N; P = .018 and P = .025). No significant differences were found for repairs with the fibrin clot as an interposition (426.9 ± 103.6 N) and the decellularized dermis patch as an interposition (469.9 ± 148.6 N; P = .73 and P = .35). CONCLUSION: Scaffold augmentation did not adversely affect the zero time strength of the tested standard double-row rotator cuff repairs. An increased ultimate load to failure was observed for 2 of the augmentation methods (collagen patch as an interposition and decellularized dermis patch on top of the reconstruction) compared with the nonaugmented repairs. CLINICAL RELEVANCE: Scaffolds intended for application of growth factors or cellular components in a repair situation did not adversely jeopardize the stability of the operative construct.

Platelet-rich plasma differs according to preparation method and human variability.

BACKGROUND: Varying concentrations of blood components in platelet-rich plasma preparations may contribute to the variable results seen in recently published clinical studies. The purposes of this investigation were (1) to quantify the level of platelets, growth factors, red blood cells, and white blood cells in so-called one-step (clinically used commercial devices) and two-step separation systems and (2) to determine the influence of three separate blood draws on the resulting components of platelet-rich plasma. METHODS: Three different platelet-rich plasma (PRP) separation methods (on blood samples from eight subjects with a mean age [and standard deviation] of 31.6 ± 10.9 years) were used: two single-spin processes (PRPLP and PRPHP) and a double-spin process (PRPDS) were evaluated for concentrations of platelets, red and white blood cells, and growth factors. Additionally, the effect of three repetitive blood draws on platelet-rich plasma components was evaluated. RESULTS: The content and concentrations of platelets, white blood cells, and growth factors for each method of separation differed significantly. All separation techniques resulted in a significant increase in platelet concentration compared with native blood. Platelet and white blood-cell concentrations of the PRPHP procedure were significantly higher than platelet and white blood-cell concentrations produced by the so-called single-step PRPLP and the so-called two-step PRPDS procedures, although significant differences between PRPLP and PRPDS were not observed. Comparing the results of the three blood draws with regard to the reliability of platelet number and cell counts, wide variations of intra-individual numbers were observed. CONCLUSIONS: Single-step procedures are capable of producing sufficient amounts of platelets for clinical usage. Within the evaluated procedures, platelet numbers and numbers of white blood cells differ significantly. The intra-individual results of platelet-rich plasma separations showed wide variations in platelet and cell numbers as well as levels of growth factors regardless of separation method.

Biomechanical evaluation of classic solid and novel all-soft suture anchors for glenoid labral repair.

PURPOSE: To evaluate the biomechanical performance of an all-soft suture anchor (JuggerKnot; Biomet, Warsaw, IN) in comparison with a classic solid suture anchor (2.4-mm biocomposite SutureTak; Arthrex, Naples, FL) in an in vitro labral repair model. METHODS: We dissected 12 cadaveric shoulders (mean age, 61 ± 9.4 years), leaving the labrum intact, and bone mineral density was obtained (mean, 0.375 ± 0.06 g/cm(3)). Simulated labral tears were made at the anteroinferior and posteroinferior edges of the labrum. Repairs used 2 all-soft suture anchors (JuggerKnot) or 2 solid anchors with free, high-strength No. 2 suture (FiberWire; Arthrex) spanning the operative construct to load the repair. Differential variable reluctance transducers were used to measure labral displacement for each specimen. The testing protocol consisted of a preconditioning phase at 10 N for 10 cycles (1 Hz) and then a final load-to-failure testing at a rate of 3 mm/min. Labral displacement of 2 mm was determined as the primary outcome. RESULTS: There was no statistical difference (P = .22) in ultimate load to failure and displacement at ultimate failure (anchor pullout) between the all-soft JuggerKnot (146.0 ± 43.0 N and 19.8 ± 5.4 mm, respectively) and the solid SutureTak (171.9 ± 52.6 N and 22.3 ± 6.8 mm, respectively). The solid anchor had a significantly higher ultimate load at 2 mm of labral displacement than the all-soft suture anchor (84.1 ± 19.0 N and 39.2 ± 10.6 N, respectively; P < .001). CONCLUSIONS: Whereas both the solid SutureTak and the all-soft JuggerKnot displayed similar results on ultimate load-to-failure testing, the solid anchor required significantly greater load for 2 mm of labral displacement than the all-soft anchor. CLINICAL RELEVANCE: The all-soft anchor (JuggerKnot) is similar in biomechanical performance to the classic solid anchor (SutureTak) with the exception of load at 2 mm of labral displacement, suggesting micromotion of the device.

Corticosteroids and local anesthetics decrease positive effects of platelet-rich plasma: an in vitro study on human tendon cells.

PURPOSE: To determine the effects of mixing anesthetics or corticosteroids with platelet-rich plasma (PRP) on human tenocytes in vitro. METHODS: Two separate protocols (double spin and single spin) were used to obtain homologous PRP from the blood of 8 healthy volunteers. Discarded tendon acquired during biceps tenodesis served as tendon specimens for all experiments. After cell isolation, tenocytes were treated in culture with PRP alone or in combination with corticosteroids and/or anesthetics. Fetal bovine serum in concentrations of 2% and 10% served as controls. Cell exposure times of 5, 10, and 30 minutes were used. Radioactive thymidine and luminescence assays were obtained to examine cell proliferation and viability. RESULTS: The presence of lidocaine, bupivacaine, or methylprednisolone resulted in significantly less proliferation than the negative 2% fetal bovine serum control (P < .05). When we compared groups, both lidocaine and bupivacaine had a greater inhibitory effect than methylprednisolone (P < .05). At all time points, viability was significantly decreased in the presence of lidocaine, bupivacaine, or methylprednisolone compared with the negative control (P < .05). CONCLUSIONS: The addition of either anesthetics or corticosteroids to PRP resulted in statistically significant decreases in tenocyte proliferation and cell viability. These results suggest that incorporation of anesthetics or corticosteroids, either alone or in combination, with PRP injection may compromise the potentially beneficial in vitro effects of isolated PRP on tendon cells and compromise cell viability at the site of tendon injury. CLINICAL RELEVANCE: Anesthetics or corticosteroids either alone or in combination should be used carefully to preserve the proposed positive effects of PRP in the treatment of tendon injury.

Biomechanical properties of repairs for dislocated AC joints using suture button systems with integrated tendon augmentation.

PURPOSE: To evaluate the biomechanical performance of different techniques for CC reconstruction using suture button systems with integrated tendon augmentation. Hypothesis was that (1) reconstructions using a cortical button combined with a biological augmentation (semitendinosus allograft) will demonstrate improved stability than a modified Weaver-Dunn procedure and (2) constructs using two tunnels at the clavicle for fixation will show superior horizontal stability than single-tunnel constructs. METHODS: The acromioclavicular joints of 47 cadaveric shoulders were tested for anterior, posterior, and superior translations (70 N load) and maximal load to failure (superior). Shoulders were assigned to 4 groups: (1) native (n = 18) and after sectioning the AC and CC ligaments; (2) CC reconstruction with 1 clavicular and 1 coracoid tunnel (GR-ST) augmented with semitendinosus graft (n = 15); (3) CC reconstruction augmented with semitendinosus tendon (GR-DT) with 2 clavicular and 1 coracoid tunnel (n = 8); and (4) modified Weaver-Dunn reconstruction (n = 6). RESULTS: The Weaver-Dunn demonstrated statistically more translation than the native joint for posterior direction (P = 0.038). The GR-ST had significantly less translation than the Weaver-Dunn for anterior and posterior translations (P = 0.003, P = 0.004) and compared to the native for superior translation (P = 0.028). The GR-DT differed significantly in anterior and posterior translations compared to the Weaver-Dunn (P = 0.002, P = 0.001). The modified Weaver-Dunn failed at significantly less load to failure compared to all other groups (P = 0.002, P = 0.002, P = 0.005). There was no significant difference between the native and the other reconstructions. CONCLUSION: The evaluated techniques for isolated CC ligament reconstruction (GR-ST) in AC joint dislocation showed biomechanical stability superior to the modified Weaver-Dunn procedure and obtained similar measures compared to the native control. A modified technique (GR-DT), which used two fixation points at the clavicle, did not result in decreased horizontal or vertical translation and therefore no superiority of the GR-DT technique could be shown compared to the GR-ST. LEVEL OF EVIDENCE: Controlled laboratory study.

Rapid isolation of human stem cells (connective progenitor cells) from the distal femur during arthroscopic knee surgery.

PURPOSE: (1) To safely obtain bone marrow aspirates from the distal femur during arthroscopic knee surgery, (2) to purify and efficiently concentrate connective tissue progenitor cells (CTPs) in the operating room (OR), and (3) to confirm that these are CTPs through their ability to differentiate into bone cells. METHODS: Bone marrow aspirates were harvested from the distal femur during arthroscopic knee surgery in 26 patients. Twenty-five matched control subjects were selected to evaluate for increased incidence of complications. CTPs were isolated using a rapid method designed for use in the OR compared with 2 accepted methods. Cytochemical and molecular analysis was used to assess osteogenic potential. RESULTS: Osteogenic potential of the CTPs was confirmed by reverse transcription polymerase chain reaction analysis and cellular staining. Bone marrow was successfully aspirated in 25 cases, with 3 incidences of stiffness in the aspirate group compared with 2 in the control group, 1 incidence of a wound irregularity in the aspirate group compared with 1 in the control group, and 3 incidences of hemarthrosis/persistent effusion in the aspirate group compared with 1 in the control group. The rate of complications for the aspirate group was 36% compared with 25% in the control group. CONCLUSIONS: Our intention was to develop a technique for extracting and purifying bone marrow so that the orthopaedic surgeon would have a simple, safe, and efficient process by which to isolate CTPs during arthroscopic knee surgery. This method of aspiration did not lead to a significant increase in complications. Further bone marrow aspirate was successfully purified in the OR, with only a slight increase in surgery time, and resulted in a fractionated layer rich with CTPs. These cells showed osteogenic potential, as evidenced by their osteoblastic differentiation. These CTPs may have future use in enhancing the incorporation of the graft into the bone. LEVEL OF EVIDENCE: Level III, matched case-control study.

In vitro changes in human tenocyte cultures obtained from proximal biceps tendon: multiple passages result in changes in routine cell markers.

PURPOSE: Results of in vitro cell models are commonly used to promote new therapies (e.g., platelet-rich plasma), and clinicians have to be aware of the specific limitations of such models. To gain a sufficient and effective cell load, many current in vitro models use cells multiplied through various passages. This is especially important in tendon-like cell (TLC) models, since native tendon tissue is not available unlimited and contains limited amount of tenocytes. The purpose was to determine the occurrence of phenotypic changes following extended monolayer culture of TLCs, according to cell-passage number. METHODS: Tendon samples were obtained from 15 healthy patients undergoing biceps tenodesis. Tendons were digested and cultured (monolayer) for six passages. Tendon-specific markers (collagens I and III, decorin, tenascin-C, and tenomodulin) and their histology were analyzed using gene expression and protein content assays. Native cells, the cells cultured and cells passaged one to six times were analyzed at each passage. RESULTS: Gene expression of types I and III collagen of cultured TLCs significantly decreased after two passages. Gene expression of decorin, tenascin-C, and tenomodulin exhibited a trend of decreased gene expression with increased passage. Protein levels of types I and III collagen and decorin decreased after four passages. CONCLUSIONS: The significant findings let conclude that tenocyte-like cells obtained from human LHB can be maintained in monolayer culture at low passages, before the signs of phenotypic drift are present. But researchers must be aware of rapid phenotypic drift at higher passage numbers. Therefore, only cells within the first 3 passages should be used as a precaution for in vitro monolayer cell models, and one has to be aware of the phenotypic changes if TLCs passaged multiple times are used. The clinical relevance of this data is that understanding of in vitro TLC models, and their limitations may finally help the clinician to judge the potential of experimental data of new biologic treatment options.

Sequential resection of the distal clavicle and its effects on horizontal acromioclavicular joint translation.

BACKGROUND: Reconstructions of the coracoclavicular (CC) ligament in an open or arthroscopically assisted procedure are often combined with a resection of the distal clavicle to prevent or treat osteoarthritic degenerations of the acromioclavicular (AC) joint. However, increased horizontal translation leading to symptomatic instability may be associated with resection of the distal clavicle. HYPOTHESIS: Horizontal translation increases in direct correlation to subsequent resection of the distal clavicle. STUDY DESIGN: Controlled laboratory study. METHODS: Twelve fresh-frozen specimens (mean age, 82.0 ± 5 years) were examined using a servohydraulic testing system. A 70-N force was applied in the anterior, posterior, and superior directions, and the translations were quantified for all directions. After native testing, sequential resection of the distal clavicle at 5 and 10 mm was performed after incision of the inferior capsule, and anterior-posterior loads were retested. Finally, the AC ligaments were incised, and repeated testing was performed. RESULTS: Native mean (± standard deviation) anterior translation was 7.90 ± 2.62 mm. After section of the inferior AC capsule, anterior translation was 8.05 ± 2.62 mm (P = 1.0). After clavicle resection of 5 mm, anterior translation was 10.75 ± 2.38 mm (P = .122), and after resection of 10 mm, anterior translation was significantly increased to 11.6 ± 2.37 mm (P = .012). Complete AC capsule sectioning led to significantly greater amounts of anterior translation (12.12 ± 3.0 mm; P = .003). Posterior translation for the native specimen was 8.88 ± 2.63 mm. Sectioning of the inferior AC capsule resulted in 9.21 ± 2.71-mm translation (P = 1.0). After clavicle resection of 5 mm, posterior translation was 10.42 ± 2.42 mm (P = 1.0), and after 10-mm resection, it was 11.31 ± 2.86 mm (P = .39). Sectioning of the complete AC capsule led to significantly greater amounts of posterior translation (12.31 ± 3.12 mm; P = .043). CONCLUSION: Ten millimeters of resection increased anterior translation of the distal clavicle with both the superior and posterior AC capsules as well as the CC ligaments intact. Both anterior and posterior translations increased after 10-mm resection and complete AC capsule sectioning. CLINICAL RELEVANCE: These results suggest that resection of the distal clavicle leads to increased horizontal translation despite an intact superior and posterior AC capsule. Only sparing resection of the distal clavicle should be performed and only if strictly indicated. Violation of the AC capsule further increases horizontal translation and should therefore be avoided, or if indicated in AC joint dislocations, a reconstruction of the AC capsule should be considered.

Bone marrow-derived mesenchymal stem cells obtained during arthroscopic rotator cuff repair surgery show potential for tendon cell differentiation after treatment with insulin.

PURPOSE: The purpose of this study was to determine whether a one-time physiologic dose of insulin when compared with the growth factors insulin-like growth factor 1, ß-fibroblastic growth factor, and growth differentiation factor 5 is capable of differentiating bone marrow-derived mesenchymal stem cells (MSCs) into tendon. METHODS: Eleven patients undergoing arthroscopic rotator cuff repair consented to undergo aspiration of bone marrow. A dose-response curve was calculated to determine the optimal dose of insulin needed to differentiate MSCs into tendon. After purification of bone marrow in the operating room, MSCs were exposed to either insulin or tendon-inducing growth factors or were left untreated to serve as a control. The potential for MSCs in each of these groups to differentiate into tendon was evaluated with a multistep process that included determination of the genetic upregulation for tendon-specific proteins, confirmation that the levels of these proteins were actually increased, staining of the MSCs with antibodies for these proteins to ensure that they were expressed on the cell surface, and finally, evaluation of cell morphology to verify the MSCs' tendon-like appearance. RESULTS: MSCs treated with insulin showed increased gene expression of tendon-specific markers (P < .05), increased content of tendon-specific proteins (P < .05), and increased receptors on the cell surface (P < .05) compared with control cells. Histologic analysis showed a tendon-like appearance compared with the control cells. CONCLUSIONS: Bone marrow-derived MSCs treated with a single physiologic dose of insulin differentiated into cells with characteristics consistent with tendon. CLINICAL RELEVANCE: The potential for MSCs to differentiate into tendon after a 1-time dose of insulin may assist in developing practical biologic options for augmentation of rotator cuff repairs.

Biomechanical comparison of arthroscopic repairs for acromioclavicular joint instability: suture button systems without biological augmentation.

BACKGROUND: Arthroscopic procedures for reconstruction of acromioclavicular (AC) joint separations are increasingly used in clinical practice. Multiple surgical techniques exist, but there are still few data on biomechanical performances of commonly used arthroscopic techniques and fixation methods. HYPOTHESIS: Single and double clavicular tunnel reconstructions show comparable primary stability with a modified Weaver-Dunn procedure, and double tunnel constructs show superior horizontal stability. STUDY DESIGN: Controlled laboratory study. METHODS: The AC joints of 40 cadaveric shoulders were tested for anterior, posterior, and superior translation (70-N load) and maximal load to failure (superior) with the MTS 858 Bionix II Servohydraulic testing system. Shoulders were assigned to 4 groups: (1) native (n = 18), (2) coracoclavicular (CC) reconstruction with 1 clavicular and 1 coracoid tunnel (SCT) fixed with a suture pulley and 2 buttons (n = 8), (3) CC reconstruction with 2 clavicular and 1 coracoid tunnel (DCT) fixed with a suture pulley and 3 buttons (n = 8), and (4) modified Weaver-Dunn reconstruction (n = 6). RESULTS: Native specimens showed a mean anterior translation of 7.92 mm (±1.69 mm), a mean posterior translation of 7.84 mm (±2.09 mm), and a superior translation of 4.28 mm (±1.81 mm). Maximal load to failure was 579.44 N (±148.01 N). The SCT technique showed a mean anterior translation of 5.81 mm (±1.16 mm), posterior translation of 8.30 mm (±1.94 mm), and a superior translation of 2.28 mm (±0.52 mm). The maximal load to failure was 591.35 N (±231.17 N). Anterior and superior translations were significantly less compared with the native specimen (P = .005 and P = .003). The DCT technique had an anterior translation of 4.68 mm (±0.6 mm), posterior translation of 6.85 mm (±0.83 mm), and superior translation of 2.09 mm (±0.86 mm). The mean maximal load to failure was 651.16 N (±226.93 N). Anterior and superior translations were significantly less compared with the native specimens (P = .000 and P = .001). No statistically significant differences were shown between SCT and DCT reconstruction for all measurements (P > .05). One reconstruction of the modified Weaver-Dunn procedure failed directly after mounting it into the testing device. The remaining 5 showed a mean anterior translation of 11.36 mm (±3.17 mm), a mean posterior translation of 13.51 mm (±2.21 mm), and a mean superior translation of 3.31 mm (±0.47 mm). Anterior and posterior translations were significantly increased compared with the native specimen (P = .019 and P = .000). The mean maximal load to failure measured 311.13 N (±52.2 N) and was significantly less compared with the native specimen (P = .000). The Weaver-Dunn technique showed significantly less maximal load to failure and more anterior and posterior translation compared with SCT and DCT (P ≤ .05). CONCLUSION: Isolated reconstruction of the CC ligaments using single and double clavicular tunnel techniques results in a high load to failure for superior translation, which is equal to the native stability, and less translation in all 3 directions as well as higher superior stability when compared with the modified Weaver-Dunn procedure. A potential drawback is the risk of coracoid fracture, as the high load to failure of the device may exceed load to failure of cortical bone prior to device breakage. CLINICAL RELEVANCE: Single clavicular tunnel arthroscopic reconstructions of the coracoacromial ligaments show good biomechanical results.

Effects of adding epinephrine to arthroscopic irrigation fluid on cultured chondrocyte survival in vitro.

PURPOSE: This study evaluates the effect of low doses of epinephrine contained in common arthroscopic irrigation solutions on viability of in vitro human articular chondrocytes during short-term exposure. METHODS: Isolated cultured human chondrocytes were treated with culture medium, normal saline solution, 1:300,000 epinephrine solution (equivalent to 10 mL of 1:1,000 epinephrine added to a 3-L saline solution bag), or 1:3,000,000 epinephrine solution (equivalent to 1 mL of 1:1,000 epinephrine added to a 3-L saline solution bag) for 1 hour (N = 84). Twenty-four hours after treatment, chondrocyte viability was measured. Statistical analysis was performed with an analysis of variance with Bonferroni post-test. RESULTS: Chondrocyte viability was significantly better when exposed to normal saline solution alone versus high-dose 1:300,000 epinephrine (87.9% ± 5.4% v 74.6% ± 9.4%, P < .05). Exposure to low-dose 1:3,000,000 epinephrine had significantly better survival versus high-dose 1:300,000 epinephrine (85.0% ± 8.3% v 74.6% ± 9.4%, P < .05). There was no difference in viability after exposure to low-dose 1:3,000,000 epinephrine versus normal saline solution (85.0% ± 8.3% v 87.9% ± 5.4%, P > .05). CONCLUSIONS: In vitro, normal saline solution and low-dose 1:3,000,000 epinephrine are significantly less toxic than high-dose 1:300,000 epinephrine to cultured human articular chondrocytes. CLINICAL RELEVANCE: This in vitro study suggests that arthroscopic irrigation fluid containing 1:3,000,000 epinephrine is less chondrotoxic than solutions containing 1:300,000 epinephrine. Surgeons may wish to use the least amount of epinephrine required for adequate visual clarity during surgery. This study does not establish a contraindication to the use of higher doses of epinephrine.

Human tendon cell response to 7 commercially available extracellular matrix materials: an in vitro study.

PURPOSE: To evaluate the response of human tenocytes in culture to 7 commercially available extracellular matrix (ECM) patches. METHODS: Four samples of each ECM were incubated in human tenocyte cultures by use of standard methods. Cell adhesion, cell proliferation, and cellular production of type I and type III collagen, decorin, and scleraxis were measured for each sample according to established experimental methods. Histologic samples were examined to measure the migration of the tenocytes into the ECM. RESULTS: Tenocytes adhered more to samples of layered submucosal pig intestine than the 6 other ECM materials (P < .002). Tenocytes proliferated more and produced more matrix proteins when cultured on ECM derived from unaltered dermal specimens of human or porcine origin (P < .001). Cells were not seen to have migrated into the matrix of any ECM sample. CONCLUSIONS: Human tenocytes reacted most favorably to dermal ECM samples that were not chemically cross-linked by the manufacturer. Less favorable responses of the human cells were seen when cultured with equine or synthetic ECM, which showed favorable biologic responses in nonhuman models. Cellular migration into the matrix of the ECM is a complex process and cannot be replicated in this model entirely. CLINICAL RELEVANCE: The results of this study suggest that dermal ECM may more favorably react with human tendon tissue than ECM of other origins. This may have great relevance as research continues in the field of augmenting surgical soft-tissue repair.

Material properties and composition of soft-tissue fixation.

Surgical interference screws and suture anchors for attaching soft tissue, such as ligaments and tendons, to bone are routinely used in arthroscopic surgery and sports medicine. Interference screw fixation provides a press fit between bone, graft/tendon, and screw and is frequently used to attach replacement ligaments in tunnels drilled for anterior and posterior cruciate ligament reconstruction. Suture anchors are used in surgical procedures wherein it is necessary for a surgeon to attach (tie) tissue to the surface of the bone, for example, during joint reconstruction and ligament repair or replacement. The composition of these implants ranges from metals to polymers and composites. Typically, because of the relatively large amount of torque that must be applied during insertion, these screws are constructed from metal. However, interference screws and suture anchors have also been constructed from bioabsorbable polymers and composites. The ideal material would (1) provide adequate mechanical fixation, (2) completely degrade once no longer needed, and (3) be completely replaced by bone. Because no material has been shown to be superior for all applications, the surgeon must weigh the advantages and disadvantages of each to evaluate the optimum material for a given application and patient. The purpose of this article is to present a comprehensive review of the commercially available interference screws and suture anchors, with an emphasis on implant composition, interaction, and design. This article provides the orthopaedic surgeon with a background on biomaterials, specifically those used in interference screws and suture anchors. Because there is no material that is perfect for all surgical situations, this review can be used to make educated decisions on a case-by-case basis.