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Objectives: Post-reconstruction imaging of the anterior cruciate ligament (ACL) is needed to non-invasively assess in vivo graft maturity before release to play. The purpose of this study was to evaluate autograft remodeling up to 12 months after ACL reconstruction using 3D ultra-short time to echo (UTE) T2* MRI. We hypothesized that the T2* values of ACL autografts would progressively change over time, initially resembling the values for the native contralateral graft source and gradually approaching that of the intact contralateral ACL. Methods: After IRB approval, 12 patients (ages 14-45 years) who underwent primary ACL reconstruction (ACLR) with semitendinosus (SemiT) or bone-patellar tendon-bone (BTB) autograft were enrolled. Patients with a history of prior injury or surgery to either knee were excluded. Subjects returned for UTE MRIs at 1, 3, 6, 9, and 12 months after ACLR. Imaging at 1-month included the contralateral knee. All scans were performed on a Philips Ingenia 3T system. MRI pulse sequences included high-resolution 3D T2 (slice thickness: 0.6mm, TR: 18.7ms; TE: 11.5ms) and a four-echo T2 UTE (slice thickness: 1mm, TR: 20ms; TE: 0.3, 3.3, 6.3, and 9.3ms). Using high-resolution 3D T2 sequences at 1 month, all slices containing the intra-articular ACL were segmented semi-automatically to generate volumetric regions of interest (ROIs) (Materialise, Inc.; Leuven, Belgium). ROIs were divided into proximal/distal and core/peripheral sub-ROIs using standardized methods. Each ROI was co-registered voxel-by-voxel to T2* maps also obtained at 1 month. These T2* maps were then co-registered with those of subsequent time points. The segmentation process was repeated for the 1-, 3-, and 6-month time points by a second reviewer for inter-observer reliability (κ). Statistical differences among the ROIs and sub-ROIs of ACL autografts were assessed with repeated-measures ANOVA and two-tailed non-parametric t-tests. PResults: Twelve subjects were enrolled in this prospective study, and after 2 subjects withdrew, 10 subjects were included in the analysis (n=7 SemiT; n=3 BTB). Nine of 10 subjects attended all postoperative imaging sessions at the time of this writing. Inter-observer reliability for T2* values was found to be excellent (κ=0.832; 95% CI [0.70-0.91]; pConclusions: ACL autografts exhibited progressively increasing T2* values and T2*[inj]/T2*[intact] ratios up to 12 months after ACLR, and these changes appear to be region-dependent. UTE T2* MR imaging can provide unique insights into the condition of remodeling ACL grafts, and may improve our ability to non-invasively assess graft maturity before allowing patients to resume high-intensity activities.

Background Augmentation of soft-tissue repairs with an autologous fibrin clot has been used clinically for nearly four decades; however, fibrin clots tend to produce an abundance of scar tissue, which is known to inhibit soft-tissue regeneration. Mesenchymal stem cells (MSCs) embedded in fibrin clots before repair could reduce scar tissue deposition and facilitate soft-tissue regeneration. To our knowledge, no published studies have directly evaluated the viability or bioactivity of MSCs in fresh human fibrin clots over time. The purpose of this study was to evaluate the viability and bioactivity of human MSCs inside human fibrin clots over time in nutritive and non-nutritive culture media. Questions/purposes We hypothesized that human MSCs would (1) be captured inside fibrin clots and retain their proliferative capacity, (2) remain viable for at least 7 days in the fibrin clots, (3) maintain their proliferative capacity for at least 7 days in the fibrin clots without evidence of active apoptosis, and (4) display similar viability and proliferative capacity when cultured in a non-nutritive medium over the same time periods. Methods Twelve patients (mean age 33.7 years; range 4-72 years) who underwent elective knee surgery were approached between February 2016 and October 2017; all patients agreed to participate and were enrolled. MSCs isolated from human skeletal muscle and banked after prior studies were used for this analysis. On the day of surgery and after expansion of the MSC population, 3-mL aliquots of phosphate-buffered saline containing approximately 600,000 labeled with anti-green fluorescent protein (GFP) antibodies were transported to the operating room, mixed in 30 mL of venous blood from each enrolled patient, and stirred at 95 rpm for 10 minutes to create MSC-embedded fibrin clots. The fibrin clots were transported to the laboratory with their residual blood for analysis. Eleven samples were analyzed after exclusion of one sample because of a processing error. MSC capture was qualitatively demonstrated by enzymatically digesting half of each clot specimen, thus releasing GFP-positive MSCs into culture. The released MSCs were allowed to culture for 7 days. Manual counting of GFP-positive MSCs was performed at 2, 3, 4, and 7 days using an inverted microscope at 100 x magnification to document the change in the number of GFP-positive MSCs over time. The intact remaining half of each clot specimen was immediately placed in proliferation media and allowed to culture for 7 days. On Days 1, 2, 3, 4, and 7, a small portion of the clot was excised, flash-frozen, cryosectioned (8-μm thickness), and immunostained with antibodies specific to GFP, Ki67 (indicative of active proliferation), and cleaved caspase-3 ([CC3]; indicative of active apoptosis). Using an inverted microscope, we obtained MSC cell counts manually at time zero and after 1, 2, 3, 4, and 7 days of culture. Intact fresh clot specimens were immediately divided in half; one half was placed in nutritive (proliferation media) and the other was placed in non-nutritive (saline) media for 1, 2, 3, 4, and 7 days. At each timepoint, specimens were processed in an identical manner as described above, and a portion of each clot specimen was excised, immediately flash-frozen with liquid nitrogen, cryosectioned (8-μm thickness), and visualized at 200 x using an inverted microscope. The numbers of stain-positive MSCs per field of view, per culture condition, per timepoint, and per antibody stain type were counted manually for a quantitative analysis. Raw data were statistically compared using t-tests, and time-based correlations were assessed using Pearson's correlation coefficients. Two-tailed p values of less than 0.05 (assuming unequal variance) were considered statistically significant. Results Green fluorescence, indicative of viable GFP-positive MSCs, was absent in all residual blood samples after 48 hours of culturing; GFP-positive MSCs were visualized after enzymatic digestion of clot matrices. The number of GFP-positive MSCs per field of view increased between the 2-day and 7-day timepoints (mean 5.4 ± 1.5; 95% confidence interval, 4.7-6.1 versus mean 17.0 ± 13.6; 95% CI, 10.4-23.5, respectively; p = 0.029). Viable GFP-positive MSCs were present in each clot cryosection at each timepoint up to 7 days of culturing (mean 6.2 ± 4.3; 95% CI, 5.8-6.6). There were no differences in MSC counts between any of the timepoints. There was no visible evidence of GFP +/CC3 + double-positive MSCs. Combining all timepoints, there were 0.34 ± 0.70 (95% CI, 0.25-0.43) GFP+/Ki67+ double-positive MSCs per field of view. The mitotic indices at time zero and Day 7 were 7.5% ± 13.4% (95% CI, 3.0%-12.0%) and 7.2% ± 14.3% (95% CI, 3.3%-12,1%), respectively (p = 0.923). There was no visible evidence of GFP +/CC3 + double-positive MSCs (active apoptosis) at any timepoint. For active proliferation in saline-cultured fibrin clots, we found averages of 0.1 ± 0.3 (95% CI, 0.0-0.2) and 0.4 ± 0.9 (95% CI, 0.0-0.8) GFP/Ki67 double-positive MSCs at time zero and Day 7, respectively (p = 0.499). The mitotic indices in saline culture at time zero and Day 7 were 2.9% ± 8.4% (95% CI, 0.0%-5.8%) and 9.1% ± 20.7% (95% CI, 1.2%-17.0%; p = 0.144). There was no visible evidence of GFP +/CC3 + double-positive MSCs (active apoptosis) at any timepoint in either culturing condition. Conclusion These preliminary in vitro results show that human MSCs mixed in unclotted fresh human venous blood were nearly completely captured in fibrin clots and that seeded MSCs were capable of maintaining their viability, proliferation capacity, and osteogenic differentiation capacity in the fibrin clot for up to 7 days, independent of external sources of nutrition. Clinical relevance Fresh human fibrin clots have been used clinically for more than 30 years to improve soft-tissue healing, albeit with scar tissue. Our results demonstrate that allogenic human MSCs, which reduce soft-tissue scarring, can be captured and remain active inside human fibrin clots, even in the absence a nutritive culture medium.

Objectives: Chronic rotator cuff tears still represent a significant source of morbidity and functional decline in the general population. The purpose of this study was to establish protocols for isolation and expansion of bursa-derived mesenchymal stem cells (BDSCs) and to evaluate their differentiation capacity, including tenogenesis. We hypothesized that BDSCs would be capable of multilineage differentiation (including tenogenesis) and represent an important source for autologous stem cells for patients undergoing rotator cuff repair. Methods: After IRB approval, 10 patients (ages 43-65 years) scheduled to undergo arthroscopic repair for chronic rotator cuff tears were enrolled. During diagnostic arthroscopy, subacromial bursa tissue was harvested using an arthroscopic shaver and collected by attaching the outflow tubing to a specialized specimen cup. Tissue specimens were transported to our laboratory for analysis. BDSCs were isolated via adherent culture and plated in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% Fetal Bovine Serum (FBS). Chondrogenic, adipogenic, and osteogenic induction media were used to induce differentiation. Tenogenic induction was performed using DMEM supplemented with varying concentrations of BMP-12, ascorbic acid, and human tenocyte-conditioned media. Alcian Blue staining was used to evaluate chondrogenesis, Oil Red O staining for adipogenesis, and Alkaline Phosphatase staining for osteogenesis. Gene expression markers for adipogenesis (ADIPOQ, FABP4, PPARγ), chondrogenesis (COL2A1 and SOX5), and osteogenesis (osteocalcin, osterix), along with primary antibodies to tenogenic markers (scleraxis, tenomodulin), were used to verify each cell lineage. Results: BDSCs isolated by adherent culture without collagen exhibited a spindle-shaped morphology characteristic of mesenchymal stem cells (MSCs), formed colonies, and demonstrated great expandability for six to eight passages without morphology changes (Figure 1A). After 3 weeks of culture, 95% (pConclusion: Our results demonstrate that subacromial bursa represents a viable source of mesenchymal stem cells. We developed a reliable protocol for isolation of BDSCs from patient bursa samples. We show that BDSCs in the presence of BMP-12 and ascorbic acid can differentiate toward a tenogenic lineage. Our work provides strong evidence that BDSCs may be a potent tool for cellular therapy and may benefit future patients who undergo surgical repair of chronic rotator cuff tears. [Figure: see text][Figure: see text]

This study characterized the soft tissue insertion morphometrics on the tibial plateau and their inter-relationships as well as variabilities. The outlines of the cruciate ligament and meniscal root insertions along with the medial and lateral cartilage on 20 cadaveric tibias (10 left and 10 right knees) were digitized and co-registered with corresponding CT-based 3D bone models. Generalized Procrustes Analysis was employed in conjunction with Principal Components Analysis to first create a geometric consensus based on tibial cartilage and then determine the means and variations of insertion morphometrics including shape, size, location, and inter-relationship measures. Step-wise regression analysis was conducted in search of parsimonious models relating the morphometric measures to the tibial plateau width and depth, and basic anthropometric and gender factors. The analyses resulted in statistical morphometric representations for Procrustes-superimposed cruciate ligament and meniscus insertions, and identified only a few moderate correlations (R 2: 0.37–0.49). The study provided evidence challenging the isometric scaling based on a single dimension frequently employed in related morphometric studies, and data for evaluating cruciate ligament reconstruction strategies in terms of re-creating the native anatomy and minimizing the risk of iatrogenic injury. It paved the way for future development of computer-aided personalized orthopaedic surgery applications improving the quality of care and patient safety, and biomechanical models with a better population or average representation.

Objectives: In clinical practice and research, the magnitude of change in patient-reported outcome (PRO) is often used to assess outcome of treatment. However, a clinically meaningful change in PRO may not be associated with an acceptable state that corresponds to “feeling well”, which is also called the Patient Acceptable Symptom State (PASS). Thresholds for common PRO measures for achieving a PASS after ACL reconstruction have not been determined. The purpose of this observational cross section study was to determine thresholds for the International Knee Documentation Committee Subjective Knee Form (IKDC-SKF) and the Knee Injury and Osteoarthritis Outcome Scale (KOOS) that indicate achievement of the PASS. Methods: Patients 1 to 5 years after primary ACL reconstruction completed a survey that included the IKDC-SKF and KOOS. Patients assessed satisfaction with their current state by answering the question “Taking into account all the activity you have during your daily life, your level of pain and also your activity limitations and participation restrictions, do you consider the current state of your knee satisfactory?” The IKDC-SKF and KOOS scores associated with a positive and negative response to this question were determined. Additionally, receiver operator characteristic (ROC) curves and the Youden Index were used to determine the PASS thresholds with the best sensitivity and specificity for each PRO that differentiated patients with an acceptable versus an unacceptable symptom state. Results: One hundred and sixty seven patients (mean age, 28.8±10.9 years) completed the survey. One hundred and forty six patients indicated they were in an acceptable symptom state (PASS-Y) and 21 were not (PASS-N) (Table 1). All PROs were significantly different between the PASS-Y and PASS-N groups (pConclusion: Patients who did not achieve a PASS have worse IKDC-SKF and KOOS scores, indicating limitations in function, sports participation, and knee-related quality of life (QoL). The differences between PASS-Y and PASS-N for the KOOS pain, symptoms and activities of daily living (ADL) subscales were smaller than differences for sports & recreation, knee-related QoL, and the IKDC-SKF. This indicates that after ACL reconstruction, achieving an acceptable symptom state is more dependent on the ability to participate in sports than on the absence of symptoms or participation in ADL. The threshold values identified will aid in determining if patients after ACL reconstruction have achieved an acceptable symptom state based on their IKDC-SKF and KOOS scores.

Objectives: The purpose of the study was to examine the nature and frequency of complications following the most common arthroscopic knee procedures, with particular attention to fellowship training, geographic location of practice, and age and sex of the patient. Methods: Data were obtained from the ABOS database for orthopaedic surgeons who sat for the part II examination from 2003-2009. The database was queried to determine the type and frequency of complications for patients who underwent knee arthroscopy and for those who underwent sports medicine knee arthroscopy including arthroscopic partial meniscectomy, meniscal repair, chondroplasy, microfracture, anterior cruciate ligament reconstruction, or posterior cruciate ligament reconstruction. Factors affecting complication rates that were investigated included type of procedure, fellowship training status, geographic location of practice and age and sex of the patient. Results: There were 4435 complications out of 92,565 knee arthroscopic procedures obtained from the ABOS database for an overall candidate-reported complication rate of 4.8%. The complication rate was highest for PCL reconstruction (20.1%) and ACL reconstruction (9.7%). The complication rates for meniscectomy, meniscal repair, and chondroplasty were 2.8%, 7.7%, and 3.5%, respectively. The complication rate for sports fellowship trained candidates was higher than for non-sports trained candidates (5.1%: sports, 4.1%: no sports) and for male patients (4.9% vs. 4.3%). The complication rate was highest for surgeons in the Northwest (4.9%) and lowest in the South (4.2%). Younger patients (60, 3.6%). The overall rate of pulmonary embolus was 0.11%. Surgical complications were more common than medical or anesthetic complications (Table 1), and infection was the most common complication overall (0.84%). Conclusion: The overall self-reported complication rate for arthroscopic knee procedures was 4.7%, which may be even higher given that the data are self-reported. PCL and ACL reconstruction had the highest complication rate amongst arthroscopic procedures in this study. The increased risk of complications for sports fellowship trained surgeons may be explained by a higher percentage of complex procedures being performed by fellowship trained surgeons. Knee arthroscopy is not a benign procedure and patients should be aware of the risk of complications. In order to reduce the number of complications, surgeons should be aware of the complexity of the procedure, patient factors, and regional differences in care. Limitations of the study include self-reporting of data, which may underestimate their actual complication rate, and that surgeons in their board collection period are more likely to be earlier in their career, which may overestimate the complication rate for more experienced surgeons.

Background: Surgeries employing arthroscopic techniques are among the most commonly performed in orthopaedic clinical practice; however, valid and reliable methods of assessing the arthroscopic skill of orthopaedic surgeons are lacking. Hypothesis: The Arthroscopic Surgery Skill Evaluation Tool (ASSET) will demonstrate content validity, concurrent criterion-oriented validity, and reliability when used to assess the technical ability of surgeons performing diagnostic knee arthroscopic surgery on cadaveric specimens. Study Design: Cross-sectional study; Level of evidence, 3. Methods: Content validity was determined by a group of 7 experts using the Delphi method. Intra-articular performance of a right and left diagnostic knee arthroscopic procedure was recorded for 28 residents and 2 sports medicine fellowship–trained attending surgeons. Surgeon performance was assessed by 2 blinded raters using the ASSET. Concurrent criterion-oriented validity, interrater reliability, and test-retest reliability were evaluated. Results: Content validity: The content development group identified 8 arthroscopic skill domains to evaluate using the ASSET. Concurrent criterion-oriented validity: Significant differences in the total ASSET score ( P < .05) between novice, intermediate, and advanced experience groups were identified. Interrater reliability: The ASSET scores assigned by each rater were strongly correlated ( r = 0.91, P < .01), and the intraclass correlation coefficient between raters for the total ASSET score was 0.90. Test-retest reliability: There was a significant correlation between ASSET scores for both procedures attempted by each surgeon ( r = 0.79, P < .01). Conclusion: The ASSET appears to be a useful, valid, and reliable method for assessing surgeon performance of diagnostic knee arthroscopic surgery in cadaveric specimens. Studies are ongoing to determine its generalizability to other procedures as well as to the live operating room and other simulated environments.