Biomechanical And Biological Evaluation Of A Model Of Post-Traumatic Osteoarthritis Following Noninvasive, Traumatic Rupture Of The Anterior Cruciate Ligament

Post-traumatic osteoarthritis (PTOA) is a prevalent condition following rupture of the anterior cruciate ligament (ACL). While numerous animal models of PTOA exist, most are based on surgical disruption of a stabilizing structure. In the rat, surgical ACL transection is the most commonly employed model, but it may introduce confounding biological factors due to surgery. The purpose of this dissertation was to utilize the tibial compression model of ACL injury to induce a noninvasive ACL rupture in the rat. First, a biomechanical characterization of four different loading protocols was undertaken, and a high-speed, high-displacement protocol was deemed optimal for inducing a repeatable, complete ACL injury. Tibiofemoral joint motion was found to be representative of motion during human injury. Next, a chronic, biological comparison of the noninvasive injury model to the surgical ACL transection model was performed. Results indicate that the two models both cause extensive degenerative joint changes, and articular cartilage degeneration was most profound in the medial compartment of the femur. The two models yield in a differential bony remodeling response, and surgical ACL transection causes more drastic degenerative changes of articular cartilage. Furthermore, the ACL transection group had elevated biomarkers of cartilage breakdown compared to the noninvasive rupture group. Lastly, the acute response following noninvasive and surgical injury was investigated. Both injuries cause the systemic mobilization of mesenchymal stem cells (MSCs) and elevated stromal-cell derived factor (SDF-1) concentrations in the joint acutely. Biomarkers of cartilage breakdown and metabolism are elevated only slightly immediately after injury. In conclusion, while some studies may benefit from the more rapid onset of PTOA in the surgical ACL transection model, a noninvasive injury model avoids confounding biological factors and may be beneficial for future studies assessing pathology or potential treatment strategies

Error introduced by common reorientation algorithms in the assessment of rodent trabecular morphometry using micro‐computed tomography

Quantitative analyses of bone using micro‐computed tomography (μCT) are routinely employed in preclinical research, and virtual image reorientation to a consistent reference frame is a common processing step. The purpose of this study was to quantify error introduced by common reorientation algorithms in μCT‐based characterization of bone. Mouse and rat tibial metaphyses underwent μCT scanning at a range of resolutions (6–30 μm). A trabecular volume‐of‐interest (VOI) was manually selected. Image stacks were analyzed without rotation, following 45° In‐Plane axial rotation, and following 45° Triplanar rotation. Interpolation was performed using Nearest‐Neighbor, Linear, and Cubic interpolations. Densitometric (bone volume fraction, tissue mineral density, bone mineral density) and morphometric variables (trabecular thickness, trabecular spacing, trabecular number, structural model index) were computed for each combination of voxel size, rotation, and interpolation. Significant reorientation error was measured in all parameters, and was exacerbated at higher voxel sizes, with relatively low error at 6 and 12 μm (max. reorientation error in BV/TV was 2.9% at 6 μm, 7.7% at 12 μm and 36.5% at 30 μm). Considering densitometric parameters, Linear and Cubic interpolations introduced significant error while Nearest‐Neighbor interpolation caused minimal error, and In‐Plane rotation caused greater error than Triplanar. Morphometric error was strongly and intricately dependent on the combination of rotation and interpolation employed. Reorientation error can be eliminated by avoiding reorientation altogether or by “de‐rotating” VOIs from reoriented images back to the original reference frame prior to analysis. When these are infeasible, reorientation error can be minimized through sufficiently high resolution scanning, careful selection of interpolation type, and consistent processing of all images. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2762–2770, 2018.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/146417/1/jor24039_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/146417/2/jor24039.pd

Articular cartilage surface roughness as an imaging‐based morphological indicator of osteoarthritis: A preliminary investigation of osteoarthritis initiative subjects

Current imaging‐based morphometric indicators of osteoarthritis (OA) using whole‐compartment mean cartilage thickness (MCT) and volume changes can be insensitive to mild degenerative changes of articular cartilage (AC) due to areas of adjacent thickening and thinning. The purpose of this preliminary study was to evaluate cartilage thickness‐based surface roughness as a morphometric indicator of OA. 3D magnetic resonance imaging (MRI) datasets were collected from osteoarthritis initiative (OAI) subjects with Kellgren–Lawrence (KL) OA grades of 0, 2, and 4 (n = 10/group). Femoral and tibial AC volumes were converted to two‐dimensional thickness maps, and MCT, arithmetic surface roughness (Sa), and anatomically normalized Sa (normSa) were calculated. Thickness maps enabled visualization of degenerative changes with increasing KL grade, including adjacent thinning and thickening on the femoral condyles. No significant differences were observed in MCT between KL grades. Sa was significantly higher in KL4 compared to KL0 and KL2 in the whole femur (KL0: 0.55 ± 0.10 mm, KL2: 0.53 ± 0.09 mm, KL4: 0.79 ± 0.18 mm), medial femoral condyle (KL0: 0.42 ± 0.07 mm, KL2: 0.48 ± 0.07 mm, KL4: 0.76 ± 0.22 mm), and medial tibial plateau (KL0: 0.42 ± 0.07 mm, KL2: 0.43 ± 0.09 mm, KL4: 0.68 ± 0.27 mm). normSa was significantly higher in KL4 compared to KL0 and KL2 in the whole femur (KL0: 0.22 ± 0.02, KL2: 0.22 ± 0.02, KL4: 0.30 ± 0.03), medial condyle (KL0: 0.17 ± 0.02, KL2: 0.20 ± 0.03, KL4: 0.29 ± 0.06), whole tibia (KL0: 0.34 ± 0.04, KL2: 0.33 ± 0.05, KL4: 0.48 ± 0.11) and medial plateau (KL0: 0.23 ± 0.03, KL2: 0.24 ± 0.04, KL4: 0.40 ± 0.10), and significantly higher in KL2 compared to KL0 in the medial femoral condyle. Surface roughness metrics were sensitive to degenerative morphologic changes, and may be useful in OA characterization and early diagnosis. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:2755–2764, 2017.A custom algorithm was used to create two‐dimensional articular cartilage thickness maps of patients from the Osteoarthritis Initiative. Thickness maps demonstrate significantly increased surface roughness as a function of increasing Kellgren–Lawrence (KL) osteoarthritis (OA) grade, particularly in the medial femoral condyle, though mean cartilage thickness was not found to differ significantly between KL grades. Surface roughness‐based metrics have potential utility as morphological indicators of OA.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/141486/1/jor23588_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/141486/2/jor23588.pd

Metabolomic serum profiling after ACL injury in rats: A pilot study implicating inflammation and immune dysregulation in post‐traumatic osteoarthritis

ACL rupture is a major risk factor for post‐traumatic osteoarthritis (PTOA) development. Little information exists on acute systemic metabolic indicators of disease development. Thirty‐six female Lewis rats were randomized to Control or noninvasive anterior cruciate ligament rupture (ACLR) and to three post‐injury time points: 72 h, 4 weeks, 10 weeks (n = 6). Serum was collected and analyzed by 1H nuclear magnetic resonance (NMR) spectroscopy and combined direct injection and liquid chromatography (LC)‐mass spectrometry (MS)/MS (DI‐MS). Univariate and multivariate statistics were used to analyze metabolomic data, and predictive biomarker models were analyzed by receiver operating characteristic (ROC) analysis. Topological pathway analysis was used to identify perturbed pathways. Two hundred twenty‐two metabolites were identified by 1H NMR and DI‐MS. Differences in the serum metabolome between ACLR and Control were dominated by medium‐ and long‐chain acylcarnitine species. Further, decreases in several tryptophan metabolites were either found to be significantly different in univariate analysis or to play important contributory roles to multivariate model separation. In addition to acylcarnitines and tryptophan metabolites, glycine, carnosine, and D‐mannose were found to differentiate ACLR from Control. Glycine, 9‐hexadecenoylcarnitine, trans‐2‐Dodecenoylcarnitine, linoelaidyl carnitine, hydroxypropionylcarnitine, and D‐Mannose were identified as biomarkers with high area under ROC curve values and high predictive accuracies. Our analysis provides new information regarding the potential contribution of inflammatory processes and immune dysregulation to the onset and progression of PTOA following ACL injury. As these processes have most commonly been associated with inflammatory arthropathies, larger‐scale studies elucidating their involvement in PTOA development and progression are necessary. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:1969–1979, 2018.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/145352/1/jor23854.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/145352/2/jor23854_am.pd

Sex differences in clinical outcomes following surgical treatment of femoroacetabular impingement

BACKGROUND: Sex-based differences in clinical outcomes following surgical treatment of femoroacetabular impingement remain largely uncharacterized; this prospective, multicenter study evaluated these differences both directly and adjusted for covariates. METHODS: Hips undergoing surgical treatment of symptomatic femoroacetabular impingement were prospectively enrolled in a multicenter cohort. Patient demographics, radiographic parameters, intraoperatively assessed disease severity, and history of surgical procedures, as well as patient-reported outcome measures, were collected preoperatively and at a mean follow-up of 4.3 years. A total of 621 (81.6%) of 761 enrolled hips met the minimum 1 year of follow-up and were included in the analysis; 56.7% of analyzed hips were female. Univariate and multivariable statistics were utilized to assess the direct and adjusted differences in outcomes, respectively. RESULTS: Male hips had greater body mass index and larger α angles. Female hips had significantly lower preoperative and postoperative scores across most patient-reported outcome measures, but also had greater improvement from preoperatively to postoperatively. The preoperative differences between sexes exceeded the threshold for the minimal clinically important difference of the modified Harris hip score (mHHS) and all Hip disability and Osteoarthritis Outcome Score (HOOS) domains except quality of life. Preoperative sex differences in mHHS, all HOOS domains, and Short Form-12 Health Survey physical function component score were greater than the postoperative differences. A greater proportion of female hips achieved the minimal clinically important difference for the mHHS, but male hips were more likely to meet the patient acceptable symptom state for this outcome. After adjusting for relevant covariates with use of multiple regression analysis, sex was not identified as an independent predictor of any outcome. Preoperative patient-reported outcome scores were a strong and highly significant predictor of all outcomes. CONCLUSIONS: Significant differences in clinical outcomes were observed between sexes in a large cohort of hips undergoing surgical treatment of femoroacetabular impingement. Despite female hips exhibiting lower baseline scores, sex was not an independent predictor of outcome or reoperation. LEVEL OF EVIDENCE: Prognostic Level II. See Instructions for Authors for a complete description of levels of evidence

An efficient R1Ï dispersion imaging method for human knee cartilage using constant magnetization prepared turbo- FLASH

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/167754/1/nbm4500.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/167754/2/nbm4500_am.pd

Small-Animal Compression Models of Osteoarthritis.

The utility of nonsurgical, mechanical compression-based joint injury models to study osteoarthritis pathogenesis and treatments is increasing. Joint injury may be induced via cyclic compression loading or acute overloading to induce anterior cruciate ligament rupture. Models utilizing mechanical testing systems are highly repeatable, require little expertise, and result in a predictable onset of osteoarthritis-like pathology on a rapidly progressing timeline. In this chapter, we describe the procedures and equipment needed to perform mechanical compression-induced initiation of osteoarthritis in mice and rats

Early patellofemoral cartilage and bone pathology in a rat model of noninvasive anterior cruciate ligament rupture

OBJECTIVE: Anterior cruciate ligament rupture (ACLR) is a risk factor for the development of post-traumatic osteoarthritis (PTOA). While PTOA in the tibiofemoral joint compartment is well-characterized, very little is known about pathology in the patellofemoral compartment after ACL injury. Here, we evaluated the extent to which ACLR induces early patellofemoral joint damage in a rat model. METHODS: Adult female Lewis rats were randomized to noninvasive ACLR or Sham. Two weeks post-injury, contrast-enhanced micro-computed tomography (µCT) of femoral and patellar cartilage was performed using 20% v/v ioxaglate. Morphometric parameters of femoral trochlear and patellar cartilage, subchondral bone, and trabecular bone were derived from µCT. Sagittal Safranin-O/Fast-Green-stained histologic sections were graded using the OARSI score in a blinded fashion. RESULTS: Cartilage and bone remodelling consistent with an early PTOA phenotype were observed in both femoral trochleas and patellae. ACLR caused osteophyte formation of the patella and pathology in the superficial zone of articular cartilage, including surface fibrillation, fissures, increased cellularity, and abnormal chondrocyte clustering. There were significant increases in thickness of patellar and trochlear cartilage. Loss of subchondral bone thickness, bone volume fraction, and tissue mineral density, as well as changes to patellar and trochlear trabecular microarchitecture, were indicative of catabolic bone remodelling. Several injury-induced changes, including increased cartilage thickness and trabecular spacing and decreased trabecular number were more severe in the patella compared to the trochlea. CONCLUSION: The patellofemoral joint develops mild but evident pathology in the early period following ACL rupture, extending the utility of this model to the study of patellofemoral PTOA

Nondestructive, indirect assessment of the biomechanical properties of the rat intervertebral disc using contrast‐enhanced μCT

Mechanical characterization of the intervertebral disc involves labor‐intensive and destructive experimental methodology. Contrast‐enhanced micro‐computed tomography is a nondestructive imaging modality for high‐resolution visualization and glycosaminoglycan quantification of cartilaginous tissues. The purpose of this study was to determine whether anionic and cationic contrast‐enhanced micro‐computed tomography of the intervertebral disc can be used to indirectly assess disc mechanical properties in an ex vivo model of disc degeneration. L3/L4 motion segments were dissected from female Lewis rats. To deplete glycosaminoglycan, samples were treated with 0 U/ml (Control) or 5 U/ml papain. Contrast‐enhanced micro‐computed tomography was performed following incubation in 40% Hexabrix (anionic) or 30 mg I/ml CA4+ (cationic) for 24 h (n = 10/contrast agent/digestion group). Motion segments underwent cyclic mechanical testing to determine compressive and tensile modulus, stiffness, and hysteresis. Glycosaminoglycan content was determined using the dimethylmethylene blue assay. Correlations between glycosaminoglycan content, contrast‐enhanced micro‐computed tomography attenuation, and mechanical properties were assessed via the Pearson correlation. The predictive accuracy of attenuation on compressive properties was assessed via repeated random sub‐sampling cross validation. Papain digestion produced significant decreases in glycosaminoglycan content and corresponding differences in attenuation and mechanical properties. Attenuation correlated significantly to glycosaminoglycan content and to all compressive mechanical properties using both Hexabrix and CA4+. Predictive linear regression models demonstrated a predictive accuracy of attenuation on compressive modulus and stiffness of 79.8–86.0%. Contrast‐enhanced micro‐computed tomography was highly predictive of compressive mechanical properties in an ex vivo simulation of disc degeneration and may represent an effective modality for indirectly assessing disc compressive properties. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2030–2038, 2018.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/145375/1/jor23850_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/145375/2/jor23850.pd

EPDR1 Governs Metabolic and Immunologic Re-programming of Human Mesenchymal Stem Cells During Osteoblast Differentiation

EPDR1 is a novel human osteoblast regulatory gene previously identified by intersecting BMD GWAS datasets with promoter-focused Capture-C and ATAC-seq generated in differentiating human osteoblasts. Recently, we showed that the open chromatin region harboring the BMD variants rs1524068, rs6975644 and rs940347 functions as an osteoblast specific EPDR1 enhancer in hFOB1.19 cells. However, the precise molecular processes controlled by EPDR1 is unknown. Herein, we knocked-down EPDR1 expression in three unique bone-marrow derived human mesenchymal stem cell donor lines (hMSC) using small interfering RNA and evaluated the global transcriptomic changes with or without BMP2 stimulation using osteopermissive media. Globally, 3,861 genes were differentially regulated (1,888 upregulated and 1,973 downregulated) by BMP in non-targeted cells, whereas 2,744 genes were differentially regulated (1,528 upregulated and 1,216 downregulated) in EPDR1 silenced cells. As expected, genes associated with osteoblast differentiation, BMP signaling pathway, Notch signaling pathway and tissue morphogenesis were expressed in non-targeted cells; however, none of these biological processes achieved significance in EPDR1 silenced cells, suggesting major pathway perturbations. To understand these changes, we performed a donor-specific comparison with EPDR1-silenced cells. Of the ~2,300 genes differentially regulated, we observed an upregulation of genes related to inflammatory response, cytokine-mediated signaling pathway, and regulation of T cell activity, while genes related to cell division were decreased. Additionally, we observed differentially expressed genes enriched for pathways involved in regulation of fatty acid metabolic process, regulation of bone resorption, arachidonic acid metabolic process, negative regulation of collagen biosynthetic process and positive regulation of tumor necrosis factor, interferon-gamma, and interleukin-8 production upon BMP2 stimulation. The crystal-structure of EPDR1 protein suggests human EPDR1 folds into a dimer using a monomeric subunit consisting of a deep hydrophobic pocket to bind to hydrophobic fatty acids and function as a lipoprotein carrier. Since EPDR1 is a secreted protein, an immune-reactive profile driven by loss of EPDR1 could be metabolically mediated via disturbances in cellular lipoprotein and fatty acid trafficking. We conclude that EPDR1 could play an important role in pathophysiological bone turnover later in life and therefore warrants further investigation.http://deepblue.lib.umich.edu/bitstream/2027.42/191456/2/EPDR1 siR RNA SEQ V2 kdh.pd