Increased tissue modulus and hardness in the TallyHO mouse model of early onset type 2 diabetes mellitus.

Individuals with type 2 diabetes mellitus (T2DM) have a higher fracture risk compared to those without T2DM despite having higher bone mineral density (BMD). Thus, T2DM may alter other aspects of resistance to fracture beyond BMD such as bone geometry, microarchitecture, and tissue material properties. We characterized the skeletal phenotype and assessed the effects of hyperglycemia on bone tissue mechanical and compositional properties in the TallyHO mouse model of early-onset T2DM using nanoindentation and Raman spectroscopy. Femurs and tibias were harvested from male TallyHO and C57Bl/6J mice at 26 weeks of age. The minimum moment of inertia assessed by micro-computed tomography was smaller (-26%) and cortical porosity was greater (+490%) in TallyHO femora compared to controls. In three-point bending tests to failure, the femoral ultimate moment and stiffness did not differ but post-yield displacement was lower (-35%) in the TallyHO mice relative to that in C57Bl/6J age-matched controls after adjusting for body mass. The cortical bone in the tibia of TallyHO mice was stiffer and harder, as indicated by greater mean tissue nanoindentation modulus (+22%) and hardness (+22%) compared to controls. Raman spectroscopic mineral:matrix ratio and crystallinity were greater in TallyHO tibiae than in C57Bl/6J tibiae (mineral:matrix +10%, p < 0.05; crystallinity +0.41%, p < 0.10). Our regression model indicated that greater values of crystallinity and collagen maturity were associated with reduced ductility observed in the femora of the TallyHO mice. The maintenance of structural stiffness and strength of TallyHO mouse femora despite reduced geometric resistance to bending could potentially be explained by increased tissue modulus and hardness, as observed at the tibia. Finally, with worsening glycemic control, tissue hardness and crystallinity increased, and bone ductility decreased in TallyHO mice. Our study suggests that these material factors may be sentinels of bone embrittlement in adolescents with T2DM.

Chronic kidney disease and aging differentially diminish bone material and microarchitecture in C57Bl/6 mice.

Chronic kidney disease (CKD) is a common disease of aging and increases fracture risk over advanced age alone. Aging and CKD differently impair bone turnover and mineralization. We thus hypothesize that the loss of bone quality would be greatest with the combination of advanced age and CKD. We evaluated bone from young adult (6 mo.), middle-age (18 mo.), and old (24 mo.) male C57Bl/6 mice three months following either 5/6th nephrectomy, to induce CKD, or Sham procedures. CKD exacerbated losses of cortical and trabecular microarchitecture associated with aging. Aging and CKD each resulted in thinner, more porous cortices and fewer and thinner trabeculae. Bone material quality was also reduced with CKD, and these changes to bone material were distinct from those due to age. Aging reduced whole-bone flexural strength and modulus, micrometer-scale nanoindentation modulus, and nanometer-scale tissue and collagen strain (small-angle x-ray scattering [SAXS]. By contrast, CKD reduced work to fracture and variation in bone tissue modulus and composition (Raman spectroscopy), and increased percent collagen strain. The increased collagen strain burden was associated with loss of toughness in CKD. In addition, osteocyte lacunae became smaller, sparser, and more disordered with age for Sham mice, yet these age-related changes were not clearly observed in CKD. However, for CKD, larger lacunae positively correlated with increased serum phosphate levels, suggesting that osteocytes play a role in systemic mineral homeostasis. This work demonstrates that CKD reduces bone quality, including microarchitecture and bone material properties, and that loss of bone quality with age is compounded by CKD. These findings may help reconcile why bone mass does not consistently predict fracture in the CKD population, as well as why older individuals with CKD are at high risk of fragility.

Altered Tissue Composition, Microarchitecture, and Mechanical Performance in Cancellous Bone From Men With Type 2 Diabetes Mellitus.

People with type 2 diabetes mellitus (T2DM) have normal-to-high BMDs, but, counterintuitively, have greater fracture risks than people without T2DM, even after accounting for potential confounders like BMI and falls. Therefore, T2DM may alter aspects of bone quality, including material properties or microarchitecture, that increase fragility independently of bone mass. Our objective was to elucidate the factors that influence fragility in T2DM by comparing the material properties, microarchitecture, and mechanical performance of cancellous bone in a clinical population of men with and without T2DM. Cancellous specimens from the femoral neck were collected during total hip arthroplasty (T2DM: n = 31, age = 65 ± 8 years, HbA1c = 7.1 ± 0.9%; non-DM: n = 34, age = 62 ± 9 years, HbA1c = 5.5 ± 0.4%). The T2DM specimens had greater concentrations of the advanced glycation endproduct pentosidine (+ 36%, P < 0.05) and sugars bound to the collagen matrix (+ 42%, P < 0.05) than the non-DM specimens. The T2DM specimens trended toward a greater bone volume fraction (BV/TV) (+ 24%, NS, P = 0.13) and had greater mineral content (+ 7%, P < 0.05) than the non-DM specimens. Regression modeling of the mechanical outcomes revealed competing effects of T2DM on bone mechanical behavior. The trend of higher BV/TV values and the greater mineral content observed in the T2DM specimens increased strength, whereas the greater values of pentosidine in the T2DM group decreased postyield strain and toughness. The long-term medical management and presence of osteoarthritis in these patients may influence these outcomes. Nevertheless, our data indicate a beneficial effect of T2DM on cancellous microarchitecture, but a deleterious effect of T2DM on the collagen matrix. These data suggest that high concentrations of advanced glycation endproducts can increase fragility by reducing the ability of bone to absorb energy before failure, especially for the subset of T2DM patients with low BV/TV. © 2019 American Society for Bone and Mineral Research.

Glycation of type I collagen selectively targets the same helical domain lysine sites as lysyl oxidase-mediated cross-linking.

Nonenzymatic glycation of collagen has long been associated with the progressive secondary complications of diabetes. How exactly such random glycations result in impaired tissues is still poorly understood. Because of the slow turnover rate of most fibrillar collagens, they are more susceptible to accumulate time-dependent glycations and subsequent advanced glycation end-products. The latter are believed to include cross-links that stiffen host tissues. However, diabetic animal models have also displayed weakened tendons with reduced stiffness. Strikingly, not a single experimentally identified specific molecular site of glycation in a collagen has been reported. Here, using targeted MS, we have identified partial fructosyl-hydroxylysine glycations at each of the helical domain cross-linking sites of type I collagen that are elevated in tissues from a diabetic mouse model. Glycation was not found at any other collagen lysine residues. Type I collagen in mouse tendons is cross-linked intermolecularly by acid-labile aldimine bonds formed by the addition of telopeptide lysine aldehydes to hydroxylysine residues at positions α1(I)Lys87, α1(I)Lys930, α2(I)Lys87, and α2(I)Lys933 of the triple helix. Our data reveal that site-specific glycations of these specific lysines may significantly impair normal lysyl oxidase-controlled cross-linking in diabetic tendons. We propose that such N-linked glycations can hinder the normal cross-linking process, thus altering the content and/or placement of mature cross-links with the potential to modify tissue material properties.

A new open-source tool for measuring 3D osteocyte lacunar geometries from confocal laser scanning microscopy reveals age-related changes to lacunar size and shape in cortical mouse bone.

Osteocytes can participate in systemic mineral homeostasis through perilacunar maintenance and remodeling, where changes to osteocyte lacunar morphology may affect bone structural integrity, tissue strains, and osteocyte mechanosensitivity. Though aging is associated with both decreased bone quality and altered mineral metabolism, it is not known if osteocyte lacunae undergo age-related changes in geometry. In order to survey lacunar changes with age, we developed an open-source program whereby 3D osteocyte lacunae are automatically segmented and then subsequently reconstructed from confocal laser scanning microscopy (CLSM) depth stacks for quantitative analysis of geometry and orientation. This approach takes advantage of the availability and speed of CLSM while avoiding time-consuming and bias-prone manual segmentation. Unlike conventional approaches used to quantify osteocyte lacunar morphology, CLSM enables facile analysis in three-dimensions with clear identification of osteocyte lacunae. We report that 3D osteocyte lacunae measured by CLSM become smaller, more spherical, more oblate, more spatially disorganized, and more sparsely populated with increased age in C57Bl/6 mouse cortical bone in groups spanning 6-24 months old. Critically, these age-related changes are in large part not observed in 2D analyses from the same samples. These results (1) demonstrate proof-of-concept of an efficient method to quantitatively assess osteocyte lacunae in 3D for application to a wide range of studies and (2) motivate further inquiry into how changes to osteocyte lacunar geometries and perilacunar material contribute to diminished bone quality in aging.

Bone Tissue Collagen Maturity and Mineral Content Increase With Sustained Hyperglycemia in the KK-Ay Murine Model of Type 2 Diabetes.

Type 2 diabetes mellitus (T2DM) increases fracture risk for a given bone mineral density (BMD), which suggests that T2DM changes bone tissue properties independently of bone mass. In this study, we assessed the effects of hyperglycemia on bone tissue compositional properties, enzymatic collagen crosslinks, and advanced glycation end-products (AGEs) in the KK-Ay murine model of T2DM using Fourier transform infrared (FTIR) imaging and high-performance liquid chromatography (HPLC). Compared to KK-aa littermate controls (n = 8), proximal femoral bone tissue of KK-Ay mice (n = 14) exhibited increased collagen maturity, increased mineral content, and less heterogeneous mineral properties. AGE accumulation assessed by the concentration of pentosidine, as well as the concentrations of the nonenzymatic crosslinks hydroxylysylpyridinoline (HP) and lysyl pyridinoline (LP), did not differ in the proximal femurs of KK-Ay mice compared to controls. The observed differences in tissue-level compositional properties in the KK-Ay mice are consistent with bone that is older and echo observations of reduced remodeling in T2DM. © 2017 American Society for Bone and Mineral Research.

The Effect of Diabetes Mellitus on Total Joint Arthroplasty Outcomes.

Diabetes has negative effects on the outcomes of total joint arthroplasty, including increased numbers of complications, decreased function, early revision, and higher costs. The prevalence of diabetes is increasing rapidly worldwide; therefore, the orthopaedic surgeon should have an understanding of how diabetes affects surgical outcomes.

Moderate chronic kidney disease impairs bone quality in C57Bl/6J mice.

Chronic kidney disease (CKD) increases bone fracture risk. While the causes of bone fragility in CKD are not clear, the disrupted mineral homeostasis inherent to CKD may cause material quality changes to bone tissue. In this study, 11-week-old male C57Bl/6J mice underwent either 5/6th nephrectomy (5/6 Nx) or sham surgeries. Mice were fed a normal chow diet and euthanized 11weeks post-surgery. Moderate CKD with high bone turnover was established in the 5/6 Nx group as determined through serum chemistry and bone gene expression assays. We compared nanoindentation modulus and mineral volume fraction (assessed through quantitative backscattered scanning electron microscopy) at matched sites in arrays placed on the cortical bone of the tibia mid-diaphysis. Trabecular and cortical bone microarchitecture and whole bone strength were also evaluated. We found that moderate CKD minimally affected bone microarchitecture and did not influence whole bone strength. Meanwhile, bone material quality decreased with CKD; a pattern of altered tissue maturation was observed with 5/6 Nx whereby the newest 60µm of bone tissue adjacent to the periosteal surface had lower indentation modulus and mineral volume fraction than more interior, older bone. The variance of modulus and mineral volume fraction was also altered following 5/6 Nx, implying that tissue-scale heterogeneity may be negatively affected by CKD. The observed lower bone material quality may play a role in the decreased fracture resistance that is clinically associated with human CKD.

In vivo model to measure bone repair efficacy of nanoparticle-based drug delivery.

Bone repair required for successful arthroplasty can be compromised in patients with comorbid conditions, such as osteoporosis, diabetes mellitus, and chronic kidney disease. Biological compounds have been proposed to promote bone health and repair. The authors have designed a new animal model for testing bone promoting compounds in the in vivo environment. For initial validation of this model, they used a synthetic agonist of a nuclear receptor, liver X receptor, which has been postulated to play a regulatory role in modulating bone growth. A distal femoral unicortical osteotomy was surgically created on skeletally mature C57Bl/6 male and female mice. A nanoparticle carrier delivery system was used to directly introduce N,N-dimethyl-3ß-hydroxycholenamide into the osteotomy. At 35 days post-procedure, the femora were harvested and specimens were obtained for histologic processing and qualitative analysis. The results indicate that the carrier nanoparticles entered the osteotomy defect. Results also indicate that bone repair occurred, although significant differences between groups were not detected in the current study. This study validates the mouse model for testing bone repair promoting compounds. This model can be combined with transgenic or other mouse models to simulate problematic bone repair environments, can be used with a variety of drug carriers, and can test many types of interventional compounds to evaluate potential orthopedic therapeutic applications.

Relationship of relaxin hormone and thumb carpometacarpal joint arthritis.

BACKGROUND: The female predominance in thumb carpometacarpal (CMC) joint arthritis has led to speculation that reproductive hormones or hypermobility are responsible. Evidence shows that patients with pathologic laxity have a higher rate of thumb CMC arthritis. Relaxin hormone increases laxity in the pelvic ligaments through upregulation of matrix metalloproteases (MMPs). It is thus a hormone of interest in the development of thumb CMC arthritis. QUESTIONS/PURPOSES: Our goals were to identify demographic and hormonal factors associated with joint laxity in patients with CMC arthritis and to evaluate the relationship among serum relaxin, relaxin receptors, and MMPs in the anterior oblique ligament (AOL) of the thumb. We hypothesized that serum relaxin was correlated with joint laxity as well as with relaxin receptors and MMPs in the AOL. METHODS: Forty-nine patients undergoing thumb CMC arthroplasty underwent laxity examination, blood draw, and AOL sampling. Ligaments were analyzed for relaxin receptor and MMPs 1 and 3 using quantitative reverse-transcriptase polymerase chain reaction. RESULTS: Women demonstrated more joint laxity than men (p < 0.001). RNA analysis confirmed relaxin receptors in the AOL as well as MMPs 1 and 3. There was a significant correlation between serum relaxin and MMP-1 (p = 0.04). Detectable serum relaxin was negatively correlated with relaxin receptors in the AOL (p = 0.02). CONCLUSIONS: Further studies are needed to evaluate the role of laxity and sex hormones in thumb CMC arthritis. CLINICAL RELEVANCE: Relaxin hormone may play a role in the development of arthritis at the thumb CMC joint. LEVEL OF EVIDENCE: Level I, prognostic study. See Guidelines for Authors for a complete description of levels of evidence.

Veterans with diabetes receive arthroplasty more frequently and at a younger age.

BACKGROUND: A future increase in total joint arthroplasties in patients with diabetes seems likely considering the prevalence of osteoarthritis and diabetes mellitus are increasing. However, the rates of arthroplasty in the population of patients with diabetes are unclear. QUESTIONS/PURPOSES: We sought to determine whether lower extremity arthroplasties in a veteran population with diabetes is different from a similar population without diabetes. The following specific questions were asked: (1) Is the rate of TKA in veterans with diabetes higher than in those without diabetes? (2) Is the rate of THA in veterans with diabetes higher than in those without diabetes? (3) Are arthroplasty revision rates greater in veterans with diabetes than in veterans without diabetes? METHODS: The US Department of Veterans Affairs Health administrative data from fiscal year 2000 was used to identify persons with primary or secondary TKA or THA. The rate of surgeries among a diabetic population was compared with that among a nondiabetic population. RESULTS: The diabetic cohort received total joint arthroplasties at a higher rate than the nondiabetic cohort at all ages younger than 66 years, with a range of odd ratios from 1.3 to 3.4. In answer to our specific questions, (1) the rate of TKA (95% CI, 2.1-3.7), (2) the rate of THA (95% CI, 1.0-2.6), and (3) the rates of arthroplasty revision (95% CI, 0.9-5.8 TKA and 0.7-6.8 THA) were higher in veterans with diabetes. Furthermore, those with diabetes in the youngest age group studied received total joint arthroplasties and revision surgeries at approximately double the rates of those without diabetes. CONCLUSIONS: If these findings hold true for the population as a whole, they imply that clinicians in the United States may see a sharp increase in younger diabetic candidates for joint arthroplasty.

Relationship of serum relaxin to generalized and trapezial-metacarpal joint laxity.

PURPOSE: The reproductive hormone relaxin acts to loosen pelvic ligaments in preparation for childbirth and is thought to be a mediator of joint laxity. The purpose of this study was to evaluate the correlation of serum relaxin with radiographic laxity at the trapezial-metacarpal joint and with generalized joint laxity. METHODS: We enrolled 289 healthy subjects prospectively. Participants completed a demographic questionnaire and were examined for generalized joint hypermobility using the Beighton-Horan scale. Stress radiographs of the trapezial-metacarpal joint were obtained in 163 subjects (56%). Blood samples were collected, and serum relaxin was measured for 287 subjects using enzyme-linked immunosorbent assay for human relaxin-2. RESULTS: The mean serum relaxin level among all subjects was 1.84 pg/mL (range, 0-45.25 pg/mL). Relaxin was not detectable in 166 of 287 samples, whereas the mean serum relaxin level among the 121 subjects with a detectable relaxin level (of 287 total relaxin samples) was 4.37 pg/mL (range, 0.46-45.25 pg/mL). Mean trapezial-metacarpal subluxation ratio scores were higher among those with a detectable relaxin level compared to those without a detectable relaxin level (0.34 vs 0.30 pg/mL). The average Beighton-Horan laxity score was 1.8 (range, 0-9). There was no correlation between generalized joint laxity measures and serum relaxin levels. CONCLUSIONS: In a large volunteer population, we demonstrated a relationship between circulating relaxin and trapezial-metacarpal joint laxity. However, we were unable to show a direct link between serum relaxin and generalized joint laxity. TYPE OF STUDY/LEVEL OF EVIDENCE: Prognostic II.

Acute repetitive lumbar syndrome: a multi-component insight into the disorder.

PURPOSE: Repetitive Lumbar Injury (RLI) is common in individuals engaged in long term performance of repetitive occupational/sports activities with the spine. The triggering source of the disorder, tissues involved in the failure and biomechanical, neuromuscular, and biological processes active in the initiation and development of the disorder, are not known. The purpose is, therefore, to test, using in-vivo feline model and healthy human subjects, the hypothesis that RLI due to prolonged exposure to repetitive lumbar flexion-extension is triggered by an acute inflammation in the viscoelastic tissues and is characterized by lingering residual creep, pronounced changes in neuromuscular control and transient changes in lumbar stability. This report, therefore, is a summary of a lengthy research program consisting of multiple projects. METHODS: A series of experimental data was obtained from in-vivo feline groups and normal humans subjected to prolonged cyclic lumbar flexion-extension at high and low loads, high and low velocities, few and many repetitions, as well as short and long in-between rest periods, while recording lumbar displacement and multifidi EMG. Neutrophil and cytokines expression analysis were performed on the dissected feline supraspinous ligaments before loading (control) and 7 h post-loading. A comprehensive, time based model was designed to represent the creep, motor control, tissue biology and stability derived from the experimental data. RESULTS: Prolonged cyclic loading induced creep in the spine, reduced muscular activity, triggered spasms and reduced stability followed, several hours later, by acute inflammation/tissue degradation, muscular hyperexcitability and hyperstability. Fast movement, high loads, many repetitions and short rest periods, triggered the full disorder, whereas low velocities, low loads, long rest and few repetitions, triggered only minor but statistically significant pro-inflammatory tissue degradation and significantly reduced stability. CONCLUSION: Viscoelastic tissue failure via inflammation is the source of RLI and is also the process which governs the mechanical and neuromuscular characteristic symptoms of the disorder. The experimental data validates the hypothesis and provides insights into the development of potential treatments and prevention.

Osteoblasts derived from induced pluripotent stem cells form calcified structures in scaffolds both in vitro and in vivo.

Reprogramming somatic cells into an ESC-like state, or induced pluripotent stem (iPS) cells, has emerged as a promising new venue for customized cell therapies. In this study, we performed directed differentiation to assess the ability of murine iPS cells to differentiate into bone, cartilage, and fat in vitro and to maintain an osteoblast phenotype on a scaffold in vitro and in vivo. Embryoid bodies derived from murine iPS cells were cultured in differentiation medium for 8­12 weeks. Differentiation was assessed by lineage-specific morphology, gene expression, histological stain, and immunostaining to detect matrix deposition. After 12 weeks of expansion, iPS-derived osteoblasts were seeded in a gelfoam matrix followed by subcutaneous implantation in syngenic imprinting control region (ICR) mice. Implants were harvested at 12 weeks, histological analyses of cell and mineral and matrix content were performed. Differentiation of iPS cells into mesenchymal lineages of bone, cartilage, and fat was confirmed by morphology and expression of lineage-specific genes. Isolated implants of iPS cell-derived osteoblasts expressed matrices characteristic of bone, including osteocalcin and bone sialoprotein. Implants were also stained with alizarin red and von Kossa, demonstrating mineralization and persistence of an osteoblast phenotype. Recruitment of vasculature and microvascularization of the implant was also detected. Taken together, these data demonstrate functional osteoblast differentiation from iPS cells both in vitro and in vivo and reveal a source of cells, which merit evaluation for their potential uses in orthopedic medicine and understanding of molecular mechanisms of orthopedic disease.

Arthroplasty in veterans: analysis of cartilage, bone, serum, and synovial fluid reveals differences and similarities in osteoarthritis with and without comorbid diabetes.

Osteoarthritis patients with diabetes who receive total knee arthroplasty are more vulnerable to complications, including aseptic loosening and need for revision surgery. To elucidate mechanisms related to arthroplasty failure in diabetes, we examined serum and synovial fluid markers as well as collagen crosslinks in bone and cartilage of 20 patients (10 with diabetes, 10 controls without) undergoing this procedure. Hemoglobin A1c, body mass index, bone alkaline phosphatase, leptin, osteocalcin, and pyridinium were analyzed along with tissue content of the crosslinks hydroxylysylpyridinoline, lysylpyridinoline, and pentosidine. Pentosidine levels in tissue specimens from diabetic subjects were higher than in control subjects. Osteocalcin levels negatively correlated with hydroxylysylpyridinoline levels in cartilage. Osteocalcin levels also negatively correlated with pentosidine levels in cartilage, but only in subjects with diabetes. This study suggests potential metabolic mechanisms for arthroplasty failure in patients with diabetes.

High-frequency loading of lumbar ligaments increases proinflammatory cytokines expression in a feline model of repetitive musculoskeletal disorder.

BACKGROUND CONTEXT: Cumulative (repetitive) lumbar disorder is common in the workforce, and the associated epidemiology points out high risk for lifting heavy loads, performing many repetitions, and performing movements at high velocity. Experimental verification of viscoelastic tissue degradation and a neuromuscular disorder exist for cyclic work under heavy loads. Experimental validation for a disorder because of cyclic loads under high-velocity movement is missing. PURPOSE: Obtain experimental verification that high-velocity lumbar flexion-extension results in significant increase of proinflammatory cytokines in the viscoelastic tissues. STUDY DESIGN: Laboratory experiments using two in vivo feline model groups subjected to cyclic flexion-extension at low and high velocity. METHODS: Seven hours after cumulative 60 minutes of cyclic flexion-extension at moderate load of 40 N and 0.25 Hz for first group and 0.5 Hz for the second group, the supraspinous ligaments of L3-L4 to L5-L6 were harvested and subjected to cytokines (interleukin [IL]-1ß, IL-6, IL-8, tumor necrosis factor-α, and transforming growth factor-ß1) analysis. Two-way mixed model analysis of variance with a post hoc analysis were used to assess any significant differences (p<.05) in cytokines expression level between the two groups as well as main effect and interaction with lumbar levels. RESULTS: Expression levels of the five cytokines were significantly increased in the group subjected to the high-frequency loading. CONCLUSIONS: Exposure of the lumbar spine to high-velocity flexion-extension triggers a significant increase in proinflammatory cytokines, indicating pronounced changes consistent with an acute inflammation. Further exposure to activity over prolonged periods may trigger chronic inflammation and tissue degeneration as the source of cumulative lumbar disorder.

Change in serum COMP concentration due to ambulatory load is not related to knee OA status.

The aim of this study was to test the hypothesis that a change in serum cartilage oligomeric matrix protein (COMP) concentration is related to joint load during a 30-min walking exercise in patients with medial compartment knee osteoarthritis (OA) and in age-matched control subjects. Blood samples were drawn from 42 patients with medial compartment knee OA and from 41 healthy age-matched control subjects immediately before, immediately after, and 0.5, 1.5, 3.5, and 5.5 h after a 30-min walking exercise on a level outdoor walking track at self-selected normal speed. Serum COMP concentrations were determined using a commercial ELISA. Basic time-distance gait variables were recorded using an activity monitor. Joint loads were measured using gait analysis. Serum COMP concentrations increased immediately after the walking exercise (+6.3% and +5.6%; p < 0.001) and decreased over 5.5 h after the exercise (-11.1% and -14.6%; p < 0.040 and p = 0.001) in patients and control subjects, respectively. The magnitude of increase in COMP concentration did not differ between groups (p = 0.902) and did not correlate with any variables describing ambulatory loads at the joints of the lower extremity. These results, taken together with a previous study of a younger healthy population, suggest the possibility that the influence of ambulatory loads on cartilage turnover is dependent on age.

Evaluation of gene expression through qRT-PCR in cyclically loaded tendons: an in vivo model.

An in vivo rabbit animal model for the tendinopathy, epicondylitis, was used to examine the effects of repetitive load on the expression of various genes associated with matrix remodeling. Following 80 h of cumulative load, tissue from the distal and proximal regions of the flexor digitorum profundus tendon was collected. Quantitative RT-PCR was used to asses mRNA levels of collagenase-1 (MMP-1), stromelysin (MMP-3), vascular endothelial growth factor (VEGF), connective tissue growth factor (CTGF), cyclooxygenase-2 (COX-2), interleukin-1beta (IL-1beta), type III collagen (COL-III) and fibronectin (FBRN). No significant differences in expression levels were found between loaded and unloaded limbs at either region of the tendon. The findings were unexpected as the same model has already demonstrated an increase in the density of cells staining for VEGF and CTGF. Different regulatory mechanisms between mRNA and protein expression or localized changes missed due to homogenization of the tissue samples, may explain the discrepancy in findings.

Effect of repetition rate on the formation of microtears in tendon in an in vivo cyclical loading model.

We reported previously the formation of microtears in an in vivo loaded Flexor Digitorum Profundus (FDP) rabbit tendon with a repetition rate of 60 repetitions per minute and a peak force of 15% of maximum peak tetanic force for 80 cumulative hours. Tear area as a percent of tendon area, tear density (tears/mm(2)), and mean tear size (microm(2)) were higher in tendons from the loaded limb compared to the unloaded control limb. The purpose of the present study was to compare those results to results obtained with a repetition rate of 10 while maintaining the same peak force and force-time integral (n = 8). Due to a strain gradient between the inner and outer sides of the FDP tendon, microtears were quantified in four regions, two regions each along the inner and outer sides of the tendon. The tear area as a percent of total tendon area and the mean tear size were significantly greater in the loaded limb compared to the unloaded limb (p < 0.03). However, the effects were less than those observed at 60 repetitions/min. The higher repetition loading pattern resulted in an increase in tear measures in all four regions, while the lower rate produced changes only in the outer regions of the tendon. This finding may establish where the initial sites of damage occur in tendons that insert into bone in a similar arrangement as the FDP. The results suggest that repetition rate or number of loading cycles is associated with increased tendon microtears or fragility in a dose-response pattern.