Dianne Little

Acute joint pathology and synovial inflammation is associated with increased intra-articular fracture severity in the mouse knee

OBJECTIVE Post-traumatic arthritis is a frequent cause of disability and occurs most commonly and predictably after articular fracture. The objective of this investigation was to examine the effect of fracture severity on acute joint pathology in a novel murine model of intra-articular fracture. DESIGN Low and high energy articular fractures (n=25 per group) of the tibial plateau were created in adult male C57BL/6 mice. The acute effect of articular fracture severity on synovial inflammation, bone morphology, liberated fracture area, cartilage pathology, chondrocyte viability, and systemic cytokines and biomarkers levels was assessed at 0, 1, 3, 5, and 7 days post-fracture. RESULTS Increasing intra-articular fracture severity was associated with greater acute pathology in the synovium and bone compared to control limbs, including increased global synovitis and reduced periarticular bone density and thickness. Applied fracture energy was significantly correlated with degree of liberated cortical bone surface area, indicating greater comminution. Serum concentrations of hyaluronic acid (HA) were significantly increased 1 day post-fracture. While articular fracture significantly reduced chondrocyte viability, there was no relationship between fracture severity and chondrocyte viability, cartilage degeneration, or systemic levels of cytokines and biomarkers. CONCLUSIONS This study demonstrates that articular fracture is associated with a loss of chondrocyte viability and increased levels of systemic biomarkers, and that increased intra-articular fracture severity is associated with increased acute joint pathology in a variety of joint tissues, including synovial inflammation, cortical comminution, and bone morphology. Further characterization of the early events following articular fracture could aid in the treatment of post-traumatic arthritis.

Electrospun cartilage-derived matrix scaffolds for cartilage tissue engineering.

Macroscale scaffolds created from cartilage-derived matrix (CDM) demonstrate chondroinductive or chondro-inductive properties, but many fabrication methods do not allow for control of nanoscale architecture. In this regard, electrospun scaffolds have shown significant promise for cartilage tissue engineering. However, nanofibrous materials generally exhibit a relatively small pore size and require techniques such as multilayering or the inclusion of sacrificial fibers to enhance cellular infiltration. The objectives of this study were (1) to compare multilayer to single-layer electrospun poly(ɛ-caprolactone) (PCL) scaffolds for cartilage tissue engineering, and (2) to determine whether incorporation of CDM into the PCL fibers would enhance chondrogenesis by human adipose-derived stem cells (hASCs). PCL and PCL-CDM scaffolds were prepared by sequential collection of 60 electrospun layers from the surface of a grounded saline bath into a single scaffold, or by continuous electrospinning onto the surface of a grounded saline bath and harvest as a single-layer scaffold. Scaffolds were seeded with hASCs and evaluated over 28 days in culture. The predominant effects on hASCs of incorporation of CDM into scaffolds were to stimulate sulfated glycosaminoglycan synthesis and COL10A1 gene expression. Compared with single-layer scaffolds, multilayer scaffolds enhanced cell infiltration and ACAN gene expression. However, compared with single-layer constructs, multilayer PCL constructs had a much lower elastic modulus, and PCL-CDM constructs had an elastic modulus approximately 1% that of PCL constructs. These data suggest that multilayer electrospun constructs enhance homogeneous cell seeding, and that the inclusion of CDM stimulates chondrogenesis-related bioactivity.

Management of drug-resistant cyathostominosis on a breeding farm in central North Carolina.

REASONS FOR PERFORMING STUDY Possible anthelmintic resistance on a breeding farm where a rapid rotation anthelmintic programme had been implemented for 9 years was investigated. Cyathostomins resistant to fenbendazole and pyrantel were documented by faecal worm egg count reduction test (FWECRT). OBJECTIVES To 1) manage small strongyle transmission in a herd of horses in which resistance to both pyrantel pamoate and fenbendazole was identified and thereby reduce the risk of clinical disease in the individual animal, 2) monitor the change in resistance patterns over time and 3) monitor the efficacy of ivermectin over the study period. METHODS Targeted ivermectin treatment of horses on the farm was instituted for mature horses with faecal worm egg counts (FWEC) > 200 eggs/g (epg) and for horses < age 2 years with FWEC > 100 epg. RESULTS Over a 30 month period, targeted ivermectin treatment achieved acceptable control in mares, as judged by FWEC, and improved control of patent cyathostome infection in consecutive foal crops. Egg reappearance time (ERT) after treatment with ivermectin was < 8 weeks in mares and foals more frequently in the second year of the study than in the first year. Numbers of anthelmintic treatments were reduced by 77.6 and 533% in the mare and foal group, respectively. CONCLUSIONS Targeted ivermectin treatment may be an economically viable method of managing multiple drug resistant cyathostominosis. POTENTIAL RELEVANCE Use of ivermectin should be monitored closely for development of resistance.

Factors associated with development of ileal impaction in horses with surgical colic: 78 cases (1986-2000)

Deal impaction is prevalent in the south-eastern USA, where feeding of Coastal Bermuda hay has been implicated as a risk factor. Alternatively, infection with the tapeworm Anoplocephala perfoliata has been identified as a risk factor for ileal impaction in the UK. We hypothesised that feeding Coastal Bermuda hay and failure to administer routinely an anthelmintic with efficacy against tapeworms would place horses at risk of developing ileal impaction in the USA. Seventy-eight horses, with surgically confirmed ileal impaction and 100 horses admitted for colic that did not have an ileal impaction, were selected retrospectively for logistic regression analysis. Using odds ratios (OR) as an index of risk, feeding Coastal Bermuda hay (OR = 2.9) and failure to administer a pyrantel salt within 3 months of admission (OR = 3.1) placed horses at risk of development of ileal impaction. This study confirms the belief that feeding Coastal Bermuda hay places horses at risk of ileal impaction, although the quality of the hay may also play a role. Periodic administration of anthelmintics with efficacy against tapeworms should be considered to reduce risk of ileal impaction.

Dietary fatty acid content regulates wound repair and the pathogenesis of osteoarthritis following joint injury

Objective The mechanisms linking obesity and osteoarthritis (OA) are not fully understood and have been generally attributed to increased weight, rather than metabolic or inflammatory factors. Here, we examined the influence of fatty acids, adipokines, and body weight on OA following joint injury in an obese mouse model. Methods Mice were fed high-fat diets rich in various fatty acids (FA) including saturated FAs (SFAs), ω-6 polyunsaturated FAs (PUFAs), and ω-3 PUFAs. OA was induced by destabilising the medial meniscus. Wound healing was evaluated using an ear punch. OA, synovitis and wound healing were determined histologically, while bone changes were measured using microCT. Activity levels and serum cytokines were measured at various time-points. Multivariate models were performed to elucidate the associations of dietary, metabolic and mechanical factors with OA and wound healing. Results Using weight-matched mice and multivariate models, we found that OA was significantly associated with dietary fatty acid content and serum adipokine levels, but not with body weight. Furthermore, spontaneous activity of the mice was independent of OA development. Small amounts of ω-3 PUFAs (8% by kcal) in a high-fat diet were sufficient to mitigate injury-induced OA, decreasing leptin and resistin levels. ω-3 PUFAs significantly enhanced wound repair, SFAs or ω-6 PUFAs independently increased OA severity, heterotopic ossification and scar tissue formation. Conclusions Our results indicate that with obesity, dietary FA content regulates wound healing and OA severity following joint injury, independent of body weight, supporting the need for further studies of dietary FA supplements as a potential therapeutic approach for OA.

Stem cell therapies for knee cartilage repair: the current status of preclinical and clinical studies.

Background: Articular cartilage damage of the knee is common, causing significant morbidity worldwide. Many adult tissues contain cells that are able to differentiate into multiple cell types, including chondrocytes. These stem cells have gained significant attention over the past decade and may become frontline management for cartilage defects in the very near future. Purpose: The role of stem cells in the treatment of knee osteochondral defects was reviewed. Recent animal and clinical studies were reviewed to determine the benefits and potential outcomes of using stem cells for cartilage defects. Study Design: Literature review. Methods: A PubMed search was undertaken. The key phrase “stem cells and knee” was used. The search included reviews and original articles over an unlimited time period. From this search, articles outlining animal and clinical trials were selected. A search of current clinical trials in progress was performed on the clinicaltrials.gov website, and “stem cells and knee” was used as the search phrase. Results: Stem cells have been used in many recent in vitro and animal studies. A number of cell-based approaches for cartilage repair have progressed from preclinical animal studies into clinical trials. Conclusion: The use of stem cells for the treatment of cartilage defects is increasing in animal and clinical studies. Methods of delivery of stem cells to the knee’s cartilage vary from direct injection to implantation with scaffolds. While these approaches are highly promising, there is currently limited evidence of a direct clinical benefit, and further research is required to assess the overall outcome of stem cell therapies for knee cartilage repair.

Aligned multilayered electrospun scaffolds for rotator cuff tendon tissue engineering.

UNLABELLED The rotator cuff consists of several tendons and muscles that provide stability and force transmission in the shoulder joint. Whereas most rotator cuff tears are amenable to suture repair, the overall success rate of repair is low, and massive tears are prone to re-tear. Extracellular matrix (ECM) patches are used to augment suture repair, but they have limitations. Tissue-engineered approaches provide a promising solution for massive rotator cuff tears. Previous studies have shown that, compared to nonaligned scaffolds, aligned electrospun polymer scaffolds exhibit greater anisotropy and exert a greater tenogenic effect. Nevertheless, achieving rapid cell infiltration through the full thickness of the scaffold is challenging, and scaling to a translationally relevant size may be difficult. Our goal was to evaluate whether a novel method of alignment, combining a multilayered electrospinning technique with a hybrid of several electrospinning alignment techniques, would permit cell infiltration and collagen deposition through the thickness of poly(ε-caprolactone) scaffolds following seeding with human adipose-derived stem cells. Furthermore, we evaluated whether multilayered aligned scaffolds enhanced collagen alignment, tendon-related gene expression, and mechanical properties compared to multilayered nonaligned scaffolds. Both aligned and nonaligned multilayered scaffolds demonstrated cell infiltration and ECM deposition through the full thickness of the scaffold after only 28days of culture. Aligned scaffolds displayed significantly increased expression of tenomodulin compared to nonaligned scaffolds and exhibited aligned collagen fibrils throughout the full thickness, the presence of which may account for the increased yield stress and Youngs modulus of cell-seeded aligned scaffolds along the axis of fiber alignment. STATEMENT OF SIGNIFICANCE Rotator cuff tears are an important clinical problem in the shoulder, with over 300,000 surgical repairs performed annually. Re-tear rates may be high, and current methods used to augment surgical repair have limited evidence to support their clinical use due to inadequate initial mechanical properties and slow cellular infiltration. Tissue engineering approaches such as electrospinning have shown similar challenges in previous studies. In this study, a novel technique to align electrospun fibers while using a multilayered approach demonstrated increased mechanical properties and development of aligned collagen through the full thickness of the scaffolds compared to nonaligned multilayered scaffolds, and both types of scaffolds demonstrated rapid cell infiltration through the full thickness of the scaffold.

Ligament-derived matrix stimulates a ligamentous phenotype in human adipose-derived stem cells.

Human adipose stem cells (hASCs) can differentiate into a variety of phenotypes. Native extracellular matrix (e.g., demineralized bone matrix or small intestinal submucosa) can influence the growth and differentiation of stem cells. The hypothesis of this study was that a novel ligament-derived matrix (LDM) would enhance expression of a ligamentous phenotype in hASCs compared to collagen gel alone. LDM prepared using phosphate-buffered saline or 0.1% peracetic acid was mixed with collagen gel (COL) and was evaluated for its ability to induce proliferation, differentiation, and extracellular matrix synthesis in hASCs over 28 days in culture at different seeding densities (0, 0.25 x 10(6), 1 x 10(6), or 2 x 10(6) hASC/mL). Biochemical and gene expression data were analyzed using analysis of variance. Fishers least significant difference test was used to determine differences between treatments following analysis of variance. hASCs in either LDM or COL demonstrated changes in gene expression consistent with ligament development. hASCs cultured with LDM demonstrated more dsDNA content, sulfated-glycosaminoglycan accumulation, and type I and III collagen synthesis, and released more sulfated-glycosaminoglycan and collagen into the medium compared to hASCs in COL (p <or= 0.05). Increased seeding density increased DNA content incrementally over 28 days in culture for LDM but not COL constructs (p <or= 0.05). These findings suggest that LDM can stimulate a ligament phenotype by hASCs, and may provide a novel scaffold material for ligament engineering applications.

Recovery of ischaemic injured porcine ileum: evidence for a contributory role of COX-1 and COX-2

Background: We have previously shown that the non-selective cyclooxygenase (COX) inhibitor indomethacin retards recovery of intestinal barrier function in ischaemic injured porcine ileum. However, the relative role of COX-1 and COX-2 elaborated prostaglandins in this process is unclear. Aims: To assess the role of COX-1 and COX-2 elaborated prostaglandins in the recovery of intestinal barrier function by evaluating the effects of selective COX-1 and COX-2 inhibitors on mucosal recovery and eicosanoid production. Methods: Porcine ileal mucosa subjected to 45 minutes of ischaemia was mounted in Ussing chambers, and transepithelial electrical resistance was used as an indicator of mucosal recovery. Prostaglandins E1 and E2 (PGE) and 6-keto-PGF1α (the stable metabolite of prostaglandin I2 (PGI2)) were measured using ELISA. Thromboxane B2 (TXB2, the stable metabolite of TXA2) was measured as a likely indicator of COX-1 activity. Results: Ischaemic injured tissues recovered to control levels of resistance within three hours whereas tissues treated with indomethacin (5×10−6 M) failed to fully recover, associated with inhibition of eicosanoid production. Injured tissues treated with the selective COX-1 inhibitor SC-560 (5×10−6 M) or the COX-2 inhibitor NS-398 (5×10−6 M) recovered to control levels of resistance within three hours, associated with significant elevations of PGE and 6-keto-PGF1α compared with untreated tissues. However, SC-560 significantly inhibited TXB2 production whereas NS-398 had no effect on this eicosanoid, indicating differential actions of these inhibitors related to their COX selectivity. Conclusions: The results suggest that recovery of resistance is triggered by PGE and PGI2, which may be elaborated by either COX-1 or COX-2.

Tendon mechanobiology: Current knowledge and future research opportunities

Tendons mainly function as load‐bearing tissues in the muscloskeletal system; transmitting loads from muscle to bone. Tendons are dynamic structures that respond to the magnitude, direction, frequency, and duration of physiologic as well as pathologic mechanical loads via complex interactions between cellular pathways and the highly specialized extracellular matrix. This paper reviews the evolution and current knowledge of mechanobiology in tendon development, homeostasis, disease, and repair. In addition, we review several novel mechanotransduction pathways that have been identified recently in other tissues and cell types, providing potential research opportunities in the field of tendon mechanobiology. We also highlight current methods, models, and technologies being used in a wide variety of mechanobiology research that could be investigated in the context of their potential applicability for answering some of the fundamental unanswered questions in this field. The article concludes with a review of the major questions and future goals discussed during the recent ORS/ISMMS New Frontiers in Tendon Research Conference held on September 10 and 11, 2014 in New York City.