Ashleigh M. Philp

Long Intergenic Noncoding RNAs Mediate the Human Chondrocyte Inflammatory Response and Are Differentially Expressed in Osteoarthritis Cartilage

To identify long noncoding RNAs (lncRNAs), including long intergenic noncoding RNAs (lincRNAs), antisense RNAs, and pseudogenes, associated with the inflammatory response in human primary osteoarthritis (OA) chondrocytes and to explore their expression and function in OA.

Developing anti-inflammatory therapeutics for patients with osteoarthritis

OA is the most common joint disorder in the world, but there are no approved therapeutics to prevent disease progression. Historically, OA has been considered a wear-and-tear joint disease, and efforts to identify and develop disease-modifying therapeutics have predominantly focused on direct inhibition of cartilage degeneration. However, there is now increasing evidence that inflammation is a key mediator of OA joint pathology, and also that the link between obesity and OA is not solely due to excessive load-bearing, suggesting therefore that targeting inflammation in OA could be a rewarding therapeutic strategy. In this review we therefore re-evaluate historical clinical trial data on anti-inflammatory therapeutics in OA patients, highlight some of the more promising emerging therapeutic targets and discuss the implications for future clinical trial design.

IL-6 secretion in osteoarthritis patients is mediated by chondrocyte-synovial fibroblast cross-talk and is enhanced by obesity

Increasing evidence suggests that inflammation plays a central role in driving joint pathology in certain patients with osteoarthritis (OA). Since many patients with OA are obese and increased adiposity is associated with chronic inflammation, we investigated whether obese patients with hip OA exhibited differential pro-inflammatory cytokine signalling and peripheral and local lymphocyte populations, compared to normal weight hip OA patients. No differences in either peripheral blood or local lymphocyte populations were found between obese and normal-weight hip OA patients. However, synovial fibroblasts from obese OA patients were found to secrete greater amounts of the pro-inflammatory cytokine IL-6, compared to those from normal-weight patients (p < 0.05), which reflected the greater levels of IL-6 detected in the synovial fluid of the obese OA patients. Investigation into the inflammatory mechanism demonstrated that IL-6 secretion from synovial fibroblasts was induced by chondrocyte-derived IL-6. Furthermore, this IL-6 inflammatory response, mediated by chondrocyte-synovial fibroblast cross-talk, was enhanced by the obesity-related adipokine leptin. This study suggests that obesity enhances the cross-talk between chondrocytes and synovial fibroblasts via raised levels of the pro-inflammatory adipokine leptin, leading to greater production of IL-6 in OA patients.

The Effect of Vancomycin and Gentamicin Antibiotics on Human Osteoblast Proliferation, Metabolic Function, and Bone Mineralization.

Study Design. The present study investigates the effect of vancomycin and gentamicin antibiotics on primary human osteoblasts. Osteoblasts were incubated with vancomycin, gentamicin, or with povidone-iodine (PVI), at concentrations advocated for wound irrigation. Osteoblast proliferation, metabolic function, and bone mineralization were measured. Objective. The aim of the study was to model gentamicin and vancomycin wound irrigation in vitro and to examine the effect on osteoblast viability and cellular function in comparison to 0.35% PVI. Summary of Background Data. Vancomycin, gentamicin, and dilute PVI are employed as wound irrigants in spinal surgery to reduce infection. We have, however, recently demonstrated that 0.35% PVI has a detrimental effect on osteoblast cellular function and bone mineralization. Studies to determine the effects of antibiotic wound irrigation solutions on osteoblasts and bone mineralization are therefore warranted. Methods. Primary human osteoblasts were exposed for 20 minutes to phosphate buffered saline (PBS) control, vancomycin (35 or 3.5 mmol/L), gentamicin (34 or 3.4 mmol/L), or 0.35% PVI for 3 minutes. Cellular proliferation was measured during 7 days by MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) assay. Osteoblast metabolic function was determined using a Seahorse XFe24 Bioanalyzer. Mineralized bone nodules were quantified using Alizarin red. Results. At concentrations advocated for wound irrigation, both gentamicin (3.4 mmol/L) and vancomycin (3.5 mmol/L) induced a transient 15% to 20% reduction in osteoblast proliferation, which returned to control values within 72 hours. This was in marked contrast to the effect of 0.35% PVI, which resulted in a sustained reduction in osteoblast proliferation of between 40% and 50% during 7 days. Neither gentamicin nor vancomycin at concentrations up to 10× clinical dose had any effect on osteoblast oxygen consumption rate, or significantly affected mineralized bone nodule formation. Conclusion. Vancomycin and gentamicin solutions, at concentrations advocated for intrawound application in spinal surgery, have a small but transient effect on osteoblast proliferation, and no effect on either osteoblast metabolic function or bone nodule mineralization. Level of Evidence: N/A

Povidone-Iodine Has a Profound Effect on In Vitro Osteoblast Proliferation and Metabolic Function and Inhibits Their Ability to Mineralize and Form Bone.

Study Design. A study examining the clinical protocol of scoliosis wound irrigation, demonstrating povidone-iodines (PVI) effect on human osteoblast cells. Primary and immortal cell line osteoblasts were treated with 0.35% PVI for 3 minutes, and analyzed for proliferation rate, oxidative capacity, and mineralization. Objective. To model spinal wound irrigation with dilute PVI in vitro, in order to investigate the effect of PVI on osteoblast proliferation, metabolism, and bone mineralization. Summary of Background Data. Previously PVI irrigation has been proposed as a safe and effective practice to avoid bacterial growth after spinal surgery. However, recent evidence in multiple cell types suggests that PVI has a deleterious effect on cellular viability and cellular function. Methods. Primary and immortal human osteoblast cells were exposed to either phosphate buffered saline control or with 0.35% PVI for 3 minutes. Cellular proliferation was measured over the duration of 7 days by MTS assay. Oxygen consumption rate, extracellular acidification rate, and proton production rate were analyzed using a Seahorse XFe24 Bioanalyzer. Protein expression of the electron transport chain subunits CII-SDHB, CIII-UQRCR2, and CV-ATP5A was measured via Western blotting. Mineralized bone nodules were stained with alizarin red. Results. Expressed as a percentage of normal osteoblast proliferation, osteoblasts exposed to 0.35% PVI exhibited a significant 24% decrease in proliferation after 24 hours. This was a sustained response, resulting in a 72% decline in cellular proliferation at 1 week. There was a significant reduction in oxygen consumption rate, extracellular acidification rate, and proton production rate (P < 0.05), in osteoblasts that had been exposed to 0.35% PVI for 3 minutes, coupled with a marked reduction in the protein expression of CII-SDHB. Osteoblasts exposed to 0.35% PVI exhibited reduced bone nodule mineralization compared to control phosphate buffered saline exposed osteoblasts (P < 0.01). Conclusion. PVI has a rapid and detrimental effect on human osteoblast cellular proliferation, metabolic function, and bone nodule mineralization. Level of Evidence: NA