The effect of conditional macrophage depletion on development of injury-induced knee OA in obese transgenic MAFIA mice

Abstract

s / Osteoarthritis and Cartilage 23 (2015) A82eA416 A287 Conclusions: Contrary to our hypothesis, we found that systemic MF depletion in obese MAFIA mice did not mitigate cartilage degradation after joint injury; instead, it enhanced joint synovitis by increasing infiltration of CD3þ T cells and neutrophils into the operated joint. Our study is significant for elucidating the immunomodulatory activity of MFs in inflammation and joint injury in obesity. 451 MECHANICALLY AIDED TRANSPORT OF ANTIBODIES THROUGH ARTICULAR CARTILAGE C. DiDomenico, Z. Wang, L. Bonassar. Cornell Univ., Ithaca, NY, USA Purpose: Recently, significant advances in the clinical treatment of rheumatoid arthritis (RA) have been made using therapeutic antibodies that target and inhibit tumor necrosis factor alpha (TNF-a) found in the synovium of the joint. Additionally, TNF-a is also known to play a critical role in the progression of osteoarthritis (OA); however, TNF-a blocking antibodies are not currently used for OA therapy. Besides total joint replacement, there is no long-term effective treatment for OA. In contrast to RA, in which synovial inflammation is easily targeted by intravenous injection of therapeutics, TNF-a activity in OA occurs primarily within articular cartilage. Because of the high density of the cartilage extracellular matrix, it is not clear whether large proteins, like antibodies, will be able to sufficiently penetrate the cartilage tissue on a time scale that is clinically relevant. Previously, mechanical stimulation of cartilage tissue has been shown to aid in the penetration of proteins ( 100 kDa). The objective of this study is to characterize the effect of physiologically relevant mechanical loading on the transport of antibodies through cartilage. Methods: For each experiment, cylindrical articular cartilage plugs (4 mm diameter, 1.15 mm thick) were extracted from 1-3 day old bovids and randomly assigned to loaded and unloaded groups. Fluorescently labeled antibodies (MW: 150 kDa) in PBS were added to each groupspecific well plates while only PBS were added to negative controls. By compressing each sample 15% with an impermeable platen array, only the outside cylindrical (radial) surface was exposed to the antibody solution (Figure 1 inset). Each group is then placed into a 37 C incubator where only the loaded group undergo unconfined cyclic axial compressionwith a custom-built, displacement-controlled compressor. The transducer then applies a set dynamic strain to the loaded samples via the platen array for 3 hours at a set frequency. After loading, all plugs were bisected axially and the cut surface was viewed using confocal microscopy to characterize antibody penetration from the radial edges of the cut surface towards the center (Figure 1 inset). Average fluorescence intensity profiles, from each radial edge of the cut surface to the center, were fitted to a 1D diffusion model derived from Fick's 2nd law to obtain an average effective diffusivity for each sample (Figure 1). The diffusivities of loaded samples were then normalized to the average diffusivity of the unloaded samples for each experiment. These “transport enhancement ratios” were then analyzed across several loading amplitudes applied a 1 Hz (Figure 2). Results: Experiments showed that cyclic compression enhances transport of antibodies into articular cartilage. The 1D diffusion model accurately described the fluorescence profiles of both the unloaded and loaded samples (coefficient of variation between 5-20% and R2 between 0.980 and 0.995). For the unloaded samples, diffusivities ranged from 2.8 to 6.0 10-8 cm2/s, while loaded diffusivities ranged from 6.8 to 15.4 10-8 cm2/s. The convective enhancement ratios obtained ranged from 1.20 for the 0.25% strain (N.S. compared to an enhancement ratio of 1, i.e., no enhancement), 1.47 for 1.3% strain (p < 0.01), 1.80 for 2.5% strain (p < 0.001), and 2.61 for 5% strain (p < 0.001) (Figure 2). Further, for this range in dynamic strain amplitude there was linear trend of increasing effective diffusivity with strain amplitude (r2 1⁄4 0.98, p < 0.005). Conclusions: Overall, despite the small pore size of cartilage and the high MW of the antibody used, mechanical loading enhanced antibody transport to a large degree. More research will have to be done to determine how well antibodies diffuse across the articular surface and how other antibodies affect transport in more mature cartilage tissue. Given these results, there is promise that antibody therapy for OA could be enhanced by an exercise regimen or physical therapy, mimicking physiological loading conditions from these experiments. To our knowledge, this is the first study to test how mechanical stimulation affects the diffusion of antibodies in cartilage and suggest further study into how more efficient antibody therapy for OA can be developed. Figure 1. Normalized fluorescent intensity vs. radial depth for a 5% dynamic strain loaded sample (blue) and unloaded (red) sample to which ID diffusion models were fit (solid lines). (Inset): Experimental setup of an individual well and an example of a confocal radial antibody profile.