Continuous low-intensity ultrasound attenuates IL-6 and TNFα-induced catabolic effects and repairs chondral fissures in bovine osteochondral explants

Abstract

Background Cartilage repair outcomes are compromised in a pro-inflammatory environment; therefore, the mitigation of pro-inflammatory responses is beneficial. Treatment with continuous low-intensity ultrasound (cLIUS) at the resonant frequency of 5 MHz is proposed for the repair of chondral fissures under pro-inflammatory conditions. Methods Bovine osteochondral explants, concentrically incised to create chondral fissures, were maintained under cLIUS (14 kPa (5 MHz, 2.5 Vpp), 20 min, 4 times/day) for a period of 28 days in the presence or absence of cytokines, interleukin-6 (IL-6) or tumor necrosis factor (TNF)α. Outcome assessments included histological and immunohistochemical staining of the explants; and the expression of catabolic and anabolic genes by qRT-PCR in bovine chondrocytes. Cell migration was assessed by scratch assays, and by visualizing migrating cells into the hydrogel core of cartilage-hydrogel constructs. Results Both in the presence and absence of cytokines, higher percent apposition along with closure of fissures were noted in cLIUS-stimulated explants as compared to non-cLIUS-stimulated explants on day 14. On day 28, the percent apposition was not significantly different between unstimulated and cLIUS-stimulated explants exposed to cytokines. As compared to non-cLIUS-stimulated controls, on day 28, cLIUS preserved the distribution of proteoglycans and collagen II in explants despite exposure to cytokines. cLIUS enhanced the cell migration irrespective of cytokine treatment. IL-6 or TNFα-induced increases in MMP13 and ADAMTS4 gene expression was rescued by cLIUS stimulation in chondrocytes. Under cLIUS, TNFα-induced increase in NF-κB expression was suppressed, and the expression of collagen II and TIMP1 genes were upregulated. Conclusion cLIUS repaired chondral fissures, and elicited pro-anabolic and anti-catabolic effects, thus demonstrating the potential of cLIUS in improving cartilage repair outcomes. Electronic supplementary material The online version of this article (10.1186/s12891-019-2566-4) contains supplementary material, which is available to authorized users.

Sustained Acoustic Medicine Combined with A Diclofenac Ultrasound Coupling Patch for the Rapid Symptomatic Relief of Knee Osteoarthritis: Multi-Site Clinical Efficacy Study

“…Recent preclinical and clinical studies have shown that ultrasound therapy has chondroprotective effects by inhibiting inflammatory factors [ 36 , 52 , 64 , 70 – 72 ]. The combined enhanced drug delivery of diclofenac and ultrasound chondroprotective effect makes SAM combined with diclofenac an effective treatment for arthritis associated pain.…”

Can Ultrasound Therapy Be an Environmental-Friendly Alternative to Non-Steroidal Anti-Inflammatory Drugs in Knee Osteoarthritis Treatment?

“…Continuous LIUS, at 300 mW/cm 2 intensity, 1 MHz frequency, suppressed TNF-α synthesis in OA cartilage in rabbit [ 99 ]. In bovine osteochondral explants with chondral fissures, continuous LIUS at frequency of 5 MHz, intensity <20 mW/cm 2 , at a constant pressure amplitude of 14 kPa, at input voltage of 2.5 Vpp (peak-to-peak voltage), suppressed TNF-α-induced increase in nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) expression and upregulated the expression of collagen II and tissue inhibitor of metalloproteinase-1 genes [ 100 ].…”

Continuous Low-Intensity Ultrasound Preserves Mitochondrial Potential, Inhibits NFκB Activation and Rescues Chondrogenesis of Mesenchymal Stromal Cells

“…All cytokines were purchased from R&D systems (Minneapolis, MN) and were employed at a concentration of 10 ng/ml. Non-focused immersion transducers (Panametrics V300, 12.7 mm diameters, Panametrix, Waltham, MA, USA) were used to apply cLIUS to plated MSCs using procedure detailed elsewhere 21 . MSCs were exposed to cytokines and cLIUS was applied one time for 10 or 20 min at 5 MHz (2.5 Vpp) with constant pressure amplitude of 14 kPa.…”

“…Cell viability in hydrogels was assessed by Live/Dead TM Viability/Cytotoxicity kit (Molecular Probes, USA) according to manufacturer's instructions 21 and visualized with the Zeiss LSM 700 Confocal Microscope. All the images were collected at 10x magni cation (z step size = 12 μm).…”

“…Low-intensity continuous ultrasound (cLIUS) when employed at 14 kPa (5 MHz and 2.5 Vpp) was shown to be a positive regulator of chondrogenesis in vitro 19,20 . Further, the chondroprotective effects of cLIUS (14 kPa) against proin ammatory cytokines on intact cartilage explants was demonstrated where cytokine-induced increases in NFκB expression and its downstream targets were suppressed, and the expression of COLII and TIMP1 genes were upregulated 21 . Recently the chondroinductive ability of cLIUS (14 kPa) was demonstrated in the absence of exogenously added growth factors 19,20,22 .…”