Botulinum neurotoxin type A does not exert concentration-dependent effects on equine articular cartilage in vitro

Objective: To determine whether Botulinum neurotoxin type A (BoNT-A) ameliorates the effects of interleukin 1 (IL-1) on equine articular cartilage, or exerts negative effects on normal equine articular cartilage homeostasis in vitro.

Sample: Articular cartilage explants from 6 healthy femoropatellar joints of 3 adult horses.

Methods: Explants were allocated to the IL-1 challenged or unchallenged group, then exposed to 1 of 6 concentrations of BoNT-A (0, 1, 10, 50, 100, or 500 pg/mL) for 96 hours. To assess BoNT-A's effects on inflammation, prostaglandin E2 (PGE2) was measured in media via ELISA. Matrix degradation was determined as the percentage of sulfated glycosaminoglycans (sGAG) released from explants via dimethylmethylene blue assay. Aggrecan synthesis was estimated using CS846 ELISA and collagen type II degradation was estimated using C2C ELISA on media. Chondrocyte apoptosis was assessed via in-situ TUNEL assay. Generalized linear mixed models were fitted to determine treatment effects using α = 0.05.

Results: The challenge with IL-1 resulted in increased concentrations of PGE2 and CS846 in media and increased release of sGAG from explants. BoNT-A did not significantly impact PGE2 or CS846 concentration in media, percentage of sGAG released, or chondrocyte apoptosis in IL-1 challenged or unchallenged cartilage explants. The concentration of C2C in media was below the quantifiable limit of the ELISA in all samples.

Clinical relevance: BoNT-A did not show chondroprotective effects or have negative effects on cartilage homeostasis in vitro at the concentrations tested. While chondroprotective effects were not observed, BoNT-A may be safe for intraarticular use. In vivo testing is warranted before clinical use.