| A series of experiments was conducted for the purpose of developing in vitro and in vivo methods for the evaluation of equine anti-inflammatory nutraceuticals. We have combined a cartilage explant culture system with a simulated digestion and ultrafiltration protocol to test dietary anti-inflammatories derived from a commercial nutraceutical, Sasha's EQ (SEQ; Interpath, Pty Ltd.). SEQ and its constituents [New Zealand Green Lipped Mussel (NZGLM), Abalone (AB), Shark Cartilage (SC) and Biota oil (BO)] were individually exposed to simulated gastric and intestinal fluid, then ultrafiltered to remove all particles > 50kDa. The ultrafiltrate was added to tissue culture medium within which cartilage explants were stimulated with 1L-1 to initiate an inflammatory condition in vitro. Simulated digest of SEQ significantly inhibited IL-1-induced prostaglandin E2 (PGE2) and nitric oxide (NO) while stimulating cell viability within cartilage explants; simulated digests of SC and NZGLM both decreased IL-1-induced PGE2 and glycosaminoglycan (GAG) release, and stimulated cell viability; AB significantly increased PGE 2 while decreasing NO; and simulated digest of BO strongly inhibited PGE2 and increased cell viability. In preliminary investigations of the effect of SEQ and all its constituents on gene expression, all constituents, with the exception of AB, decreased expression of genes encoding Cox1, Cox2 and iNOS, and increased gene expression of aggrecan. AB did not appear to affect Cox 1 or aggrecan expression, but may upregulate Cox 2 and downregulated iNOS expression; these preliminary gene expression data provide support for the observed increase in media [PGE2] and decreased media [NO] observed in in vitro experiments with AB. The in vitro data suggest a role for SEQ and its constituents in modulating the pathophysiological and molecular mechanisms involved in articular inflammation. SEQ was then tested in a novel, sub-clinical model of equine articular inflammation. IL-1 was injected into the intercarpal joint of 11 horses, and an equal volume of saline was injected into the contra-lateral joints. Dietary SEQ (n=5) for 2 weeks prior to IL-1 challenge prevented significant IL-1-induced elevation in synovial fluid PGE2, while increasing synovial fluid GAG. This provides evidence for potential anti-inflammatory role of SEQ in horses with mild inflammation. Before a definitive chondroprotective role of SEQ can be suggested, there is a need to investigate whether the increase in synovial fluid [GAG] resulted from an increase in accumulation of dietary GAGs or whether the synovial fluid GAGs originated from the cartilage tissue. The product was not associated with adverse effects as identifiable through blood biochemistry and hematology analysis. The in vitro and in vivo models used in these experiments provide useful insight into the mechanism of action of dietary nutraceuticals. Furthermore, Sasha's EQ and its constituents show some anti-inflammatory and/or chondroprotective characteristics further investigation into this product as a dietary intervention for joint inflammation in horses is warranted. |
