Type I Cystatin Derived from Fasciola gigantica Suppresses Macrophage-Mediated Inflammatory Responses

Abstract

There is an inverse relationship between the high incidence of helminth infection and the low incidence of inflammatory disease. Hence, it may be that helminth molecules have anti-inflammatory effects. Helminth cystatins are being extensively studied for anti-inflammatory potential. Therefore, in this study, the recombinant type I cystatin (stefin-1) of Fasciola gigantica (rFgCyst) was verified to have LPS-activated anti-inflammatory potential, including in human THP-1-derived macrophages and RAW 264.7 murine macrophages. The results from the MTT assay suggest that rFgCyst did not alter cell viability; moreover, it exerted anti-inflammatory activity by decreasing the production of proinflammatory cytokines and mediators, including IL-1β, IL-6, IL-8, TNF-α, iNOS, and COX-2 at the gene transcription and protein expression levels, as determined by qRT-PCR and Western blot analysis, respectively. Further, the secretion levels of IL-1β, IL-6, and TNF-α determined by ELISA and the NO production level determined by the Griess test were decreased. Furthermore, in Western blot analysis, the anti-inflammatory effects involved the downregulation of pIKKα/β, pIκBα, and pNF-κB in the NF-κB signaling pathway, hence reducing the translocation from the cytosol into the nucleus of pNF-κB, which subsequently turned on the gene of proinflammatory molecules. Therefore, cystatin type 1 of F. gigantica is a potential candidate for inflammatory disease treatment.

Molecular and biochemical characterizations of a Fasciola gigantica retinoid X receptor-α isoform A (FgRXRα-A)

“…All collected fractions were run on 12.5% SDS-PAGE and stained with Coomassie brilliant blue G-250 for evaluation. At the same time, the recombinant thioredoxin (rTrx) was produced from BL21(DE3) E. coil containing pET32a(+) and purified under native conditions as previously described 17 , 35 , 36 . The purified r Fg RXRα-A + Trx and rTrx were transferred to a micro-dialysis bag (Sigma-Aldrich, St. Louis, MO, USA) for dialysis against PBS, pH 7.4.…”

Cystatins from the Human Liver Fluke Opisthorchis viverrini: Molecular Characterization and Functional Analysis

“…Proteinase inhibitors are of interest as they not only regulate the proteinases of the parasite but also those of the host. A few cystatins have been molecularly characterized in the trematodes Schistosoma sp., Fasciola sp., and Clonorchis sinensis , for example, two Fasciola gigantica cystatins, both classified as type 1 cystatin, were found to be involved with regulating intracellular cysteine proteinase activity and protecting against extracellular proteolytic processes [ 8 , 9 , 10 , 11 , 12 , 13 , 14 ]. From our previous study of type 3 multi-domain cystatin in Fasciola gigantica , we found that processed cystatin domains inhibited both parasite and mammalian cysteine proteinases.…”