TY - JOUR
T1 - A novel cystatin derived from trichinella spiralis suppresses macrophage-mediated inflammatory responses
AU - Kobpornchai, Porntida
AU - Flynn, Robin J.
AU - Reamtong, Onrapak
AU - Rittisoonthorn, Nonglucksanawan
AU - Kosoltanapiwat, Nathamon
AU - Boonnak, Kobporn
AU - Boonyuen, Usa
AU - Ampawong, Sumate
AU - Jiratanh, Montakan
AU - Tattiyapong, Muncharee
AU - Adisakwattana, Poom
N1 - Funding Information:
This research project was supported by Mahidol University (Fiscal Year 2015-2017) and the Thailand Research Fund through a Royal Golden Jubilee Ph.D. scholarship to Porntida Kobpornchai (PHD/0209/2559). This study was also supported by ICTM Grants from the Faculty of Tropical Medicine, Mahidol University. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. We gratefully thank the Faculty of Tropical Medicine, Mahidol University for supporting all facilities and services. Our gratitude also goes to the Central Equipment Unit, Faculty of Tropical Medicine, Mahidol University for facilitating us all necessary instruments.
Publisher Copyright:
© 2020 Kobpornchai et al.
PY - 2020/4
Y1 - 2020/4
N2 - Trichinella spiralis can modulate host immune responses to retain a suitable environment for its long-term survival. Incidentally, the parasite elicits regulatory effects through immuno-modulatory molecule release, which can suppress host inflammation and may be used for the treatment of unrelated inflammatory diseases in someday. Here we identified and characterized a novel T. spiralis cystatin (TsCstN), which inhibits inflammation mediated by LPS-treated macrophages.Proteins contained in the excretory–secretory (ES) product of muscle-stage T. spiralis (ES-L1) were fractionated, and each was treated with mouse bone marrow-derived macrophages (mBMDMs) before LPS stimulation. The fractions that exhib-ited high immunomodulatory property by decreasing pro-inflammatory cytokines or increas-ing anti-inflammatory cytokines were identified by mass spectrometry. Incidentally, the conserved hypothetical protein (Tsp_04814) was selected for further characterization as it presented the most significant MS score. An annotation of Tsp_04814 using protein structural homology comparison suggested that it has high structural similarity to human cystatin E/M (TM score 0.690). The recombinant T. spiralis novel cystatin (rTsCstN) was expressed in Escherichia coli at a molecular weight of approximately 13 kDa. Mouse anti-rTsCstN poly-clonal antibody (pAb) could detect native TsCstN in crude worm antigens (CWA) and ES-L1 and be predominantly localized in the stichosome and subcuticular cells. rTsCstN inhibited cysteine proteases in vitro, especially cathepsin L, at an optimal pH of 6. Besides, rTsCstN could be internalized into mBMDMs, which were mostly distributed in the cytoplasm and lysosome both before and after LPS stimulation. To evaluate the rTsCstN immunomodula-tory properties on mBMDMs, rTsCstN was incubated with mBMDM before LPS stimulation; this demonstrated that rTsCstN suppressed pro-inflammatory cytokine production and MHC class II expression.T. spiralis L1-derived TsCstN was characterized as a novel cysteine protease inhibitor. The protein elicits an anti-inflammatory property by suppressing pro-inflammatory cytokines and interfering with the antigen presentation process through deple-tion of MHC class II expression.
AB - Trichinella spiralis can modulate host immune responses to retain a suitable environment for its long-term survival. Incidentally, the parasite elicits regulatory effects through immuno-modulatory molecule release, which can suppress host inflammation and may be used for the treatment of unrelated inflammatory diseases in someday. Here we identified and characterized a novel T. spiralis cystatin (TsCstN), which inhibits inflammation mediated by LPS-treated macrophages.Proteins contained in the excretory–secretory (ES) product of muscle-stage T. spiralis (ES-L1) were fractionated, and each was treated with mouse bone marrow-derived macrophages (mBMDMs) before LPS stimulation. The fractions that exhib-ited high immunomodulatory property by decreasing pro-inflammatory cytokines or increas-ing anti-inflammatory cytokines were identified by mass spectrometry. Incidentally, the conserved hypothetical protein (Tsp_04814) was selected for further characterization as it presented the most significant MS score. An annotation of Tsp_04814 using protein structural homology comparison suggested that it has high structural similarity to human cystatin E/M (TM score 0.690). The recombinant T. spiralis novel cystatin (rTsCstN) was expressed in Escherichia coli at a molecular weight of approximately 13 kDa. Mouse anti-rTsCstN poly-clonal antibody (pAb) could detect native TsCstN in crude worm antigens (CWA) and ES-L1 and be predominantly localized in the stichosome and subcuticular cells. rTsCstN inhibited cysteine proteases in vitro, especially cathepsin L, at an optimal pH of 6. Besides, rTsCstN could be internalized into mBMDMs, which were mostly distributed in the cytoplasm and lysosome both before and after LPS stimulation. To evaluate the rTsCstN immunomodula-tory properties on mBMDMs, rTsCstN was incubated with mBMDM before LPS stimulation; this demonstrated that rTsCstN suppressed pro-inflammatory cytokine production and MHC class II expression.T. spiralis L1-derived TsCstN was characterized as a novel cysteine protease inhibitor. The protein elicits an anti-inflammatory property by suppressing pro-inflammatory cytokines and interfering with the antigen presentation process through deple-tion of MHC class II expression.
UR - http://www.scopus.com/inward/record.url?scp=85083546389&partnerID=8YFLogxK
U2 - 10.1371/journal.pntd.0008192
DO - 10.1371/journal.pntd.0008192
M3 - Article
C2 - 32236093
AN - SCOPUS:85083546389
VL - 14
SP - 1
EP - 24
JO - PLoS Neglected Tropical Diseases
JF - PLoS Neglected Tropical Diseases
SN - 1935-2727
IS - 4
M1 - e0008192
ER -
