Effects of Gnathostoma spinigerum infective stage larva excretory-secretory products on NK cells in peripheral blood mononuclear cell culture: focused on expressions of IFN-γ and killer cell lectin-like receptors
Human gnathostomiasis is mainly caused by third-stage larvae of Gnathostoma spinigerum (G. spinigerum L3). Excretory-secretory products (ES) released from infective helminthic larvae are associated with larval migration and host immunity modulation. Natural killer (NK) cells have important immune functions against helminth infection. Currently, the effects of ES from G. spinigerum L3 (G. spinigerum ES) on NK cell activity are unclear. This study investigated whether G. spinigerum ES affected human NK cells. Human normal peripheral blood mononuclear cell (PBMC) cultures were used to mimic immune cells within the circulation. PBMC were co-cultured with G. spinigerum ES (0.01–0.05 μg/ml) for 5 or 7 days. Levels of IFN-γ in cultured supernatants were measured by enzyme-linked immunosorbent assay. The expressions of mRNA encoding NK cell receptors, especially the C type killer cell lectin-like family (KLR; NKG2A, NKG2C, and NKG2D) and IFN-γ in ES induced PBMC were determined by quantitative reverse transcription-polymerase chain reaction (RT-qPCR). ES induced PBMC markedly decreased the levels of IFN-γ and increased the expressions of NKG2A and NKG2D on NK cells. In conclusion, low amounts of G. spinigerum ES modulated NK cells by downregulating the transcription of IFN-γ and upregulating the expressions of KLR (NKG2A and NKG2D receptors) during the 7-day observation period. These findings indicate more in-depth studies of NK cell function are required to better understand the mechanism involved in immune evasive strategies of human gnathostomiasis.
KeywordsGnathostoma spinigerum Excretory-secretory products NK cell Killer cell lectin-like receptors IFN-γ
The authors gratefully acknowledge the Mahidol-Oxford Tropical Medicine Research Unit (MORU) for providing access to the FACSCaliber flow cytometer and CellQuest software. We thank Mrs. Supaporn Nuamtanong for the collection of third-stage G. spinigerum larva cultures and preparation of ES. We also thank Edanz Group (www.edanzediting.com/ac) for editing a draft of this manuscript.
TK and UC were responsible for laboratory work, performed the RNA extraction, and carried out the real-time PCR assays and analyses. YM and WD conducted PBMC culture, and sample preparation for all experiments, and carried out the FACS analysis. WD and SB were responsible for data analysis and statistical analysis. YM, UC, PD, SA, and PV participated in the study design, conceived the study, and coordination. TK and YM contributed to writing the manuscript. All authors interpreted the results and have read and approved the final manuscript.
This study was supported by the Faculty of Tropical Medicine, Mahidol University (TK). It was also supported in part by Grant-in-Aids from the Ministry of Health, Labor, and Welfare of Japan (No.H20-Shinko-Ippan-015) to YE and from the Ministry of Education, Culture, Sports, Science and Technology of Japan (Kiban B, Overseas, No. 20401050) to YE.
Compliance with ethical standards
This article does not contain any studies with human participants or animals performed by any of the authors.
Conflict of interest
The authors declare that they have no competing interests.
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