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. 2023 Sep;27(17):2614-2625.
doi: 10.1111/jcmm.17894. Epub 2023 Aug 2.

Characterisation of extracellular vesicles isolated from hydatid cyst fluid and evaluation of immunomodulatory effects on human monocytes

Mojdeh Khosravi  1 , Hanieh Mohammad Rahimi  2 , Abdoreza Nazari  3 , Kaveh Baghaei  4 , Hamid Asadzadeh Aghdaei  4 , Shabnam Shahrokh  5 , Meysam Sharifdini  6 , Ana Claudia Torrecilhas  7 , Fatemeh Mehryab  3   8 , Hamed Mirjalali  2 , Faezeh Shekari  3 , Mohammad Reza Zali  5
Affiliations

Characterisation of extracellular vesicles isolated from hydatid cyst fluid and evaluation of immunomodulatory effects on human monocytes

Mojdeh Khosravi et al. J Cell Mol Med. 2023 Sep.

Abstract

Hydatidosis is a disease caused by the larval stage of Echinococcus granulosus, which involves several organs of intermediate hosts. Evidence suggests a communication between hydatid cyst (HC) and hosts via extracellular vesicles. However, a little is known about the communication between EVs derived from HC fluid (HCF) and host cells. In the current study, EVs were isolated using differential centrifugation from sheep HCF and characterized by western blot, electron microscope and size distribution analysis. The uptake of EVs by human monocyte cell line (THP-1) was evaluated. The effects of EVs on the expression levels of pro- and anti-inflammatory cytokines were investigated using quantitative real-time PCR (RT-PCR), 3 and 24 h after incubation. Moreover, the cytokine level of IL-10 was evaluated in supernatant of THP-1 cell line at 3 and 24 h. EVs were successfully isolated and showed spherical shape with size distribution at 130.6 nm. After 3 h, the expression levels of pro-inflammatory cytokine genes (IL1Β, IL15 and IL8) were upregulated, while after 24 h, the expression levels of pro-inflammatory cytokines were decreased and IL13 gene expression showed upregulation. A statistically significant increase was seen in the levels of IL-10 after 24 h. The main mechanism of the communication between EVs derived from HCF and their host remains unclear; however, time-dependent anti-inflammatory effects in our study suggest that HC may modulate the immune responses via EVs.

Keywords: Echinococcus granulosus; extracellular vesicles; human monocyte cell line; hydatid cyst; inflammation.

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Conflict of interest statement

The authors declare that they have no conflict of interest.

Figures

FIGURE 1
FIGURE 1
The immunologic and morphometric analyses of isolated EVs and an immunologic comparison between EVs and the parasite lysate. (A) The westernblot analysis suggests the presence of CD63 marker and the lack of CD81 marker. In addition, EVs and parasite lysate exhibited similar expression of specific markers. (B) The analysis of SEM microscope represented spherical shape EVs isolated from HCF. (C) DLS analysis shows that EVs mostly presented size at 130.6 nm.
FIGURE 2
FIGURE 2
The comparison of the expression levels of inflammatory IL1Β, IL15, IL8, TNFΑ, IFNG and NLRP3 genes in THP‐1 cell line co‐incubated with HCF derived EVs, compared to the housekeeping gene (ACTB) for (A) 3 h and (B) 24 h. This picture shows also the expression of anti‐inflammatory biomarkers IL4, TGFΒ, and IL13 genes in THP‐1 cell line co‐incubated with HCF derived EVs (C) 3 h and (D) 24 h after exposure. The expression levels of pro‐inflammatory cytokines are time‐dependently decreased 24 h after exposure to HCF EVs. * p < 0.05; ** p < 0.01; *** p < 0.001. ACTB, Beta Actin; C, control; IFN: interferon; IL, Interleukin; NLRP, nod‐like receptor pyrin domain containing; TGF, tumour growth factor; TNF, tumour necrosis factor.
FIGURE 3
FIGURE 3
The expression levels of (A) IL1Β, (B) IL15, (C) IL8, (D) IFNG, (E) TNFΑ, (F) NLRP3, (G) TGFΒ, (H) IL4, and (I) IL13 genes in THP‐1 cell line co‐incubated with HCF derived EVs, compared to the housekeeping gene (ACTB), 3 and 24 h after exposure. This picture shows that the expression levels of pro‐inflammatory cytokines are time‐dependently decreased 24 h after exposure to HCF derived EVs. * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001. ACTB, Beta Actin; C, Control; IFN: Interferon; IL, Interleukin; NLRP, Nod‐like receptor pyrin domain containing; TGF, Tumour growth factor; TNF, Tumour necrosis factor.
FIGURE 4
FIGURE 4
The cytokine release of IL‐10 shows the concentration of this cytokine in 3 h and 24 h after treatment with HCF derived EVs. It seems that treatment with HCF EVs decreases the released concentration of IL‐10. * p < 0.05; *** p < 0.001; **** p < 0.0001. IL, interleukin.
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