isolation-characterization-extracellular-vesicles-from-adult

Isolation and Characterization of Extracellular Vesicles from Adult <em>Schistosoma japonicum</em>

Extracellular vesicles (EVs) are membranous vesicles released by a variety of cells into the extracellular microenvironment. EVs represent a population of ...

Ministry of Agriculture

Inhibitor of apoptosis proteins (IAP) play an important role in the regulation of apoptotic processes and are defined by the presence of baculoviral IAP repeat (BIR) domains. Here, we characterized a cDNA fragment (SjBIRP) synthesized from the RNA of Schistosoma japonicum, which was found to contain the BIR domain, suggesting that it could encode a potential IAP. Real-time PCR analyses indicated that SjBIRP transcription was detected at several stages of the schistosome's lifecycle, with increased levels present in schistosomula (7 days). In addition, the SjBIRP was highly expressed in adult females as compared to adult males. A functional assay showed that SjBIRP could inhibit caspase3/7 activity in both HeLa cells and schistosome lysates. Furthermore, SjBIRP expression profiles varied between different hosts of S. japonicum. Taken together, our preliminary studies suggest that SjBIRP may play a functional role in the regulation of apoptosis in schistosomes, and that it could be a potential drug target for schistosomiasis control. 

Molecular characterization of SjBIRP, another apoptosis inhibitor, from Schistosoma japonicum.

Schistosomiasis is a worldwide parasitic disease, and while it can be successfully treated with chemotherapy, this does not prevent reinfection with the parasite. Adenovirus vectors have been widely used for vaccine delivery, and a vaccination approach has the potential to prevent infection with Schistosoma. Here, we developed a recombinant adenoviral vector that expresses Schistosoma japonicum inhibitor apoptosis protein (Ad-SjIAP) and assessed its immunoprotective functions against schistosomiasis in mice. Murine immune responses following vaccination were investigated using enzyme-linked immunosorbent assays (ELISA), lymphocyte proliferation, and cytokine assays. The protective immunity in mice was evaluated by challenging with S. japonicum cercariae. Our results indicated that immunization with the Ad-SjIAP in mice induced a strong serum IgG response against IAP including IgG1, IgG2a, and IgG2b. In addition, lymphocyte proliferation experiments showed that mice treated with Ad-SjIAP significantly increased the lymphocyte response upon stimulation with recombinant Schistosoma japonicum inhibitor apoptosis protein (rSjIAP). Moreover, cytokine assays indicated that vaccination of Ad-SjIAP significantly increased the production of interferon (IFN)-γ and IL-2 as compared to the corresponding control group. Furthermore, following the challenge with S. japonicum cercariae, the vaccine conferred moderate protection, with an average rate of 37.95% for worm reduction and 31.7% for egg reduction. Taken together, our preliminarily results suggested that schistosoma IAP may be a potential vaccine against S. japonicum and that adenoviral vectors may serve as an alternative delivery vehicle for schistosome vaccine development.

Evaluation of protective immune response in mice by vaccination the recombinant adenovirus for expressing Schistosoma japonicum inhibitor apoptosis protein.

Schistosomes, a class of parasitic trematode worms, cause schistosomiasis. Accumulating evidence suggests that microRNAs (miRNAs)-small, non-coding RNAs that are known to play critical regulatory roles in many organisms-may be involved in schistosome development and sexual maturation, as well as the pathogenesis of schistosomiasis. Schistosoma miRNAs, such as Bantam and miR-10, may be involved in the pathological processes of schistosomiasis, and recent studies suggest that schistosome-specific miRNAs (e.g., Bantam, miR-3479-3p) in the bloodstream of a final host could be used as biomarkers for schistosomiasis diagnosis. Furthermore, aberrant miRNAs, such as miR-223 and miR-454, can be produced by a host in response to schistosome infection, and these miRNAs may contribute to the pathogenesis of schistosomiasis-associated liver injury. Here, we summarize recent progress evaluating the relationship between schistosome miRNAs and schistosomiasis and discuss how these miRNAs can mediate the pathogenesis of schistosomiasis and be used as biomarkers for schistosomiasis diagnosis. 

Role of microRNAs in schistosomes and schistosomiasis.

Dioecious flatworms of the genus Schistosoma causes schistosomiasis, which is a major public health problem in developing countries. Acquiring detailed knowledge of schistosome-host interactions may aid in the development of novel strategies for schistosomiasis control. MicroRNAs (miRNAs) are involved in processes such as development, cell proliferation, metabolism, and signal transduction. Circulating miRNAs not only serve as a novel class of biomarkers of many diseases but also regulate target gene expression in recipient cells, which are similar to hormones.

Altered levels of circulating miRNAs are associated Schistosoma japonicum infection in mice.

Schistosomes, blood flukes, are an important global public health concern. Paired adult female schistosomes produce large numbers of eggs that are primarily responsible for the disease pathology and critical for dissemination. Consequently, understanding schistosome sexual maturation and egg production may open novel perspectives for intervening with these processes to prevent clinical symptoms and to interrupt the life-cycle of these blood-flukes. microRNAs (miRNAs) are key regulators of many biological processes including development, cell proliferation, metabolism, and signal transduction. Here, we report on the identification of Schistosoma japonicum miRNAs using small RNA deep sequencing in the key stages of male-female pairing, gametogenesis, and egg production. We identified 38 miRNAs, including 10 previously unknown miRNAs. Eighteen of the miRNAs were differentially expressed between male and female schistosomes and during different stages of sexual maturation. We identified 30 potential target genes for 16 of the S. japonicum miRNAs using antibody-based pull-down assays and bioinformatic analyses. We further validated some of these target genes using either in vitro luciferase assays or in vivo miRNA suppression experiments. Notably, suppression of the female enriched miRNAs bantam and miR-31 led to morphological alteration of ovaries in female schistosomes. These findings uncover key roles for specific miRNAs in schistosome sexual maturation and egg production. 

MicroRNAs Are Involved in the Regulation of Ovary Development in the Pathogenic Blood Fluke Schistosoma japonicum.

Secreted extracellular vesicles play an important role in pathogen-host interactions. Increased knowledge of schistosome extracellular vesicles could provide insights into schistosome-host interactions and enable the development of novel intervention strategies to inhibit parasitic processes and lessen disease transmission. Here, we describe biochemical characterization of Schistosoma japonicum exosome-like vesicles (S. japonicum EVs). A total of 403 proteins were identified in S. japonicum EVs, and bioinformatics analyses indicated that these proteins were mainly involved in binding, catalytic activity, and translation regulatory activity. Next, we characterized the population of small RNAs associated with S. japonicum EVs. Further studies demonstrated that mammalian cells could internalize S. japonicum EVs and transfer their cargo miRNAs to recipient cells. Additionally, we found that a specific miRNA, likely originating from a final host, ocu-miR-191-5p, is also associated with S. japonicum EVs. Overall, our findings demonstrate that S. japonicum EVs could be implicated in the pathogenesis of schistosomiasis via a mechanism involving the transfer of their cargo miRNAs to hosts. Our findings provide novel insights into the mechanisms of schistosome-host interactions.

Lihui Zhu

Shanghai Veterinary Research Institute,
Chinese Academy of Agricultural Sciences,
Key Laboratory of Animal Parasitology,
Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Key Laboratory of Animal Parasitology

Ministry of Agriculture

Isolation and Characterization of Extracellular Vesicles from Adult Schistosoma japonicum