Background Schistosoma japonicum is a parasitic flatworm that causes human schistosomiasis. Secreted extracellular vesicles (EVs) play a key role in pathogen-host interfaces. Previous studies have shown that S. japonicum adult worms can release microRNA (miRNA)-containing EVs, which can transfer their cargo to mammalian cells and regulate gene expression in recipient cells. Tissue-trapped eggs are generally considered the major contributor to the severe pathology of schistosomiasis; however, the interactions between the host and parasite in this critical stage remain largely unknown.MethodsThe culture medium for S. japonicum eggs in vitro was used to isolate EVs. Transmission electron microscopy (TEM) analysis was used to confirm that vesicles produced by the eggs were EVs based on size and morphology. Total RNA extracted from EVs was analyzed by Solexa technology to determine the miRNA profile. The in vitro internalization of the EVs by mammalian cells was analyzed by confocal microscopy. The presence of EVs associated miRNAs in the primary hepatocytes of infected mice was determined by quantitative real-time PCR (qRT-PCR).ResultsEVs were isolated from the culture medium of in vitro cultivated S. japonicum eggs. TEM analysis confirmed that nanosized vesicles were present in the culture medium. RNA-seq analysis showed that the egg-derived EVs contained small non-coding RNA (sncRNA) populations including miRNAs, suggesting a potential role in host manipulation. This study further showed that Hepa1-6, a murine liver cell line, internalized the purified EVs and their cargo miRNAs that were detectable in the primary hepatocytes of mice infected with S. japonicum.Conclusions Schistosoma japonicum eggs can release miRNA-containing EVs, and the EVs can transfer their cargo to recipient cells in vitro. These results demonstrate the regulatory potential of S. japonicum egg EVs at the parasite-host interface.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-016-1845-2) contains supplementary material, which is available to authorized users.

Cervical cancer caused by infection with high-risk human papillomavirus remains to be the most deadly gynecologic malignancy worldwide. It is well documented that persistent expression of two oncogenes (E6/E7) plays the key roles in cervical cancer. Thus, in vivo detection of the oncoproteins is very important for the diagnosis of the cancer. Recently, affibody molecules have been demonstrated to be a powerful targeting probe for tumor–targeted imaging and diagnosis. In this study, four HPV16 E7-binding affibody molecules (ZHPV16 E7127, ZHPV16E7301, ZHPV16E7384 and ZHPV16E7745) were screened from a phage-displayed peptide library and used for molecular imaging in tumor-bearing mice. Biosensor binding analyses showed first that the four affibody molecules bound to HPV16 E7 with very high affinity and specificity. They co-localized with E7 protein only in two HPV16-positive cancer cells (SiHa and CaSki). Furthermore, affibody ZHPV16E7384 was conjugated with Dylight755 and used for in vivo tumor-imaging. Strongly high-contrast tumor retention of this affibody only occurred in HPV16-derived tumors of mice as early as 30 min post-injection, not in HPV-negative and HPV18-derived tumors. The accumulation of Dylight755-conjugated ZHPV16E7384 in tumor was achieved over a longer time period (24 h). The data here provide strong evidence that E7-specific affibody molecules have great potential used for molecular imaging and diagnosis of HPV-induced cancers.

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Cervical cancer, the second most common cause of cancer death in women worldwide, is significantly associated with infection of high-risk human papillomaviruses (HPVs), especially the most common genotype, HPV 16. To date, there is no established noninvasive therapy to treat cervical cancer.Methods: Here, we report a novel affitoxin that targets HPV16 E7 protein, one of the primary target proteins in molecular targeted therapy for HPV-induced cervical cancer. The affitoxin, ZHPV16E7 affitoxin384 was generated by fusing the modified Pseudomonas Exotoxin A (PE38KDEL) to the HPV16 E7-specific affibody. The expressed and purified ZHPV16E7 affitoxin384 was characterized using numerous methods. SPR assay, indirect immunofluorescence assay, and near-infrared (NIR) optical imaging were respectively performed to assess the targeting ability of ZHPV16E7 affitoxin384 to HPV16 E7 protein both in vitro and in vivo. Cell viability assays and SiHa tumor-bearing nude mice were used to evaluate the efficacy of ZHPV16 E7 affitoxin384 in vitro and in vivo, respectively.Results: Using in vitro methods the SPR assay and indirect immunofluorescence assay showed that ZHPV16E7 affitoxin384 targeted HPV16 E7 with high binding affinity and specificity. Significant reduction of cell viability in HPV16 positive cells was observed in the presence of ZHPV16 E7 affitoxin384. By NIR optical imaging, ZHPV16 E7 affitoxin384 specifically targeted HPV16 positive tumors in vivo. ZHPV16E7 affitoxin384 showed significant in vivo antitumor efficacy in two kinds of tumor-bearing nude mouse models.Conclusions: ZHPV16E7 affitoxin384 is a potent anti-cervical cancer therapeutic agent that could be effective against HPV16 positive tumors in humans.

In addition to regulating apoptosis via its interaction with the death domain of Fas receptor, death domain associated protein 6 (Daxx) is also known to be involved in transcriptional regulation, suggesting that the function of Daxx depends on its subcellular localization. In this study, we aimed to explore Daxx subcellular localization in gastric cancer (GC) cells and correlate the findings with clinical data in GC patients. Seventy pairs of tissue samples (GC and adjacent normal tissue) were analyzed immunohistochemically for Daxx expression and localization (nuclear and cytoplasmic). The Daxx Nuclear/Cytoplasmic ratio (Daxx NCR) values in tissue microarray data with 522 tumor samples were further analyzed. The defined Prior cohort (n = 277, treatment between 2006 and 2009) and Recent cohort (n = 245, treatment between 2010 and 2011) were then used to examine the relationship between Daxx NCR and clinical data. The Daxx NCR was found to be clinically informative and significantly higher in GC tissue. Using Daxx NCR (risk ratio = 2.0), both the Prior and Recent cohorts were divided into high‐ and low‐risk groups. Relative to the low‐risk group, the high‐risk patients had a shorter disease free survival (DFS) and overall survival (OS) in both cohorts. Importantly, postoperative chemotherapy was found having differential effect on high‐ and low‐risk patients. Such chemotherapy brought no survival benefit, (and could potentially be detrimental,) to high‐risk patients after surgery. Daxx NCR could be used as a prognosis factor in GC patients, and may help select the appropriate population to benefit from chemotherapy after surgery.

Chlamydia trachomatis is the leading cause of sexually transmitted infections worldwide. There is currently no commercially available vaccine against C. trachomatis. Major outer membrane protein (MOMP) of C. trachomatis is considered to be an ideal candidate for prophylactic vaccine. We designed a MOMP multi-epitope containing T- and B-cell epitope-rich peptides and developed hepatitis B surface antigen (HBsAg) as antigen delivery vehicle. In order to study the immunogenicity and efficacy of the candidate vaccine in a murine model of chlamydial genital infection, we engineered a recombinant plasmid expressing HBsAg and MOMP multi-epitope genes. Results of reverse transcription polymerase chain reaction and immunofluorescence assay revealed successful expression of the recombinant HBsAg/MOMP multi-epitope gene at both the transcription and translation levels. Intramuscular administration in mice was able to elicit not only antibodies against Chlamydia and HBsAg but also cytotoxic T lymphocyte activity against Chlamydia. In addition, mice inoculated with the rHBsAg were highly resistant to C. trachomatis genital infection. The rHBsAg DNA with MOMP multi-epitope appended at the C terminus of the HBsAg stimulated a stronger immune response and protective response than that appended at the N terminus. Together, our results suggested that use of a recombinant HBsAg encoding the MOMP multi-epitope could be a powerful approach to developing a safe and immunogenic C. trachomatis vaccine.

Schistosoma japonicum eggs trapped in host liver secretes microRNA (miRNA)-containing extracellular vesicles (EVs) that can be transferred to host cells. Recent studies demonstrated that miRNAs derived from plants can modulate gene expression and phenotype of mammalian cells in a cross-kingdom manner. In this study, we identified a Schistosoma japonicum miRNA (e.g., Sja-miR-3096) that is present in the hepatocytes of mice infected with the parasite and has notable antitumor effects in both in vitro and in vivo models. The Sja-miR-3096 mimics suppressed cell proliferation and migration of both murine and human hepatoma cell lines by targeting phosphoinositide 3-kinase class II alpha (PIK3C2A). We generated a murine hepatoma cell line that stably expressed the pri-Sja-miR-3096 gene and demonstrated cross-species processing of the schistosome pri-miRNA to the mature Sja-miR-3096 in the mammalian cell. Importantly, inoculation of this cell line into the scapula and livers of mice led to a complete suppression of tumorigenesis of the hepatoma cells. Moreover, tumor weight was significantly reduced on intravenous administration of Sja-miR-3096 mimics. Thus, the schistosome miRNA-mediated antitumor activity occurs in host liver cells during schistosome infection, which may strengthen resistance of host to liver cancer, and discovery and development of such miRNAs may present promising interventions for cancer therapy.

Nasopharyngeal carcinoma (NPC) induced by latent infection with Epstein-Barr virus (EBV) remains the most common head and neck cancer in Southeast Asia, especially in the southern part of China. It is well known that persistent expression of two EBV latent membrane proteins (LMP1/LMP2A) plays a key role in nasopharyngeal carcinogenesis. Therefore, the therapeutic approach of targeting the LMP1/LMP2A protein and subsequently blocking the LMP1/ LMP2A-mediated signalling pathway has been considered for treating patients with NPC. Recently, affibody molecules, a new class of small (~6.5 kDa) affinity proteins, have been confirmed to be powerful generalisable tools for developing imaging or therapeutic agents by targeting specific molecules. In this study, three EBV LMP2A N-terminal domain-binding affibody molecules (Z LMP2A-N 85, Z LMP2A-N 110 and Z LMP2A-N 252) were identified by screening a phagedisplayed peptide library, and their high affinity and specificity for the EBV LMP2A N-terminal domain were confirmed by surface plasmon resonance (SPR), indirect immunofluorescence, co-immunoprecipitation and near-infrared small animal fluorescence imaging in vitro and in vivo. Moreover, affibody molecules targeting the EBV LMP2A N-terminal domain significantly reduced the viability of the EBV-positive cell lines C666-1, CNE-2Z and B95-8. Further investigations showed that affibody Z LMP2A-N 110 could inhibit the phosphorylation of AKT, GSK-3β and β-catenin signalling proteins, leading to suppression of β-catenin nuclear translocation and subsequent inhibition of c-Myc oncogene expression, which may be responsible for the reduced viability of NPC-derived cell lines. In conclusion, our findings provide a strong evidence that three novel EBV LMP2A N-terminal domain-binding affibody molecules have great potential for utilisation and development as agents for both molecular imaging and targeted therapy of EBVrelated NPC.