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Alveolar echinococcosis (AE) is a parasitic disease caused by E. multilocularis metacestodes and it is highly prevalent in the northern hemisphere. We have previously found that vaccination with E. multilocularis Leucine aminopeptidase (EM-LAP) induced specific immune response and had an inhibiting effect on the parasites. In this study, the therapeutic effect of recombinant EM-LAP (rEM-LAP) on AE was evaluated and verified using Ubenimex, a broad-spectrum inhibitor of LAP. The results reveal that rEM-LAP could inhibit cyst growth and invasion and induce specific immunity response in BALB/c mice infected with E. multilocularis protoscoleces. The ultrasonic, MRI, and morphological results show that treatment with rEM-LAP inhibits E. multilocularis infection and reduces cyst weight, number, fibrosis and invasion. The same effect is observed for the treatment with Ubenimex by inhibiting LAP activity. The indirect ELISA shows that rEM-LAP could induce specific immunity response and produce high levels of IgG, IgG1, IgG2a, IgM, and IgA, and the serum levels of IFN-γ and IL-4 are significantly increased compared to the control groups, indicating that treatment with rEM-LAP leads to a Th1 and Th2 mixed-type immune response. This study suggests that EM-LAP could be a potential therapeutic target of E. multilocularis infection.

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Pore types and pore structure parameters are the important factors affecting the storage capacity of a shale oil reservoir. Pore morphology and mineralogical composition of shales have diverse effects on the upgrading of various phases of shale oil. To interpret the formation and distribution of different pore types and their structure parameters in the lacustrine calcareous shale, a combination of polarizing microscopy, X-ray diffraction, total organic carbon (TOC), field-emission scanning electron microscopy, and low-pressure nitrogen adsorption experiments were conducted on the Es3x shale of the Eocene Shahejie Formation in the Zhanhua Depression. The interpretations regarding pore types, pore structure parameters, and pore size distribution indicate that the pore morphology and pore size distribution in the lacustrine shale are very complicated and demonstrate strong heterogenic behavior. Inorganic pores (interparticle pores, intraparticle pores, intercrystalline pores, dissolution pores, and microfractures) are the most commonly distributed pore types in the studied shale. However, organic matter pores are poorly developed due to the lower thermal maturity of the Es3x shale. The Brunauer-Emmett-Teller specific surface and pore volume range from 0.026 to 1.282 m2/g (average 0.697 m2/g) and 0.003 to 0.008 cm3/g (average 0.005 cm3/g), respectively. The shape of the pores varies from slit-like to narrow slit. Different minerals develop different types of pores with various sizes extending from micropores (<2 nm), mesopores (2-50 nm), to macropores (>50 nm). The relationship between mineral components and pore parameters indicates that the carbonate minerals act as the main contributors to the formation and distribution of different pore types in the studied shale. Pore volume and the pore specific surface area did not show a good relationship with mineral composition and TOC due to disordered pores, but pore size shows a good relationship with mineral composition and TOC of the Es3x shale. The whole pore system description showed that the mesopores and macropores are abundantly distributed and are the main contributors to the pore system in the Es3x shale. A comprehensive understanding of the formation mechanism and structural features of various sized pores in a variety of different minerals can provide a good tool for the exploration and development of shale oil reservoirs.

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Alveolar echinococcosis, a parasitic zoonotic disease caused by the larval stage of Echinococcus multilocularis infection, is a global epidemic in Eurasia and North America. Leucine aminopeptidase (LAP) of the M17 peptidase family could act on an ideal target antigen in diagnosis and prevention of parasitic diseases (schistosomiasis, malaria, fascioliasis) because of its good immunogenicity. In this study, the bioinformatic and enzymatic characterizations of recombinant Echinococcus multilocularis LAP (rEm-LAP) were evaluated. A prokaryotic expression system for rEm-LAP protein was established and its immunogenicity and preventive efficacy were demonstrated in a BALB/c mice model. This is the first report about the LAP of Echinococcus multilocularis and with a 57.4 KD purified rEm-LAP protein successfully expressed by pCzn1-LAP in Escherichia coli BL-21 cells. Enzymatic analysis results showed optimal rEm-LAP activity at pH 9. Serum indirect ELISA demonstrated that rEm-LAP could induce a Th1 and Th2 mixed-type immunological response and produce high levels of IgG, IgG1, IgG2a, IgM, and IgA. Furthermore, serum IFN-γ and IL-4 secretion were increased compared with the control groups. Finally, vaccination with rEm-LAP significantly decreased both the number and size of the cysts in Echinococcus multilocularis metacestode infected mice model. The current study provides evidence that rEm-LAP could be a potential vaccine antigen of Echinococcus multilocularis.

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The herbal orchid Bletilla striata (Thunb.) Rchb.f. has a long cultivation history and has been widely used in medicines and cosmetics. The fungal infection leaf blight (LB) seriously threatens B. striata cultivation. Here, we systemically collected wild B. striata accessions and isolated the accessions with strong resistance against LB. We carried out proteomic profiling analysis of LB-resistant and LB-susceptible accessions, and identified a large number of differentially expressed proteins with significant gene ontology enrichment for 'oxidoreductase activity.' Of the proteins identified in the reactive oxygen species signalling pathway, the protein abundance of the Cu-Zn superoxide dismutase BsSOD1 and its gene expression level were higher in LB-resistant accessions than in LB-susceptible lines. Transient expression of the dismutase fused with yellow fluorescent protein determined that its subcellular localisation is in the cytoplasm. Our study provides new insights into the molecular markers associated with fungal infection in B. striata.

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The herbaceous peony (Paeonia lactiflora Pall.) is a perennial flowering plant of the Paeoniaceae species that is widely cultivated for medical and ornamental uses. The monoterpene glucoside paeoniflorin and its derivatives are the active compounds of the P. lactiflora roots. However, the gene regulation pathways associated with monoterpene and paeoniflorin biosynthesis in P. lactiflora are still unclear. Here, we selected three genotypes of P. lactiflora with distinct morphologic features and chemical compositions that were a result of long-term reproductive isolation. We performed an RNA-sequencing experiment to profile the transcriptome changes of the shoots and roots. Using de novo assembly analysis, we identified 36,264 unigenes, including 521 genes responsible for encoding transcription factors. We also identified 28,925 unigenes that were differentially expressed in different organs and/or genotypes. Pathway enrichment analysis showed that the P. lactiflora unigenes were significantly overrepresented in several secondary metabolite biosynthesis pathways. We identified and profiled 33 genes responsible for encoding the enzymescontrolling the major catalytic reactions in the terpenoid backbone and in monoterpenoid biosynthesis. Our study identified the candidate genes in the terpenoid biosynthesis pathways, providing useful information for metabolic engineering of P. lactiflora intended for pharmaceutical uses and facilitating the development of strategies to improve marker-assist P. lactiflora in the future.

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OBJECTIVE: To discuss the effect and mechanism of miR-34a on the proliferation, apoptosis and invasion of laryngeal carcinoma cells. METHODS: The laryngeal squamous carcinoma Hep2 cells were transiently transfected with miR-34a mimics and miR-34a NC. The MTT, colony-forming assay, Hoechst staining and AnnexinV-PI double staining flow cytometry were employed to detect the effect of miR-34a on the viability and apoptosis of laryngeal squamous carcinoma Hep2 cells; Transwell assay to defect the effect of miR-34a on the migration and invasion of laryngeal squamous carcinoma Hep2 cells; western blot and RT-PCR assay to defect the effect of miR-34a mimics on the expression of survivin and Ki-67 mRNA in laryngeal squamous carcinoma Hep2 cells. RESULTS: Compared with miR-34a NC group, the cell viability in miR-34 mimics group was significantly decreased (P < 0.01), the cell apoptosis rate was significantly increased (P < 0.01), the abilities of cell migration and invasion were significantly reduced (P < 0.01) and the expression of survivin and Ki-67 mRNA was significantly decreased (P < 0.01). CONCLUSIONS: The increased expression of miR-34a can induce the apoptosis of Hep2 laryngeal carcinoma cells and inhibit the cell proliferation and invasion, which is related to the down-regulated expression of survivin and Ki-67.

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OBJECTIVE: To investigate the expression of Survivin, p53 and Ki67 in laryngeal carcinoma and the relation with clinical data. METHOD: Immunohistochemical staining (SP) was used to detect expression of Survivin, p53 and Ki67 of 64 cases with laryngeal carcinoma, 26 cases with precancerosis, 34 cases with vocal polyps. RESULT: The positive expression rates of Survivin, p53 and Ki67 were 59.4%, 68.8%, 65.6% respectively in laryngeal carcinoma, which were significantly higher than those in precancerosis and vocal polyps (P < 0.01). The expression of Survivin, p53 and Ki67 in laryngeal carcinoma were significantly statistical different in TNM stage and lymph node metastasis (P < 0.05), but were not correlated with patients' ages, the pathological grades, 3 years and 5 years surviving rates (P > 0.05). The expression of Survivin, Ki-67 and p53 was positively correlated (r = 0.607, 0.541, 0.648, P < 0.01) in laryngeal carcinoma. CONCLUSION: Survivin, p53 and Ki-67 may play an important role in the carcinogenesis and progress of laryngeal carcinoma. They may play synergetic roles in the process of carcinogenesis of laryngeal carcinoma.

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