STANDARDIZATION OF A FLUORESCENT-BASED QUANTITATIVE ADHESION ASSAY TO STUDY ATTACHMENT OF Taenia solium ONCOSPHERE TO EPITHELIAL CELLS In Vitro

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Chile, Nancy; Evangelista, Julio; Gilman, Robert H.; Arana, Yanina; Palma, Sandra; Sterling, Charles R; Garcia, Hector H.; Gonzalez, Armando; Verastegui, Manuela

2012-01-01

To fully understand the preliminary stages of Taenia solium oncosphere attachment in the gut, adequate tools and assays are necessary to observe and quantify this event that leads to infection. A fluorescent-based quantitative adhesion assay, using biotinylated activated-oncospheres and monolayers of Chinese hamster ovary cells (CHO-K1) or human intestinal monolayer cells (INT-407, HCT-8 or HT-29), was developed to study initial events during the infection of target cells and to rapidly quantify the in vitro adhesion of T. solium oncospheres. Fluorescein streptavidin was used to identify biotinylated activated-oncospheres adhered to cells. This adherence was quantified using an automated fluorescence plate reader, and the results were expressed as fluorescence intensity values. A series of three assays were performed. The first was to identify the optimum number of biotinylated activated-oncospheres to be used in the adhesion assay. The goal of the second assay was to validate this novel method with the established oncosphere-binding system using the immunofluorescent-antibody assay (IFA) method to quantify oncosphere adhesion. A total of 10,000 biotinylated activated-oncospheres were utilized to assess the role of sera and laminin (LM) in oncosphere adherence to a CHO-K1 cell monolayer. The findings that sera and LM increase the adhesion of oncospheres to monolayer cells were similar to results that were previously obtained using the IFA method. The third assay compared the adherence of biotinylated activated-oncospheres to different types of human intestinal monolayer cells. In this case, the fluorescence intensity was greatest when using the INT-407 cell monolayer. We believe this new method of quantification offers the potential for rapid, large-scale screening to study and elucidate specific molecules and mechanisms involved in oncosphere-host cell attachment. PMID:22178422

Characterization of the carbohydrate components of Taenia solium oncosphere proteins and their role in the antigenicity.

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Arana, Yanina; Verastegui, Manuela; Tuero, Iskra; Grandjean, Louis; Garcia, Hector H; Gilman, Robert H

2013-10-01

This study examines the carbohydrate composition of Taenia solium whole oncosphere antigens (WOAs), in order to improve the understanding of the antigenicity of the T. solium. Better knowledge of oncosphere antigens is crucial to accurately diagnose previous exposure to T. solium eggs and thus predict the development of neurocysticercosis. A set of seven lectins conjugates with wide carbohydrate specificity were used on parasite fixations and somatic extracts. Lectin fluorescence revealed that D-mannose, D-glucose, D-galactose and N-acetyl-D-galactosamine residues were the most abundant constituents of carbohydrate chains on the surface of T. solium oncosphere. Lectin blotting showed that posttranslational modification with N-glycosylation was abundant while little evidence of O-linked carbohydrates was observed. Chemical oxidation and enzymatic deglycosylation in situ were performed to investigate the immunoreactivity of the carbohydrate moieties. Linearizing or removing the carbohydrate moieties from the protein backbones did not diminish the immunoreactivity of these antigens, suggesting that a substantial part of the host immune response against T. solium oncosphere is directed against the peptide epitopes on the parasite antigens. Finally, using carbohydrate probes, we demonstrated for the first time that the presence of several lectins on the surface of the oncosphere was specific to carbohydrates found in intestinal mucus, suggesting a possible role in initial attachment of the parasite to host cells.

The highly antigenic 53/25 kDa Taenia solium protein fraction with cathepsin-L like activity is present in the oncosphere/cysticercus and induces non-protective IgG antibodies in pigs

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Zimic, Mirko; Pajuelo, Mónica; Gilman, Robert H.; Gutiérrez, Andrés H.; Rueda, Luis D.; Flores, Myra; Chile, Nancy; Verástegui, Manuela; Gonzalez, Armando; García, Héctor H.; Sheen, Patricia

2011-01-01

Cathepsin L-like proteases are secreted by several parasites including Taenia solium. The mechanism used by T. solium oncospheres to degrade and penetrate the intestine and infect the host is incompletely understood. It is assumed that intestinal degradation is driven by the proteolytic activity of enzymes secreted by the oncosphere. Blocking the proteolytic activity by an antibody response would prevent the oncosphere penetration and further infection. Serine and cysteine proteases including chymotrypsin, trypsin, elastase, and cathepsin L, are secreted by T. solium and Taenia saginata oncospheres when cultured in vitro, being potential vaccine candidates. However, the purification of a sufficient quantity of proteases secreted by oncospheres to conduct a vaccine trial is costly and lengthy. A 53/25 kDa cathepsin L-like fraction partially purified from T. solium cyst fluid was described previously as an important antigen for immunodiagnostics. In this study we found that this antigen is present in the T. solium oncosphere and is also secreted by the cysticercus. This protein fraction was tested for its ability to protect pigs against an oral challenge with T. solium oncospheres in a vaccine trial. IgG antibodies against the 53/25 kDa cathepsin L-like protein fraction were elicited in the vaccinated animals but did not confer protection. PMID:22119017

Characterisation of antibody responses in pigs induced by recombinant oncosphere antigens from Taenia solium.

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Jayashi, César M; Gonzalez, Armando E; Castillo Neyra, Ricardo; Kyngdon, Craig T; Gauci, Charles G; Lightowlers, Marshall W

2012-12-14

Recombinant antigens cloned from the oncosphere life cycle stage of the cestode parasite Taenia solium (T. solium) have been proven to be effective as vaccines for protecting pigs against infections with T. solium. Previous studies have defined three different host protective oncosphere antigens, TSOL18, TSOL16 and TSOL45. In this study, we evaluated the potential for combining the antigens TSOL16 and TSOL18 as a practical vaccine. Firstly, in a laboratory trial, we compared the immunogenicity of the combined antigens (TSOL16/18) versus the immunogenicity of the antigens separately. Secondly, in a field trial, we tested the ability of the TSOL16/18 vaccine to induce detectable antibody responses in animals living under environmental stress and traditionally reared in areas where T. solium cysticercosis is endemic; and finally, we characterised the immune response of the study population. Pigs of 8-16 weeks of age were vaccinated with 200 μg each of TSOL16 and TSOL18, plus 5mg of Quil-A. Specific total IgG, IgG(1) and IgG(2) antibody responses induced by TSOL16 and TSOL18 were determined with ELISA. The immunogenicity of both antigens was retained in the combined TSOL16/18 vaccine. The combined vaccine TSOL16/18 induced detectable specific anti-TSOL18 antibody responses in 100% (113/113) and specific anti-TSOL16 in 99% (112/113) of the vaccinated animals measured at 2 weeks following the booster vaccination. From the two IgG antibody subtypes analysed we found there was stronger response to IgG(2). Copyright © 2012 Elsevier Ltd. All rights reserved.

Establishment and Biological Characterization of a Panel of Glioblastoma Multiforme (GBM) and GBM Variant Oncosphere Cell Lines.

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Binder, Zev A; Wilson, Kelli M; Salmasi, Vafi; Orr, Brent A; Eberhart, Charles G; Siu, I-Mei; Lim, Michael; Weingart, Jon D; Quinones-Hinojosa, Alfredo; Bettegowda, Chetan; Kassam, Amin B; Olivi, Alessandro; Brem, Henry; Riggins, Gregory J; Gallia, Gary L

2016-01-01

Human tumor cell lines form the basis of the majority of present day laboratory cancer research. These models are vital to studying the molecular biology of tumors and preclinical testing of new therapies. When compared to traditional adherent cell lines, suspension cell lines recapitulate the genetic profiles and histologic features of glioblastoma multiforme (GBM) with higher fidelity. Using a modified neural stem cell culture technique, here we report the characterization of GBM cell lines including GBM variants. Tumor tissue samples were obtained intra-operatively and cultured in neural stem cell conditions containing growth factors. Tumor lines were characterized in vitro using differentiation assays followed by immunostaining for lineage-specific markers. In vivo tumor formation was assayed by orthotopic injection in nude mice. Genetic uniqueness was confirmed via short tandem repeat (STR) DNA profiling. Thirteen oncosphere lines derived from GBM and GBM variants, including a GBM with PNET features and a GBM with oligodendroglioma component, were established. All unique lines showed distinct genetic profiles by STR profiling. The lines assayed demonstrated a range of in vitro growth rates. Multipotency was confirmed using in vitro differentiation. Tumor formation demonstrated histologic features consistent with high grade gliomas, including invasion, necrosis, abnormal vascularization, and high mitotic rate. Xenografts derived from the GBM variants maintained histopathological features of the primary tumors. We have generated and characterized GBM suspension lines derived from patients with GBMs and GBM variants. These oncosphere cell lines will expand the resources available for preclinical study.

Cloning and Expression of HSP of Taenia Solium Oncosphere%猪带绦虫六钩蚴 HSP 的克隆和表达

Institute of Scientific and Technical Information of China (English)

王哲; 赵权

2013-01-01

　　The HSP gene was separately amplified from total RNA of activated Taenia solium oncosphere by RT -PCR.The PCR products were cloned into pGH vector,recombinant positive clones was sequenced after restriction enzyme digestion.The HSP gene was subcloned into pET28a expression vector,the recombinant pET28a-HSP infected into E.coliBL21.IPTG was added to induce fusion expression and the expression products was identified by SDS-PAGE and Western-blot.One fusion protein band about 35 kDa was identified by SDS -PAGE after inducible expression after inducible expression,The result would lay foundations for the mechanism of invasion of between oncosphere and host,and the design of new vaccine anti-porcine cysticercosis and taeniasis.%　　提取猪带绦虫激活和未激活六钩蚴总 RNA，RT-PCR 扩增 HSP 目的基因，将目的基因与 pGH 克隆载体连接，经酶切鉴定后，将阳性重组质粒进行测序，结果扩增出激活的六钩蚴的目的片段。将目的基因亚克隆到原核表达载体 pET-28a-HSP中，并将获得的 pET28a-HSP 阳性重组子转化至宿主菌 E.coliBL21，IPTG 进行诱导表达，并对重组抗原 pET-28a-HSP 进行SDS-PAGE 和 Western-blot 检测。结果表明重组经 SDS-PAGE 分析可见一条约35 kDa 大小的融合蛋白条带的抗原，Western-blot 结果显示其能被囊虫病人阳性血清识别。这将为进一步阐明六钩蚴入侵中间宿主的机理、设计新型抗猪囊虫病和绦虫病疫苗打下基础。

Origin, differentiation and functional ultrastructure of egg envelopes in the cestode Echinococcus multilocularis Leuckart, 1863 (Cyclophyllidea: Taeniidae).

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Świderski, Zdzisław; Miquel, Jordi; Azzouz-Maache, Samira; Pétavy, Anne-Françoise

2017-07-01

The origin, differentiation and functional ultrastructure of oncospheral or egg envelopes in Echinococcus multilocularis Leuckart, 1863 were studied by transmission electron microscopy (TEM) and cytochemistry. The purpose of our study is to describe the formation of the four primary embryonic envelopes, namely vitelline capsule, outer envelope, inner envelope and oncospheral membrane, and their transformation into the oncospheral or egg envelopes surrounding the mature hexacanth. This transformation takes place in the preoncospheral phase of embryonic development. The vitelline capsule and oncospheral membrane are thin membranes, while the outer and inner envelopes are thick cytoplasmic layers formed by two specific types of blastomeres: the outer envelope by cytoplasmic fusion of two macromeres and the inner envelope by cytoplasmic fusion of three mesomeres. Both outer and inner envelopes are therefore cellular in origin and syncytial in nature. During the advanced phase of embryonic development, the outer and inner envelopes undergo great modifications. The outer envelope remains as a metabolically active layer involved in the storage of glycogen and lipids for the final stages of egg development and survival. The inner envelope is the most important protective layer because of its thick layer of embryophoric blocks that assures oncospheral protection and survival. This embryophore is the principal layer of mature eggs, affording physical and physiological protection for the differentiated embryo or oncosphere, since the outer envelope is stripped from the egg before it is liberated. The embryophore is very thick and impermeable, consisting of polygonal blocks of an inert keratin-like protein held together by a cementing substance. The embryophore therefore assures extreme resistance of eggs, enabling them to withstand a wide range of environmental temperatures and physicochemical conditions.

First ultrastructural data on the human tapeworm Taenia asiatica eggs by scanning and transmission electron microscopy (SEM, TEM).

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Galán-Puchades, M Teresa; Yang, Yichao; Marcilla, Antonio; Choe, Seongjun; Park, Hansol; Osuna, Antonio; Eom, Keeseon S

2016-09-01

Humans are definitive hosts of three species of the Taenia genus, namely Taenia solium, Taenia saginata and Taenia asiatica. The relative novelty of the latter explains the lack of knowledge concerning certain relevant aspects related to this parasite, such as its definite geographical distribution and whether its eggs can infect humans or not. So far, only the eggs of T. solium are known to be infective for humans, producing cysticercosis. Although eggs contain the infective stage, the oncosphere, there is a lack of research on the ultrastructure of eggs of human taeniids. We show, for the first time, the ultrastructure of eggs of T. asiatica by means of SEM and TEM analyses. We detected all the envelopes, namely the egg shell, vitelline layer, outer embryophoric membrane, embryophore, granular layer, basal membrane, oncospheral membrane and oncospheral tegument. Hooks surrounded by myofibrils and glycogen-like particles, the two types of secretory granules of the penetration glands, as well as several nuclei and mitochondria were also revealed in the oncospheres. In addition to the already known structures in eggs from other Taenia species, the presence of two types of small vesicles is described herein, possibly corresponding to exosomes and ectosomes because of their shape and size, which could participate in the host/parasite intercellular communication.

In Vitro Study of Taenia solium Postoncospheral Form.

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Nancy Chile

2016-02-01

Full Text Available The transitional period between the oncosphere and the cysticercus of Taenia solium is the postoncospheral (PO form, which has not yet been completely characterized. The aim of this work was to standardize a method to obtain T. solium PO forms by in vitro cultivation. We studied the morphology of the PO form and compared the expression of antigenic proteins among the PO form, oncosphere, and cysticerci stages.T. solium activated oncospheres were co-cultured with ten cell lines to obtain PO forms, which we studied at three stages of development--days 15, 30, and 60. A high percentage (32% of PO forms was obtained using HCT-8 cells in comparison to the other cell lines. The morphology was observed by bright field, scanning, and transmission electron microscopy. Morphology of the PO form changed over time, with the six hooks commonly seen in the oncosphere stage disappearing in the PO forms, and vesicles and microtriches observed in the tegument. The PO forms grew as they aged, reaching a diameter of 2.5 mm at 60 days of culture. 15-30 day PO forms developed into mature cysticerci when inoculated into rats. Antigenic proteins expressed in the PO forms are also expressed by the oncosphere and cysticerci stages, with more cysticerci antigenic proteins expressed as the PO forms ages.This is the first report of an in vitro production method of T. solium PO forms. The changes observed in protein expression may be useful in identifying new targets for vaccine development. In vitro culture of PO form will aid in understanding the host-parasite relationship, since the structural changes of the developing PO forms may reflect the parasite's immunoprotective mechanisms. A wider application of this method could significantly reduce the use of animals, and thus the costs and time required for further experimental investigations.

In Vitro Study of Taenia solium Postoncospheral Form.

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Chile, Nancy; Clark, Taryn; Arana, Yanina; Ortega, Ynes R; Palma, Sandra; Mejia, Alan; Angulo, Noelia; Kosek, Jon C; Kosek, Margaret; Gomez-Puerta, Luis A; Garcia, Hector H; Gavidia, Cesar M; Gilman, Robert H; Verastegui, Manuela

2016-02-01

The transitional period between the oncosphere and the cysticercus of Taenia solium is the postoncospheral (PO) form, which has not yet been completely characterized. The aim of this work was to standardize a method to obtain T. solium PO forms by in vitro cultivation. We studied the morphology of the PO form and compared the expression of antigenic proteins among the PO form, oncosphere, and cysticerci stages. T. solium activated oncospheres were co-cultured with ten cell lines to obtain PO forms, which we studied at three stages of development--days 15, 30, and 60. A high percentage (32%) of PO forms was obtained using HCT-8 cells in comparison to the other cell lines. The morphology was observed by bright field, scanning, and transmission electron microscopy. Morphology of the PO form changed over time, with the six hooks commonly seen in the oncosphere stage disappearing in the PO forms, and vesicles and microtriches observed in the tegument. The PO forms grew as they aged, reaching a diameter of 2.5 mm at 60 days of culture. 15-30 day PO forms developed into mature cysticerci when inoculated into rats. Antigenic proteins expressed in the PO forms are also expressed by the oncosphere and cysticerci stages, with more cysticerci antigenic proteins expressed as the PO forms ages. This is the first report of an in vitro production method of T. solium PO forms. The changes observed in protein expression may be useful in identifying new targets for vaccine development. In vitro culture of PO form will aid in understanding the host-parasite relationship, since the structural changes of the developing PO forms may reflect the parasite's immunoprotective mechanisms. A wider application of this method could significantly reduce the use of animals, and thus the costs and time required for further experimental investigations.

Construction and Identification of a Recombinant Plasmid Encoding Echinococcus granulosus Oncosphere Antigen (EG95Abstract Background: Cystic echinococcosis (CE, as a zoonotic disease cause to health threat and economic losses. Despite implemented cont

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Nahideh MAZAHERI

2017-12-01

Full Text Available AbstractBackground: Cystic echinococcosis (CE, as a zoonotic disease cause to health threat and economic losses. Despite implemented control programs, few countries have been able to decrease or eliminate this infection. Vaccination of the intermediate host offers an additional strategy to control the parasite transmission and EG95 antigen is considered more than the others in the vaccine issue. According to the high protection induced by the EG95 recombinant vaccine, this study was designed to construct recombinant plasmid formulation of EG95 antigen.Methods: In 2015, the Echinococcus granulosus eggs were recovered from an infected dog in Parasitological laboratory of Tarbiat Modares University in Tehran, Iran. Following hatching, the oncospheres of E. granulosus were activated to increase the presence of the desired mRNA. The extracted mRNA was transcribed to the cDNA which used as template in RT-PCR. Then the EG95 gene cloned into pET28a vector and the recombinant plasmids expression was  investigated in prokaryotic and eukaryotic cells.Results:  The recombinant plasmid encoding EG95 antigen was successfully constructed and identified by PCR, restriction enzyme digestion and sequencing. In vitro expression of the EG95 antigen was confirmed in prokary­otic and eukaryotic systems by SDS-PAGE and western blotting analysis.Conclusion: Because of potential advantages of DNA vaccines, including ability to induce long-term immune responses, low production cost and stability in different temperatures, this study carried out to construct the EG95 gene into a vector. This recombinant vector can be evaluated in further studies as a DNA vaccine may provide new prospects for the development of a vaccine against cystic hydatid disease.

c-orfi and Tnpnin crnssiceps Larvae I Department of Pharmaceutical ...

African Journals Online (AJOL)

satisfactorily. The putative first intermediate host is a ground-dwelling coprophagous arthropod that ingests oncospheres and accommodates the development of larvae. It is presumed that vertebrates become hosts for the larvae after ingestion of infected arthropods. This is followed by penetration of the small intestine by a.

A new anoplocephalid (Cestoda: Cyclophyllidea) from Gallotia atlantica (Reptilia, Lacertidae) in the Canary Islands, Spain.

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Foronda, Pilar; Abreu-Acosta, Néstor; Casanova, Juan Carlos; Ribas, Alexis; Valladares, Basilio

2009-06-01

A new species of anoplocephalid cestode is described from Gallotia atlantica (Reptilia, Lacertidae) on the Canary Islands, Spain. Oochoristica feliui n. sp. belongs to the group of Oochoristica spp. having circular suckers and fewer than 25 testes in a single cluster: O. lygosomae Burt, 1933; O. lygosomatis Skinker, 1935; O. elongata Dupouy et Kechemir, 1973; O. jonnesi Bursey, McAllister and Freed, 1997; O. junkea Johri, 1950; O. macallisteri Bursey and Goldberg, 1996; O. novaezelandae Schmidt and Allison, 1985; O. parvogenitalis Dupouy and Kechemir, 1973; and O. sobolevi (Spasskii, 1948) Spaskii, 1951. Oochoristica feliui n. sp. differs from these species in a variety of characters such as the number of proglottids, size of the scolex and suckers, presence of a neck, size and shape of the ovary (divided into 5-6 lobules), ovoid shape of the vitelline gland, a spined cirrus, size of eggs, oncosphere and oncosphere hooks, and the presence of 2 osmoregulatory canals. Gallotia spp. are endemic to the Canary Islands. This is the first report of an adult cestode species in this lizard.

Early intrauterine embryonic development in Khawia sinensis Hsu, 1935 (Cestoda, Caryophyllidea, Lytocestidae), an invasive tapeworm of carp (Cyprinus carpio): an ultrastructural study

Czech Academy of Sciences Publication Activity Database

Bruňanská, M.; Mackiewicz, J. S.; Młocicki, D.; Świderski, Z.; Nebesářová, Jana

2012-01-01

Roč. 110, č. 2 (2012), s. 1009-1017 ISSN 0932-0113 Institutional support: RVO:60077344 Keywords : ATRACTOLYTOCESTUS-HURONENSIS ANTHONY * WENYONIA-VIRILIS WOODLAND * DIDYMOBOTHRIUM-RUDOLPHII MONTICELLI * FARMED COMMON CARP * ONCOSPHERAL ENVELOPES * VITELLOGENESIS * PARASITE * CESTOIDEA * EUCESTODA Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 2.852, year: 2012 http://link.springer.com/content/pdf/10.1007%2Fs00436-011-2590-2

Dissecting and Targeting Latent Metastasis

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2015-09-01

underwent a rapid decrease in proliferation, as determined by dye retention (Figures 2G ), whereas the parental line showed little (H2087) or no...decrease in the ability to seed the brain (Figure 3G ). The ability of tumor-initiating cells to form colonies (oncospheres) in suspension provides a...Handgretinger, Human lymphoid and myeloid cell development in NOD/LtSz-scid IL2R gamma null mice engrafted with mobilized human hemopoietic stem

Atypical outbreak of acute coenurosis by Taenia multiceps in a sheep flock.

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Pintus, Davide; Varcasia, Antonio; Dessì, Giorgia; Tamponi, Claudia; Manunta, Maria Lucia; Carboni, Giovanni Antonio; Cancedda, Maria Giovanna; Ligios, Ciriaco; Scala, Antonio

2018-06-01

Herein, we examined the brain of adult ewes and lambs less than 30 days old which were found affected by neurological signs in a flock located in Sardinia (Italy). Gross anatomo-pathological examination of all brains of the animals revealed multiple linear reddish-yellow foci of necrotic purulent inflammation due to oncosphere migration. Histologically, we confirmed a multifocal pyo-granulomatous meningo-encephalitis both in ewes and in lambs, confirming acute coenurosis. Morphological examination and DNA sequencing identified the Taenia multiceps we isolated as Tm1 strain. This report describes for the first time a natural acute coenurosis infection in suckling lambs under 30 days of age.

LncMAPK6 drives MAPK6 expression and liver TIC self-renewal.

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Huang, Guanqun; Jiang, Hui; He, Yueming; Lin, Ye; Xia, Wuzheng; Luo, Yuanwei; Liang, Min; Shi, Boyun; Zhou, Xinke; Jian, Zhixiang

2018-05-15

Liver tumor initiating cells (TICs) have self-renewal and differentiate capacities, and largely contribute to tumor initiation, metastasis and drug resistance. MAPK signaling is a critical pathway in many biological processes, while its role in liver TICs hasn't been explored. Online-available dataset was used for unbiased screening. Liver TICs were examined CD133 FACS or oncosphere formation. TIC self-renewal was detected by oncosphere formation and tumor initiation assay. LncRNA function was detected by loss of function or gain of function assays. The molecular mechanism of lncRNA was explored by RNA pulldown, RNA immunoprecipitation, ChIP, western blot and double FISH. Here, we examined the expression profiles of MAPK components (MAPKs, MAP2Ks, MAP3Ks, MAP4Ks), and found MAPK6 is most highly expressed in liver cancer samples. Moreover, a divergent lncRNA (long noncoding RNA) of MAPK6, termed lncMAPK6 here, is also overexpressed along with liver tumorigenesis. LncMAPK6 promotes liver tumor propagation and TIC self-renewal through MAPK6. LncMAPK6 interacts with and recruits RNA polymerase II to MAPK6 promoter, and finally activates the transcription of MAPK6. Through MAPK6 transcriptional regulation, lncMAPK6 drives MARK signaling activation. LncMAPK6-MAPK6 pathway can be used for liver TIC targeting. Altogether, lncMAPK6 promotes MARK signaling and the self-renewal of liver TICs through MAPK6 expression. MAPK6 was the most highly expressed MAPK component in liver cancer and liver TICs and lncMAPK6 participated in the transcriptional regulation of MAPK6in cis. This work revealed the importance role of MAPK signaling in liver TIC self-renewal and added a new layer for liver TIC and MAPK6 expression regulation.

Excretory/secretory-products of Echinococcus multilocularis larvae induce apoptosis and tolerogenic properties in dendritic cells in vitro.

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Justin Komguep Nono

Full Text Available BACKGROUND: Alveolar echinococcosis, caused by Echinococcus multilocularis larvae, is a chronic disease associated with considerable modulation of the host immune response. Dendritic cells (DC are key effectors in shaping the immune response and among the first cells encountered by the parasite during an infection. Although it is assumed that E.multilocularis, by excretory/secretory (E/S-products, specifically affects DC to deviate immune responses, little information is available on the molecular nature of respective E/S-products and their mode of action. METHODOLOGY/PRINCIPAL FINDINGS: We established cultivation systems for exposing DC to live material from early (oncosphere, chronic (metacestode and late (protoscolex infectious stages. When co-incubated with Echinococcus primary cells, representing the invading oncosphere, or metacestode vesicles, a significant proportion of DC underwent apoptosis and the surviving DC failed to mature. In contrast, DC exposed to protoscoleces upregulated maturation markers and did not undergo apoptosis. After pre-incubation with primary cells and metacestode vesicles, DC showed a strongly impaired ability to be activated by the TLR ligand LPS, which was not observed in DC pre-treated with protoscolex E/S-products. While none of the larvae induced the secretion of pro-inflammatory IL-12p70, the production of immunosuppressive IL-10 was elevated in response to primary cell E/S-products. Finally, upon incubation with DC and naïve T-cells, E/S-products from metacestode vesicles led to a significant expansion of Foxp3+ T cells in vitro. CONCLUSIONS: This is the first report on the induction of apoptosis in DC by cestode E/S-products. Our data indicate that the early infective stage of E. multilocularis is a strong inducer of tolerance in DC, which is most probably important for generating an immunosuppressive environment at an infection phase in which the parasite is highly vulnerable to host attacks. The

Linc00210 drives Wnt/β-catenin signaling activation and liver tumor progression through CTNNBIP1-dependent manner.

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Fu, Xiaomin; Zhu, Xiaoyan; Qin, Fujun; Zhang, Yong; Lin, Jizhen; Ding, Yuechao; Yang, Zihe; Shang, Yiman; Wang, Li; Zhang, Qinxian; Gao, Quanli

2018-03-14

Liver tumor initiating cells (TICs) have self-renewal and differentiation properties, accounting for tumor initiation, metastasis and drug resistance. Long noncoding RNAs are involved in many physiological and pathological processes, including tumorigenesis. DNA copy number alterations (CNA) participate in tumor formation and progression, while the CNA of lncRNAs and their roles are largely unknown. LncRNA CNA was determined by microarray analyses, realtime PCR and DNA FISH. Liver TICs were enriched by surface marker CD133 and oncosphere formation. TIC self-renewal was analyzed by oncosphere formation, tumor initiation and propagation. CRISPRi and ASO were used for lncRNA loss of function. RNA pulldown, western blot and double FISH were used to identify the interaction between lncRNA and CTNNBIP1. Using transcriptome microarray analysis, we identified a frequently amplified long noncoding RNA in liver cancer termed linc00210, which was highly expressed in liver cancer and liver TICs. Linc00210 copy number gain is associated with its high expression in liver cancer and liver TICs. Linc00210 promoted self-renewal and tumor initiating capacity of liver TICs through Wnt/β-catenin signaling. Linc00210 interacted with CTNNBIP1 and blocked its inhibitory role in Wnt/β-catenin activation. Linc00210 silencing cells showed enhanced interaction of β-catenin and CTNNBIP1, and impaired interaction of β-catenin and TCF/LEF components. We also confirmed linc00210 copy number gain using primary hepatocellular carcinoma (HCC) samples, and found the correlation between linc00210 CNA and Wnt/β-catenin activation. Of interest, linc00210, CTNNBIP1 and Wnt/β-catenin signaling targeting can efficiently inhibit tumor growth and progression, and liver TIC propagation. With copy-number gain in liver TICs, linc00210 is highly expressed along with liver tumorigenesis. Linc00210 drives the self-renewal and propagation of liver TICs through activating Wnt/β-catenin signaling. Linc00210

[Molecular characterization of Echinococcus granulosus isolates obtained from different hosts].

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Erdoğan, Emrah; Özkan, Bora; Mutlu, Fatih; Karaca, Serkan; Şahin, İzzet

2017-01-01

Echinococcus granulosus is a parasite that can be seen throughout the world. So far, five species of genus Echinococcus have been identified as parasite in people: E.granulosus, E.multilocularis, E.vogeli, E.oligarthrus, E.shiquicus. Larval (metacestod) form of parasite settles in internal organs of hoofed animals (cattle, goats, pigs, horses, sheep, etc.) and human; the adult form is found in small intestine of final host, canine. Disease caused by parasite called as "Cystic echinococcosis" (CE) is an important health problem and causes economic losses in many countries including our country that livestock is common. Infective eggs cause infections in intermediate hosts by taking oral way and rarely inhalation. Received egg opens in the stomach and intestines of intermediate host and oncosphere is released. Oncosphere quickly reaches the lamina propria of the villus epithelium by its histolytic enzymes and hooks. It usually transported from here to the liver and lungs, less frequently, muscle, brain, spleen, kidney and to other organs through the veins. By molecular studies, five species have been validated taxonomically and 10 different variants or strains of E.granulosus have been identified. Host and developmental differences between strains may negatively affect control studies and fight against the parasite. This study aimed to determinate E.granulosus strains obtained from cyst material of different intermediate hosts from different regions of Turkey by molecular methods. In the study, 25 human, 8 cattle, 6 sheep and 2 goat cysts material has been collected. Total genomic DNA was isolated from protoscoleces in cyst fluid and analyzed by PCR with COX-1 (L) and COX-1 (S) genes specific primers. DNA sequence analysis for each PCR product has been made. DNA sequence analysis results evaluated phylogenetically by MEGA analyze and BLAST software. As a result of this study, all isolates were identified as E.granulosus sensu stricto (G1) by DNA sequence analysis. CE