ABSTRACT
Expression-library immunization has been proposed as an effective means to screen a large number of genes of the pathogen as candidate protective molecules. In this study, we examined the efficacy of expression-library immunization using a T. taeniaeformis rat model system. Total RNAs were isolated from the last 15 segments of adult T. taeniaeformis and poly A RNA was purified. cDNA library was produced using SuperScript Plasmid System, which contains a mammalian expression vector, pCMV*SPORT6. From about 3,500 clones examined, more than 800 clones were found to contain DNA fragments. About 200 clones were sequenced and the homology search was carried out. The blast search revealed that 29% of the expression genes were mitochondrial genes (rRNA; 17%, protein; 12%). Nuclear rRNA genes (10%), nuclear protein (9%) and genes from Escherichia coli were also detected. Forty-two percent of sequences did not show a significant similarity to any genes deposited in the public database. Rats were immunized with expression-library and injected orally with 1,000 T. taeniaeformis eggs. However the protective effect of expression-library vaccine was not confirmed.
Subject(s)
Animals , Antibodies, Helminth , Female , Gene Expression , Immunization , RNA, Nuclear , Rats , Taenia/genetics , Taeniasis/immunology , Vaccines, DNA/pharmacologyABSTRACT
PURPOSE: To report two cases of nasal type Natural Killer (NK)/T-cell lymphoma of the orbit. METHODS: (Case 1) A 42-year-old woman presented with a 3-month history of painful proptosis and eyelid swelling in the right eye. Under the diagnosis of inflammatory pseudotumor, she was treated with intravenous and oral steroids. She visited our clinic with a 3-day history of proptosis with decreased visual acuity. Computed tomography (CT) revealed a large infiltrative mass in the right orbit. Incisional biopsy was performed in the right lower lid. (Case 2) A 42-year-old man was referred for consultation with a 1-month history of fever of unknown origin. Two months previously uveitis had developed in the right eye and had been treated with topical steroid. CT revealed an infiltrative soft-tissue like mass in the left orbit. Incisional biopsy was performed in the orbital area. RESULTS: Histopathologically, a diffuse infiltration of atypical lymphocytes was observed with angiocentric pattern in both patients. The infiltrating cells were positive on immunohistochemical stains for CD3 and CD56. The tumor cells were negative for CD20 and CD30. In situ hybridization demonstrated Epstein-Barr virus-encoded nuclear RNA. A diagnosis was made with NK/T-cell lymphoma of nasal type associated with Epstein-Barr virus infection. CONCLUSIONS: NK/T-cell lymphoma of the orbit is rare and has poor prognosis. It is important to distinguish a NK/T-cell lymphoma from the pseudotumor or uveitis unresponsive to steroid therapy.
Subject(s)
Adult , Female , Humans , Biopsy , Coloring Agents , Diagnosis , Exophthalmos , Eyelids , Fever of Unknown Origin , Granuloma, Plasma Cell , Herpesvirus 4, Human , In Situ Hybridization , Lymphocytes , Lymphoma , Orbit , Prognosis , RNA, Nuclear , Steroids , Uveitis , Visual AcuityABSTRACT
Background. The presence of RNA in the cell nucleus is well known. However, a high resolution in situ hybridization evidence for the presence of RNA in some nuclear particles is still lacking. The aim of this work is to localize RNA in subnuclear particles using a novel ultrastructural in situ hybridization procedure. In this study, biotinylated genomic mouse DNA as a probe to localiza total RNA in the nuclei of mouse hepatocytes was used. Methods. The procedure is based on Paraformaldehyde fixation and embedding in lowicryl resin. Thin sections are mounted in formvar-coated gold grids. Hybridization is performed on non-denatured thin sections. DNA-RNA hybrids are detected with streptavidin-10 mm gold particles complex. By controlling the time of nick-translation during incorporation of biotin into the probe, labeling in the fibrillar portions of the nucleoplasm is obtained. More digested probes generate more labeling in the granular components. Nucleoli were similarly labeled. Results. As expected, no label was observed in the compact chromatin clumps. These results indicate that granular components as perichromatin granules in the nucleus contain more processed RNA than fibrillar portions. As a comparison, viral DNA sequences on denatured RNase-treated thin sections of adenovirus-2 (Ad-2)-infected human cells were detected. As previously reported, at late stages DNA was observed in the viral particles and surrounding nucleoplasm, where Ad-2 DNA is synthesized. Conclusions. The present procedure allows the study of intranuclear RNA distribution and will be useful fo the analysis of RNA processing in several types of cells