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A new micromethod for the detection of HL-A antigens on cultured human tumor cells.

A rapid microcytotoxicity assay for the detection of HL-A antigens on tissue culture cells derived from human solid tumors is described. Tumor cells were prelabeled with 125Iododeoxyuridine. Isotopically labeled tumor cells were reacted with up to 37 highly selected HL-A antisera and diluted rabbit complement. Results of the HL-A typing of nine human tumor cell lines are reported. Three melanoma cell lines showed individually distinct HL-A profiles at the first HL-A locus which agreed with the antigenic pattern of the tumor donor's autologous lymphocytes. Less reactivity was noted with HL-A antisera defining second locus specificities on the three melanoma cell lines, whereas some other cell lines showed more HL-A reactions than required to present a "full house". This method obviates the necessity for visually enumerating residual tumor target cells.

Adenocarcinoma

A study of the relationship of the dental midline to the facial median line.

This report presents the results of an original investigation designed to determine (1) the prevalence in the natural dentition of a maxillary midline located in the exact middle of the mouth using the philtrum as the most reliable guide and (2) the percentage of people in whom the maxillary and mandibular midlines precisely coincide with each other. Results indicate that the midline is situated in the exact middle of the mouth in approximately 70% of people and that the maxillary and mandibular midlines fail to coincide in almost three fourths of the population.

Adolescent

Multiplexed Dual-Color Fluorescence-Based Distinction Between Nuclear Trapping and Translocation of FOXO3.

FOXO3 is a transcription factor that mainly exerts its functions in the cell nucleus. The amino acid sequence of FOXO3 contains a nuclear localization sequence (NLS) and a nuclear export sequence (NES) allowing for nuclear/cytoplasmic shuttling that plays an important role in regulating FOXO3 activity. Nuclear accumulation of FOXO3 proteins can be the result of translocation to the nucleus triggered by upstream regulatory input or trapping of FOXO3 within the nucleus through the inhibition of its nuclear export via the receptor CRM1. In order to distinguish these two modes of FOXO3 activation, we have generated a multiplexed assay. The development of this platform includes a reporter cell line that monitors CRM1 activity by using RFP-labeled HIV-1 Rev. protein with a strong heterologous NES. Simultaneously, the intracellular localization of FOXO3 can be monitored by a second cell line stably expressing GFP-FOXO3. Here we describe a detailed protocol on how to co-culture these reporter cell lines and use them to interrogate compound-induced FOXO3 activation in order to understand the mode of action.

Forkhead Box Protein O3

Infection of cardiac suture line after ventricular aneurysmectomy. Report of two cases.

Two cases of sepsis following ventricular aneurysmectomy are presented. In both, the source of sepsis was an infected caridac suture line in which Teflon felt strips were used to reinforce the closure. One patient had a pseudoaneurysm and a chronic empyema of the left side of the chest, and the second had a ventriculocutaneous fistula. Early operation with removal of the foreign body is warranted in this major complication of a ventricular aneurysmectomy.

Candidiasis

Comparison of reports on lung function tests made by chest physicians with those made by a simple computer program.

The numerical results of 60 sets of pulmonary function tests were submitted to five consultant chest physicians for independent reporting. The chest physicians' reports were compared with the reports generated by the on-line computer reporting system in routine use in the pulmonary function laboratory at Westminster Hospital. There was good agreement between the reports of the chest physicians among themselves and with the computer. The individual reports were compared with the consensus opinions of the physicians and the computer. The computer's decisions differed from the consensus opinion in 4.6% of instances. The comparable figures for the five physicians were respectively 2.9%, 2.9%, 2.9%, 4.0%, and 4.6%. Decisions differing from the consensus were due to mistakes or actual sustained disagreements of opinion. The physicians made more mistakes than the computer even though they were performing under test conditions. They made up for this, however, by producing fewer genuine disagreements from the consensus opinions. It is concluded that in routine day-to-day practice the computer report will be as consistently useful as the chest physicians' reports and more immediately available.

Airway Obstruction

Evaluation of single-agent therapy in human colorectal tumour xenografts.

The response of 6 human colorectal tumour xenografts to 7 cytotoxic agents have been established. Tumour responses have been quantified by growth inhibition, and the time taken for 3H-thymidine fractional incorporation (TFI) to recover to the control value after treatment. The chemosensitivity of each tumour line to a spectrum of agents was individual, and no pattern of response which would allow prediction of individual agent efficacy was apparent. Cyclophosphamide, methyl-CCNU and 5-fluorouracil produced marked growth inhibition in individual tumour lines, whereas actinomycin-D, cis-dichlorodiammine platinum, doxorubicin and pentamethylmelamine showed little activity. Data presented agree with clinical evaluation for single-agent therapy. The uptake and incorporation of radiolabelled 5-fluorouracil into 4 tumour lines is reported. No marked differences between 3 FU-insensitive lines and 1 sensitive line have been observed.

Animals

On-line computer storage, retrieval, and reporting of coded angiographic data.

Interpretations of angiographic images have been coded and stored using an on-line computer terminal for seven years. Decoded angiographic information is incorporated in computer-generated reports which are printed on demand after completion of cardiac catheterization procedures. Currently, almost 5,500 cases are stored in an off-line data base which has been designed to help identify patterns with prognostic potential. Also, the program has helped standardize angiographic nomenclature, saved much clerical time, and virtually eliminated clerical errors. The program interacts with other software in the hospital, avoiding repetitious entries. The advantages and shortcomings of the program and commonly used approaches to computer storage/retrieval of radiographic information are described.

Angiography

Development of a cell-based nanoluciferase reporter system for high-throughput screening of HBV cccDNA inhibitors.

Hepatitis B virus (HBV) persistence is sustained by the viral covalently closed circular DNA (cccDNA) minichromosome, which remains a major barrier to curative antiviral therapies. The lack of reliable quantitative cccDNA detection methods and surrogate markers has hindered efforts to target cccDNA in antiviral high-throughput screening (HTS). Here, we established a novel inducible cccDNA-dependent nanoluciferase (NLuc) reporter cell line, designated HepBLE12, by inserting an in-frame 11-amino acid split-NLuc HiBiT tag into the precore (pC) coding region of an HBV transgene. The resulting 1.3-kDa HiBiT tag on pC serves as the detection module of the split NLuc system, generating quantitative luminescence upon high-affinity complementation with the cognate 18-kDa LgBiT subunit in cell lysates. Notably, the HiBiT assay enables direct detection of intracellular HiBiT-pC protein rather than secreted HBeAg, providing a reporter signal more closely linked to cccDNA activity. HepBLE12&#x202f;cells exhibited inducible and robust viral DNA replication, and the cccDNA-dependent HiBiT signal was validated under diverse experimental conditions that modulate cccDNA formation or transcription. We further miniaturized the assay to a 384-well format and optimized key parameters following standard HTS assay development practices. The assay was successfully automated and demonstrated excellent performance in a multi-day variability study and a pilot screen, with signal-to-background (S/B)&#x202f;&#x2248;&#x202f;9, coefficient of variance (CV)&#x202f;<&#x202f;10%, and average Z-factor value of 0.74, exceeding canonical HTS quality benchmarks. Together, the HepBLE12 cell-based HTS platform provides a robust and practical tool for identifying inhibitors targeting HBV cccDNA.

Hepatitis B virus

Pigment in the lining of nasopalatine duct cysts: report of two cases.

In a review of 20 nasopalatine duct cysts, two cases were encountered with a pigmented epithelial lining. In the literature we have found one similar case only. The pigment in our cases was shown to be melanin. The authors suggest that the melanin-containing epithelial cells are derived from Jacobson's organs and, therefore, most likely should be considered olfactory epithelium.

Adult

A suite of enhancer AAVs and transgenic mouse lines for genetic access to cortical cell types.

The mammalian cortex is comprised of cells classified into types according to shared properties. Defining the contribution of each cell type to the processes guided by the cortex is essential for understanding its function in health and disease. We use transcriptomic and epigenomic cortical cell-type taxonomies from mouse and human to define marker genes and putative enhancers and create a large toolkit of transgenic lines and enhancer adeno-associated viruses (AAVs) for selective targeting of cortical cell populations. We report creation and evaluation of fifteen transgenic driver lines, two reporter lines, and >1,000 different enhancer AAV vectors covering most subclasses of cortical cells. The tools reported here have been made publicly available, and along with the scaled process of tool creation, evaluation, and modification, they will enable diverse experimental strategies toward understanding mammalian cortex and brain function.

Animals

[Radiation effects in brain and spinal cord (author's transl)].

Radiation sensitivity of both the brain and spinal cord in prenatal and postnatal stages, in infancy and adult age is represented also in consideration of a combined treatment with methotrexate. In adults, application of important doses of high-energy radiation increases the risk of injurious effects to the central nervous system. If the spinal cord is involved, more than 60% of the radiolesions have a progredient course ending with death. The pathogenesis and disposing factors are referred to, and the incidence of radiation necrosis with regard to age and sex, the degrees of injury and their frequence within different ranges of dosage are analyzed on the basis of data from universal literature. An examination of "tolerance doses" for the spinal cord is made by means of Strandquist-diagrams and of the Ellis-formula. The slopes of regression lines are reported for various "degrees of response" in skin, brain and spinal cord following radiation therapy. In the Strandquist-diagram, slopes of regression lines are dependent on the "degree of response", flattening if skin and spinal cord are affected by radiation in the same degree, necroses having the same slope for both the organs.

Age Factors

Ectopic and eutopic secretion of chorionic gonadotropin and its sub-nits in vitro: comparison of clonal strains from carcinomas of lung and placenta.

We compared rates of secretion in vitro of human chorionic gonadotropin (HCG) and its subunits alpha and beta by established clonal cell lines of a bronchogenic carcinoma (ChaGo) and a choriocarcinoma (JEG). Clones showing the highest secretion rates of either HCG or its subunits were studied: ChaGo-K1, a new clonal strain, and ChaGo-C5 and JEG-3, two previously reported clonal lines. Cells were grown under identical conditions in the same laboratory. Hormone and subunit concentrations were measured by radioimmunoassays. ChaGo-K1 and ChaGo-C5 secreted only alpha-subunit whereas JEG-3 secreted only HCG. Average peak secretion rates in picomoles/day/mg protein were: for ChaGo-K1, HCG less than 0.3, alpha=290, and beta less than 0.5; for ChaGo-C5, HCG less than 0.3, alpha=21, and beta less than 0.5; and for JEG-3, HCG=18, alpha less than 0.7, and beta less than 0.5. The ChaGo-K1 secretion rate of alpha was greater than that of any of out previously reported ChaGo clones. Significant quantities of estradiol and progesterone accumulated in the media of all three cell lines; however, only JEG-3 secreted detectable quantities of placental lactogen. Thus under identical culture conditions, a bronchogenic carcinoma clonal line secreted only alpha-subunit, whereas a choriocarcinoma line secreted only HCG; these findings implied major differences in cellular control mechanisms. Moreover, the ectopic secretions of alpha exceeded the eutopic trophoblastic secretion of HCG, which suggested that in certain cases ectopic protein production may be even more efficient than nonectopic production.

Carcinoembryonic Antigen

Establishment of an Epstein-Barr virus-determined nuclear antigen-negative human B-cell line from acute lymphoblastic leukemia.

A human B-cell line designated as BALL-1 was established from the peripheral blood of a patient with acute lymphoblastic leukemia (ALL). Neither Epstein-Barr virus (EBV) particles nor EBV-determined nuclear antigen (EBNA) was detectable. The morphologic and growth characteristics were clearly distinct from those of numerous EBV-positive lymphoblastoid cell lines previously reported. BALL-1 cells probably originated from the donor's leukemia cells as judged from their cytogenetic, morphologic, and surface features. The BALL-1 line was the first EBNA-negative B-cell line established from ALL.

Antigens, Viral

Human leukemic "null" cell line (NALL-1).

A human lymphoblast cell line, NALL-1, was established from the peripheral blood of a patient with acute lymphoblastic leukemia (ALL). NALL-1 cells had neither properties of T and B cells nor Epstein-Barr virus (EBV). Many characteristics of the NALL-1 line were distinct from those of numerous EBV-positive lymphoblastoid cell lines previously reported. NALL-1 cells are considered to have originated from the donor's leukemic cells on the basis of their cytogenetic, morphologic and functional features. The NALL-1 line is the first human leukemic "null" cell line derived from ALL. The significance of this cell line is disscussed.

B-Lymphocytes

Telephone line transmission of rectilinear radionuclide scans. Report of a pilot project.

The General Electric-Elscint system for telephone line transmission of rectillinear scans has been tested and with certain limitations found to be acceptable in terms of diagnostic accuracy for many scan procedures. The system tested had some logistic disadvantage requiring batch processing of the scans, and making quality control of the scans difficult with the one video data processor available at the reference centre.

Data Display

Iron-mediated post-transcriptional regulation in Toxoplasma gondii.

Iron is required to support almost all life; however, levels must be carefully regulated to maintain homeostasis. Although the obligate parasite Toxoplasma gondii requires iron, how it responds upon iron limitation has not been investigated. Here, we show that iron depletion triggers significant transcriptional changes in the parasite, including in iron-dependent pathways. We find that a subset of T. gondii transcripts contain stem-loop structures, which have been associated with post-transcriptional iron-mediated regulation in other cellular systems. We validate one of these (found in the 3' UTR of TGME49_261720) using a reporter cell line. We show that the presence of the stem-loop-containing UTR is sufficient to confer accumulation at the transcript and protein levels under low iron. This response is dose and time-dependent and is specific for iron. The accumulation of transcript is likely driven by an increased reporter mRNA stability under low iron. Interestingly, we find iron-mediated changes in mRNA stability in around 400 genes. To examine the potential mechanism of this stability, we tested aconitase interaction with mRNA in low iron and found 43 enriched transcripts, but no specific interaction with our reporter UTR. However, the endogenous UTR led to maintenance of protein levels and increased survival of the parasite under low iron. Our data demonstrate the existence of iron-mediated post-transcriptional regulation in Toxoplasma for the first time; and suggests iron-mediated regulation may be important to the parasite in low iron environments.

Toxoplasma