Detection of reverse transcriptase activity by enzyme-linked immunosorbent assay in human immunodeficiency virus type 1.

An enzyme-linked immunosorbent assay (ELISA) using biotin-labelled oligo-dT primer and digoxigenin (Dig)-dUTP was designed to measure the reverse transcriptase (RT) activity of human immunodeficiency virus type 1 (HIV-1). The ELISA system involves the selective detection step of a newly synthesized cDNA by two specific bindings, biotin-streptavidin binding and alkaline phosphatase (AP)-conjugated anti-Dig-Dig binding, and the enzymatic amplification step to increase coloring generated by AP. This method was used to measure the activity of RT in the culture supernatants of peripheral leukocytes obtained from four anti-HIV-1-positive persons cocultivated with those from four anti-HIV-1-negative persons. RT activity was detected in all of four anti-HIV-1-positive culture supernatants but not in those cultivated with anti-HIV-1-negative supernatants alone. Thus, our improved ELISA for detection of HIV-1 appears to be sensitive enough and useful for routine laboratory work. This non-radioactive method will also be useful for detecting other retroviruses and for screening of RT inhibitors.</p

Neuroendocrine Carcinoma of the Stomach: A Case Study

Gastric neuroendocrine carcinomas are rare and have a poor prognosis, and the diagnostic criteria for this disease have recently changed. We herein report a case of sporadic gastric neuroendocrine carcinoma. A 75-year-old man was referred to our hospital with epigastric pain. Endoscopic examination revealed a localized ulcerative lesion (diameter, 4 cm) at the upper stomach. The diagnosis on biopsy was neuroendocrine carcinoma. Total gastrectomy with D2 lymphadenectomy, splenectomy, and cholecystectomy was performed. Pathologically, the tumor infiltrated the subserosal layer, and 6/49 lymph nodes were involved. The tumor was uniform in shape and arranged in a rosette-like structure to form solid nests, with medium-sized, round-to-cuboid-shaped tumor cells and intense mitosis 46/10 HPF. It was positive for synaptophysin and chromogranin A, and the Ki-67 labeling index was 70–80%. The diagnosis of neuroendocrine carcinoma was made according to the WHO 2010 criteria. The patient was followed up for three years without recurrence

H, C, and N isotopic compositions of Hayabusa category 3 organic samples

Since isotopic ratios of H, C, and N are sensitive indicators for determining extraterrestrial organics, we have measured these isotopes of Hayabusa category 3 organic samples of RB-QD04-0047-02, RA-QD02-0120, and RB-QD04-0001 with ion imaging using a NanoSIMS ion microprobe. All samples have H, C, and N isotopic compositions that are terrestrial within errors (approximately ±50‰ for H, approximately ±9‰ for C, and approximately ±2‰ for N). None of these samples contain micrometer-sized hot spots with anomalous H, C, and N isotopic compositions, unlike previous isotope data for extraterrestrial organic materials, i.e., insoluble organic matters (IOMs) and nano-globules in chondrites, interplanetary dust particles (IDPs), and cometary dust particles. We, therefore, cannot conclude whether these Hayabusa category 3 samples are terrestrial contaminants or extraterrestrial materials because of the H, C, and N isotopic data. A coordinated study using microanalysis techniques including Fourier transform infrared spectrometry (FT-IR), time-of-flight secondary ion mass spectrometry (ToF-SIMS), NanoSIMS ion microprobe, Raman spectroscopy, X-ray absorption near edge spectroscopy (XANES), and transmission electron microscopy/scanning transmission electron microscopy (TEM/STEM) is required to characterize Hayabusa category 3 samples in more detail for exploring their origin and nature.This research was supported by the JSPS Strategic Fund for Strengthening Leading-edge Research and Development to the JAMSTEC

H, C, and N isotopic compositions of Hayabusa category 3 organic samples

Since isotopic ratios of H, C, and N are sensitive indicators for determining extraterrestrial organics, we have measured these isotopes of Hayabusa category 3 organic samples of RB-QD04-0047-02, RA-QD02-0120, and RB-QD04-0001 with ion imaging using a NanoSIMS ion microprobe. All samples have H, C, and N isotopic compositions that are terrestrial within errors (approximately ±50‰ for H, approximately ±9‰ for C, and approximately ±2‰ for N). None of these samples contain micrometer-sized hot spots with anomalous H, C, and N isotopic compositions, unlike previous isotope data for extraterrestrial organic materials, i.e., insoluble organic matters (IOMs) and nano-globules in chondrites, interplanetary dust particles (IDPs), and cometary dust particles. We, therefore, cannot conclude whether these Hayabusa category 3 samples are terrestrial contaminants or extraterrestrial materials because of the H, C, and N isotopic data. A coordinated study using microanalysis techniques including Fourier transform infrared spectrometry (FT-IR), time-of-flight secondary ion mass spectrometry (ToF-SIMS), NanoSIMS ion microprobe, Raman spectroscopy, X-ray absorption near edge spectroscopy (XANES), and transmission electron microscopy/scanning transmission electron microscopy (TEM/STEM) is required to characterize Hayabusa category 3 samples in more detail for exploring their origin and nature.This research was supported by the JSPS Strategic Fund for Strengthening Leading-edge Research and Development to the JAMSTEC

ToF-SIMS analysis of carbonaceous particles in the sample catcher of the Hayabusa spacecraft

Three carbonaceous category 3 particles (RA-QD02-0180, RB-QD04-0037-01, and RB-QD04-0047-02) returned in the sample catcher from the Hayabusa spacecraft were analyzed by time of flight-secondary ion mass spectrometry (ToF-SIMS) to establish an analytical procedure for determination of their origins. By the different analytical schemes, the three particles gave distinct elemental and molecular ions, in which the organic carbons commonly appear to be associated with nitrogen, silicon, and/or fluorine. The particles could be debris of silicon rubber and fluorinated compounds and are therefore man-made artifacts rather than natural organic matter

Sequential analysis of carbonaceous materials in Hayabusa-returned samples for the determination of their origin

Preliminary results of the analyses of five carbonaceous materials (particle size of approximately 50 μm) from the Hayabusa spacecraft sample catcher, including their texture, chemistry, and chemical/isotopic compositions, are summarized. The carbonaceous particles underwent sequential analysis using a series of microanalytical instruments located at several research institutes and universities. Collected particles were initially classified into four categories: two categories containing extraterrestrial silicate particles, one category containing metal and quartz particles consistent with contamination from the sample catcher or sample manipulation tools, and a final category containing carbonaceous particles. Analysis of this final category was the main focus of this study. Through examination of the carbonaceous materials, the appropriate analytical processes for sample transportation and handling were optimized to minimize sample damage and terrestrial contamination. Particles were investigated by transmission electron microscopy/scanning transmission electron microscopy, and Ca-carbonate inclusions were found in one particle. In a different particle, a heterogeneous distribution of silicon in a uniform C, N, and O matrix was found. Though further analysis is required for a strict determination of particle origin, the differences in the microstructure and elemental distribution of the carbonaceous particles suggest multiple origins.TEM analyses were performed at JEOL Ltd. XANES analysis was conducted in UVSOR Facility in Institute for Molecular Science, supported by Nanotechnology Platform Program (Molecule and Material Synthesis) of the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan

X-ray absorption near edge structure spectroscopic study of Hayabusa category 3 carbonaceous particles

Analyses with a scanning transmission x-ray microscope (STXM) using x-ray absorption near edge structure (XANES) spectroscopy were applied for the molecular characterization of two kinds of carbonaceous particles of unknown origin, termed category 3, which were collected from the Hayabusa spacecraft sample catcher. Carbon-XANES spectra of the category 3 particles displayed typical spectral patterns of heterogeneous organic macromolecules; peaks corresponding to aromatic/olefinic carbon, heterocyclic nitrogen and/or nitrile, and carboxyl carbon were all detected. Nitrogen-XANES spectra of the particles showed the presence of N-functional groups such as imine, nitrile, aromatic nitrogen, amide, pyrrole, and amine. An oxygen-XANES spectrum of one of the particles showed a ketone group. Differences in carbon- and nitrogen-XANES spectra of the category 3 particles before and after transmission electron microscopic (TEM) observations were observed, which demonstrates that the carbonaceous materials are electron beam sensitive. Calcium-XANES spectroscopy and elemental contrast mapping identified a calcium carbonate grain from one of the category 3 particles. No fluorine-containing molecular species were detected in fluorine-XANES spectra of the particles. The organic macromolecular features of the category 3 particles were distinct from commercial and/or biological ‘fresh (non-degraded)’ polymers, but the category 3 molecular features could possibly reflect degradation of contaminant polymer materials or polymer materials used on the Hayabusa spacecraft. However, an extraterrestrial origin for these materials cannot currently be ruled out

ToF-SIMS analysis of carbonaceous particles in the sample catcher of the Hayabusa spacecraft

Three carbonaceous category 3 particles (RA-QD02-0180, RB-QD04-0037-01, and RB-QD04-0047-02) returned in the sample catcher from the Hayabusa spacecraft were analyzed by time of flight-secondary ion mass spectrometry (ToF-SIMS) to establish an analytical procedure for determination of their origins. By the different analytical schemes, the three particles gave distinct elemental and molecular ions, in which the organic carbons commonly appear to be associated with nitrogen, silicon, and/or fluorine. The particles could be debris of silicon rubber and fluorinated compounds and are therefore man-made artifacts rather than natural organic matter

Diagnostic Performance of 11C-choline PET/CT and FDG PET/CT in Prostate Cancer

We compared 11C-choline and FDG PET/CT scan findings for the staging and restaging of prostate cancer. Twenty Japanese prostate cancer patients underwent 11C-choline and FDG PET/CT before (n=5) or after (n=15) treatment. Using a five-point scale, we compared these scanning modalities regarding patient- and lesion-based diagnostic performance for local recurrence, untreated primary tumor, and lymph node and bony metastases. Of the 20 patients, documented local lesions, and node and bony metastases were present in 11 (55.0%), 9 (45.0%), and 13 (65.0%), respectively. The patient-based sensitivity/specificity/accuracy/area under the receiver-operating-characteristic curve (AUC) values for 11C-choline-PET/CT for diagnosing local lesions were 90.9% /100%/ 95.0% / 1.0, whereas those for FDG-PET/CT were 45.5% /100%/ 75.0% / 0.773. Those for 11C-choline-PET/CT for node metastasis were 88.9% /100%/ 95.0% / 0.944, and those for FDG-PET/CT were 44.4%/100%/75.0%/0.722. Those for 11C-choline-PET/CT for bone metastasis were 84.6%/100%/90.0%/0.951, and those for FDG-PET/CT were 76.9% /100%/ 85.0% / 0.962. The AUCs for local lesion and node metastasis differed significantly (p=0.0039, p=0.011, respectively). The lesion-based detection rates of 11C-choline compared to FDG PET/CT for local lesion, and node and bone metastases were 91.7% vs. 41.7%, 92.0% vs. 32.0%, and 94.8% vs. 83.0% (p=0.041, p=0.0030, p<0.0001), respectively. 11C-choline-PET/CT is more useful for the staging and restaging of prostate cancer than FDG-PET/CT in Japanese men

Tumor innate immunity primed by specific interferon-stimulated endogenous retroviruses.

Mesenchymal tumor subpopulations secrete pro-tumorigenic cytokines and promote treatment resistance1-4. This phenomenon has been implicated in chemorefractory small cell lung cancer and resistance to targeted therapies5-8, but remains incompletely defined. Here, we identify a subclass of endogenous retroviruses (ERVs) that engages innate immune signaling in these cells. Stimulated 3 prime antisense retroviral coding sequences (SPARCS) are oriented inversely in 3' untranslated regions of specific genes enriched for regulation by STAT1 and EZH2. Derepression of these loci results in double-stranded RNA generation following IFN-γ exposure due to bi-directional transcription from the STAT1-activated gene promoter and the 5' long terminal repeat of the antisense ERV. Engagement of MAVS and STING activates downstream TBK1, IRF3, and STAT1 signaling, sustaining a positive feedback loop. SPARCS induction in human tumors is tightly associated with major histocompatibility complex class 1 expression, mesenchymal markers, and downregulation of chromatin modifying enzymes, including EZH2. Analysis of cell lines with high inducible SPARCS expression reveals strong association with an AXL/MET-positive mesenchymal cell state. While SPARCS-high tumors are immune infiltrated, they also exhibit multiple features of an immune-suppressed microenviroment. Together, these data unveil a subclass of ERVs whose derepression triggers pathologic innate immune signaling in cancer, with important implications for cancer immunotherapy