Previously, we established an easy and quick construction method for obtaining a stable and highly productive gene-amplified recombinant Chinese hamster ovary (CHO) cell line. With a gradual increase in methotrexate (MTX) concentration, gene-amplified cell pools had high and stable specific growth and production rates. Moreover, the phenotype of gene-amplified cells seemed to be affected by the location of the amplified gene in chromosomal DNA. We suspected that various kinds of gene-amplified cells might appear during the long-term selection to construct gene-amplified cell pools. To clarify the behavior of gene-amplified cell pools during a stepwise increase of MTX concentration, we isolated gene-amplified clones derived from gene-amplified cell pools. We compared the characteristics of isolated clones, such as the productivity of recombinant protein, stability of amplified genes, and the location of amplified genes. As a result, telomere-type clones, in which the amplified gene was located near the telomeric region, were found to be more stable and productive than other types of clones. Telomere-type clones had over 100 copies of amplified genes in the chromosomal DNA. In contrast, a large number of other types of clones had less than 10 copies of amplified genes. During long-term cultivation in the absence of MTX, in other types of clones, amplified genes rapidly decreased in the chromosomal DNA.

Animals↗

PubMedDate not supplied

Disseminated histoplasmosis in a sea otter (Enhydra lutris).

Disseminated histoplasmosis was diagnosed in a 4.75-year-old, captive female sea otter (Enhydra lutris). At necropsy, the liver was found to be markedly swollen, with many nodules (4-12 mm in diameter). Histologically, macrophages containing numerous intracellular yeast-like organisms were noted in the liver, spleen, lung and kidney. These organisms were labelled immunohistochemically with anti-histoplasma yeast antibody. Ultrastructurally, the yeast-like organisms, 2-4 microm in diameter, were found within membranous structures in the cytoplasm of macrophages. This is the first confirmed report of disseminated histoplasmosis in sea otters.

Animals↗

PubMedDate not supplied

Malignant endocrine carcinoma of the stomach.

Small cell carcinoma and small portions of poorly differentiated adenocarcinoma of the stomach sometimes differentiate into secretory cells. We experienced a pure endocrine carcinoma of the stomach. The patient, who had a giant mass in the greater curvature of the body of the stomach and regional lymph node metastasis, underwent total gastrectomy. Immunohistologic staining of the resected specimens revealed that the tumor was an endocrine carcinoma. The tumor cell shape in the present case was polygonal or oval and the cell size was relatively large. The cell nucleus was coarse chromatic and nucleoli were seen, whereas, the nucleus of small cell carcinoma often shows a fine chromatic pattern and nucleoli are not seen. On the basis of these findings, the present case was considered to be a pure endocrine carcinoma which was not classified as a small cell carcinoma.

Biomarkers, Tumor↗

PubMedDate not supplied

Quantitative in vitro bioassay for recombinant human interleukin-11.

A cell culture-based in vitro bioassay was developed to measure the biological activity of recombinant human interleukin-11 (rhIL-11). The bioassay measures induced proliferation of T10 cells, derived from the T1165 murine plasmacytoma line. A colorimetrically detectable formazan product, obtained by cellular reduction of the tetrazolium compound, 4-[3-(4-iodophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-1,3-benzene disulfonate (WST-1), was used as an endpoint for response of clone T10 to added rhIL-11. Positions of the samples and the standards in 96-well microplates affected the precision of this bioassay, which was improved by using 2 microplates where serially diluted sample and standard lines were interleaved and their positions were alternated. The coefficient of variation for this bioassay was less than 8%. This method is suitable for quality control of rhIL-11 because of its simplicity, reproducibility, and accuracy.

Animals↗

PubMedDate not supplied

Characteristics of gastric mucus in elderly patients with gastric ulcers.

BACKGROUND/AIMS: Studies of the most important defensive factor, gastric mucus, in the treatment of gastric ulcers in elderly patients have been lacking. Therefore we focused on the changes in gastric mucus during the ulcer-healing process in elderly patients. METHODOLOGY: Twenty elderly patients (> or = 65 years old), and 20 younger patients (< 65 years old) with gastric ulcers were administered antisecretory agents for 24 weeks. Biopsies were taken from the antrum and body of the stomach, and the levels of gastric mucosal hexosamine and periodic acid-Schiff-positive gastric mucus were measured. RESULTS: In both groups, the hexosamine levels in the specimens from the body of the stomach declined during the healing process. The decrease was more marked in the elderly, and the recovery of this level was also slower than in the younger group. The periodic acid-Schiff-positive mucosal index was also lower in the elderly. A decrease in body periodic acid-Schiff-positive mucus was seen with treatment in both groups, but recovery was slower in the elderly group. CONCLUSIONS: A decrease in gastric mucus, as a gastric mucosal defensive factor, was seen in gastric ulcers in elderly patients. The potential usefulness of the administration of mucosal protective agents for elderly patients with gastric ulcers was suggested.

Adolescent↗

PubMedDate not supplied

Variables influencing the prognosis in atrioventricular block.

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Adult↗

PubMedDate not supplied

Carbonyl reductase activity for acetohexamide in human erythrocytes.

Acetohexamide is an oral antidiabetic agent and is metabolized by the reductive conversion of the acetoxy group to a secondary alcohol metabolite. In vivo, many drugs are metabolized by reductase enzymes; however, the characteristics of the enzymes that reduce carbonyl compounds need to be clarified. We tested whether reductase activity for acetohexamide can be found in human erythrocytes. Enzyme activity was monitored by formation of hydroxyhexamide using HPLC methods. In human erythrocytes, reductase activity (6.10 +/- 1.20 nmol/min/g hemoglobin) (mean +/- SD) was indeed observed, when 0.5 mM acetohexamide was used as a substrate. KM values and Vmax at the physiologically important pH 7.4 were 0.70 +/- 0.13 mM and 9.19 +/- 0.88 nmol/min/g hemoglobin, respectively. Separation of protein by gel filtration gave one major peak fraction with reductase activity whose molecular weight was estimated to be 31,000. Known substrates of carbonyl reductase such as menadione, daunorubicin, and ethacrynic acid inhibited the acetohexamide reduction. The acetohexamide reductase in erythrocyte showed characteristics of carbonyl reductase. Furthermore, acetohexamide reductase activity in erythrocyte was approximately 30% activity of that of human liver (0.17 +/- 0.05 nmol/min/mg cytosolic protein). The pattern of inhibitors in human liver was essentially the same as that in erythrocytes. It is plausible that the activity in erythrocytes may predict the activity in the liver. It was concluded that carbonyl reductase in human erythrocyte plays an important role in acetohexamide metabolism.

Acetohexamide↗