Regulation of glucose transport by angiotensin II and glucose in cultured vascular smooth muscle cells.

Glucose transport in response to angiotensin II (AII) was assessed in cultured vascular smooth muscle (VSM) cells by measuring the uptake of [3H]-2-deoxyglucose, a radiolabeled non-metabolizable glucose analog. Significant stimulation occurred by 2 hr of exposure with the maximum effect being observed between 6 and 8 hr. All effects were concentration dependent with a threshold response being detected at 0.1 nM. AII-stimulated transport was blocked by saralasin, an AII receptor antagonist, indicating that AII binding to a specific receptor is required for AII to elicit the transport response. AII-stimulated transport was also blocked when cells were incubated with cycloheximide for 6 hr, suggesting that protein synthesis is required for the long-term effects of AII on glucose transport. A specific protein synthesized in response to AII stimulation was the GLUT 1 glucose transporter as assessed by western blot analysis. Inhibition of protein kinase C (PKC) by bisindolylmaleimide and staurosporine did not affect VSM responsiveness to AII, suggesting that AII is capable of stimulating glucose transport through a PKC-independent mechanism; however, VSM responsiveness to AII did appear to be dependent upon the presence of extracellular calcium. The importance of calmodulin in mediating the response of VSM cells to AII was indicated by the inhibition of AII-stimulated glucose transport when VSM cells were incubated in the presence of the calmodulin inhibitors, calmidazolium and W7. Finally, glucose uptake increased with decreasing levels of glucose in the incubation medium. This was accompanied by a corresponding decrease in the relative effectiveness of AII in stimulating glucose uptake.

A recipe-based, diet-planning modelling system.

In a recent article in the British Journal of Nutrition, Sklan & Dariel (1993) presented a method for diet planning employing a mixed-integer programming algorithm for meeting nutritional requirements at minimum costs for institutions or individuals. They recognized that most food items are generally consumed in whole units and as such they are represented as integer variables. However, as in most previous studies, they derived the minimum cost diets by optimizing over purchased food items. The present paper presents a computer-assisted, diet-planning modelling system for individuals by optimizing over recipes instead of food items. This is accomplished by restricting the integer programming solutions to those bundles of food that represent reasonably popular meal recipes. The modelling system is composed of three main components: recipe data entry, database management, and the model. The recipe data entry component creates and stores recipes. It also provides nutritional analysis of the recipes. The database management component creates and maintains several databases necessary to build the modelling data file. The modelling component solves the user-specified model. Currently, the model component can solve for the optimal diet by minimizing cost or minimizing cooking and preparation time. The optimal diet is prepared to satisfy the recommended nutritional guidelines for a predefined group of individuals for 1 week. The system currently has 895 popular recipes found in Hawaii. Diet plans generated using this modelling system with differing objectives are discussed and compared.

Effect of the elastase inhibitor, eglin-c, on antibacterial mechanisms in experimental pneumonia. Description of a system to quantitate phagocytic and bactericidal activity of resident murine alveolar macrophages in vivo.

To evaluate the influence of the elastase inhibitor, eglin-c, on lung host defense, normal CD-1 mice were challenged intratracheally with type 3 Streptococcus pneumoniae suspended in phosphate-buffered saline alone or containing 10 mg/ml eglin-c. After infection with 5 X 10(3) colony-forming units (cfu), animals given eglin-c demonstrated a significant enhancement in their capacity to clear viable pneumococci from the lungs at 24 h after challenge; the augmented pulmonary clearance was associated with an increased influx of granulocytes at 6 and 24 h. After challenge with higher inocula (5 X 10(4) and 5 X 10(5) cfu), animals treated with eglin-c exhibited a significant impairment in pulmonary clearance at 6 h; however, in the presence of larger numbers of granulocytes within the bronchoalveolar spaces, the attenuation in pulmonary clearance resolved between 6 and 24 h. Changes in the kinetics of pulmonary clearance that were similar to those noted after infection with high pneumococcal inocula were also observed after challenge with 1 X 10(6) cfu Staphylococcus aureus. In addition, although it did not influence the in vivo phagocytic capacity of resident alveolar against S. aureus, eglin-c depressed the bactericidal activity of these cells. We conclude that in the mouse, high doses of eglin-c alter pulmonary antimicrobial mechanisms important for preventing and eradicating bacterial infection of the lower respiratory tract.

Cloning of a non-c-myc DNA fragment from the double minutes of a human colon carcinoid cell line.

The cell line COLO 320 DM, derived from an untreated human colon carcinoid tumor, was subcloned to obtain a population (Cl 11) with an average of 37 double minutes (DM) per cell. Fractionation of the chromosomes by differential centrifugation yielded a fraction enriched in DM. DNA isolated from the DM-enriched fraction was inserted into the Pst I site of pBR322. One clone, p446, representative of a number of similar clones, contained a region complementary to genomic unique sequences (region p446U). Southern blot analysis using COLO 320 DNA, and DNA from two other cell lines derived from the same biopsy, COLO 320 HSR and COLO 321 HSR, demonstrated amplification and rearrangement of sequences complementary to p446U when compared with 28 different tumor and normal cell lines, some of which contained DM or homogeneously staining regions (HSR). COLO 320 DM Cl 11 had approximately 110 copies per cell of the p446U sequence, or three copies per DM. COLO 320 HSR, which contained one HSR, had 35 copies per cell, while COLO 321 HSR, which contained two HSR, had 700 copies. In addition, p446U did not hybridize with insert sequences of recombinant plasmid pHM(E + H), which includes the human c-myc coding region, 3 kb of upstream flanking sequences and 0.5 kb of downstream flanking sequences, or with an exon 3 probe, pMYC RI-CLA. Amplification of p446U was also not seen in cell lines containing amplified c-myc or N-myc genes. These results indicate that more than one sequence may be amplified in DM or HSR containing tumor cells, but that they need not be amplified together in other tumors.

Multiple cell lines from patients with malignant melanoma: morphology, karyology, and biochemical analysis.

Of 23 cell lines representing 8 patients with malignant melanoma, one cell line from each patient has been extensively characterized by means of phase morphology, ultrastructural morphology, biochemical markers, steroid receptor protein analysis, and steroid hormone production. Extensive cytogenetic characterization was compiled from seven of the eight cell lines. The melanocytic derivation of the tumor cell lines was supported by the detection of the catecholamines epinephrine, norepinephrine, and dopamine. In addition, well-defined melanosomes were present in six of the seven lines whose ultrastructure were examined morphologically. None of the lines produced alpha-fetoprotein chorionic gonadotropin, or carcinoembryonic antigen in detectable amounts. The individuality of the cell lines was confirmed by phenotypic patterns of isozymes and by karyology.

A human gallbladder adenocarcinoma cell line.

A continuous cell line, COLO 346, was established from a liver metastasis in a patient with adenocarcinoma of the gallbladder. COLO 346 grew as an adherent monolayer of pleomorphic epithelioid cells. COLO 346 cells produced esterone, but no estradiol, progesterone, or cortisol. No adrenocorticotropic hormones, beta-subunit of human chorionic gonadotropin, carcinoembryonic antigen, or alpha-fetoprotein production by the cells was detected. Cell doubling time was 36 h. Seven allelic isozymes were assayed. COLO 346 had a chromosome mode of 74 at 21 months postestablishment with 6 marker chromosomes present in 100% of the cells analyzed. COLO 346 has been in continuous culture for over 2 yr and is available to other investigators for their studies.

Human cell line (COLO 357) of metastatic pancreatic adenocarcinoma.

A continuous human cell line, COLO 357, with exceptional characteristics was derived from a metastasis of a pancreatic adenocarcinoma. COLO 357 grew as an adhering monolayer with a cell doubling time of 21 h and grew with 10% clonal efficiency in soft agar. COLO 357 cells had numerous lamellar inclusions. The cells elaborated the pancreatic enzymes trypsin, elastase and chymotrypsin. COLO 357 also secreted appreciable amounts of carcinoembryonic antigen and human chorionic gonadotropin. COLO 357 had a chromosome mode of 53 with 20 identifiable Giemsa-banded marker chromosomes. Nine nucleolar organizing regions were found by silver-stained metaphase preparations. COLO 357 has been "fingerprinted" for seven allelic isozymes. This cell line has been maintained in active culture for over 2 years, is preserved in a cell bank, and is available to other investigators.

Characterization of cell lines from four undifferentiated human malignancies.

Tumor tissue from 4 patients with undifferentiated malignancy was studied by means of electron microscopy and cell culture. The cell lines were characterized for morphologic and ultramorphologic appearance, chromosome constitution, and cell products. Cell types established in culture were compared to histologic diagnosis of the original tumor. In only 1 case originally diagnosed as malignant melanoma were cell cultures and ultrastructure consistent with that diagnosis. Two cases in which cell cultures and ultramorphologic appearance were consistent with melanoma were originally diagnosed as carcinoma and sarcoma. The tumor of the fourth patient, diagnosed as melanoma, had no features in the cell cultures and electron micrographs consistent with melanoma. The reliability of using the presence or absence of melanosomes alone as an absolute diagnostic criterion is doubtful.

A human liposarcoma cell line producing hyaluronic acid.

A human liposarcoma cell line COLO 222, derived from a primary tumor in a 62-year-old male, elaborates hyaluronic acid. COLO 222 is characterized on the basis of histochemical, ultramorphological, and cytogenetic properties, along with isozyme phenotype and cell products. A chromosome mode of 53 predominates and unique Giemsa-banded marker chromosomes are identified. An autochthonous lymphoid cell line, COLO 143v, was established after the addition of exogenous Epstein-Barr virus. Cytogenetic analysis of Colo 143v is consistent with a normal male karyotype. COLO 143v possesses B-cell characteristics. This autochthonous system had been used for immunological studies and cytotoxicity assays.

Cell lines from human colon carcinoma with unusual cell products, double minutes, and homogeneously staining regions.

Two human colon carcinoma cell lines derived from the same tumor specimen were characterized. The cell lines, COLO 320 and COLO 321, have amine precursor uptake and decarboxylation cell properties, such as ectopic production of norepinephrine, epinephrine, serotonin, adrenocorticotropic hormone, and parathyroid hormone. The cells were morphologically different from most colon cell lines. Double minutes (DM) were initially present in nearly 100% of the metaphases. In a few subcultures of COLO 320, DM have persisted for 1.5 years. However, in COLO 321 and some subcultures of COLO 320, a loss of DM was observed and new marker chromosomes with homogeneously staining regions were observed. These unusual cell lines should be valuable for studies of apudomas of the colon and the cytogenetic phenomena of DM and homogeneously staining regions.

Comparison of four new cell lines from patients with adenocarcinoma of the ovary.

Permanent human tumor cell lines COLO 110, COLO 316, COLO 319, and COLO 330 were established from four patients with serous cystadenocarcinoma of the ovary. COLO 110 was derived from primary tumor tissue; COLO 316, COLO 319, and COLO 330 were derived from cells in malignant effusions. COLO 110 and COLO 316 grew as monolayers of epithelioid cells in culture; COLO 319 and COLO 330 grew as vermiform, floating colonies of epithelioid cells in culture. Epithelial-like morphology was confirmed by transmission electron microscopy. All four cell lines had marker chromosomes and double minute chromosomes. Giemsa banding revealed chromosomes 1, 3, 6, and 7 were involved in markers in all four lines, and chromosomes 2, 4, 5, 9, 11, and 15 were involved in markers in three of the cell lines. Marker chromosomes with possible homogeneous staining regions were observed in COLO 319. Estrone was elaborated by three of the lines, but neither chorionic gonadotropin, carcinoembryonic antigen, nor estrogen or progesterone receptor proteins were detected. Each cell line demonstrated a distinctive isozyme phenotype. These cell lines are maintained in active culture and in a cell bank for distribution to other investigators.

Tumor and lymphoid cell lines from a patient with carcinoma of the colon for a cytotoxicity model.

Three tumor cell lines (COLO 201, COLO 205, and COLO 206) have been established from ascites fluid obtained from a male patient with adenocarcinoma of the colon. In addition to the tumor lines, two lymphoid lines (COLO 197 and COLO 200) have been established from the same patient, with one line from the original biopsy and one from peripheral blood. Characterization of the tumor cell lines revealed four cell types that differ from most colon cell lines reported by others. Chromosome markers were identical in COLO 201 and COLO 205. A long-arm isochromosome 5 observed in COLO 201 and COLO 205 was absent in COLO 206. Statistical analysis of autosomal polysomy revealed that these cell lines were stable and indicated that there may be a cytogenetic basis for the three predominant types of cell morphology. The lymphoid cell line derived from the peripheral blood had a normal male karyotype. The lymphoid cell line derived from a biopsy specimen had a mode of 46 and a deleted chromosome 7 marker. Both lymphoid cell lines had B-cell characteristics. These autochthonous cell lines have been used for immunological studies in cytotoxicity assays and immunoglobulin characterization.

A transitional cell carcinoma cell line.

A transitional cell carcinoma cell line, COLO 232, was derived from a primary urinary bladder tumor in a Caucasian male. In culture, COLO 232 retained distinct uroepithelial phenotypic traits and produced both carcinoembryonic antigen and adrenocorticotropic hormone. COLO 232 had a chromosome mode of 58 and retained the X and Y chromosomes. Ten marker chromosomes were identified. COLO 232 will be of value for biochemical and immunological studies.

Marker properties of tumor and lymphoid cell lines derived from a patient with squamous cell carcinoma.

A squamous cell carcinoma tumor cell line, COLO 227, derived from a metastatic tumor in a Caucasian male, produces both parathyroid hormone and carcinoembryonic antigen. A chromosome mode of 106 predominated and the X and Y chromosomes were retained. Seven marker chromosomes were identified. Cytogenetic analysis revealed an isochromosome 8 marker similar to a HeLa cell line marker and an isochromosome 17 marker described in other cancers. An autochthonous lymphoid cell line, COLO 219, was established and characterized. COLO 219 is a normal lymphoid cell line with B-cell characteristics. This autochthonous system of both cultured tumor cells and cultured lymphocytes is of use in immunological studies.