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Biomedical subjects

D Beck

Publications and source records attributed to D Beck.

At least 37 records · Page 2Linked to original sources

Lysophosphatidylcholine-stimulated protein and glycoprotein production by human gallbladder mucosal cells.

It has been demonstrated in experimental cholecystitis in cats produced by lysophosphatidylcholine that the development of inflammation is associated with the exsorption of a large amount of protein into the gallbladder lumen. It was subsequently demonstrated that in feline experimental cholecystitis the protein produced was albumin and that its production was decreased by vesicular transport inhibitors, suggesting an active secretory process. In the present study, the effect of lysophosphatidylcholine on protein production by fresh, isolated human gallbladder mucosal cells was evaluated. Isolated gallbladder mucosal cells were incubated with [14C]leucine for 24 hr in tissue culture medium. The cells readily incorporated the radioactive label into cellular protein, a process inhibited by cycloheximide. Exposure of the cells to lysophosphatidylcholine for 1 hr in buffer solution resulted in loss of intracellular protein into the buffer solution. Exposure of the cells for 1 hr prior to lysophosphatidylcholine administration to vesicular transport inhibitors, colchicine, and cytochalasin B and to 4 degrees C culture conditions failed to alter the lysophosphatidylcholine-produced passage of the 14C label extracellularly. SDS-PAGE evaluation of the protein produced demonstrated that human gallbladder mucosal cells continuously produced a 66-kDa protein that was not increased by increasing concentration of lysophosphatidylcholine and a 14-kDa protein that increased with increasing concentrations of lysophosphatidylcholine. Employing Western blotting with specific antibodies, the 66-kDa protein was demonstrated to not be albumin but a 66-kDa glycoprotein, and the 14-kDa protein was demonstrated to contain phospholipase A2. Human gallbladder mucosal cells produced a protein and glycoprotein in response to lysophosphatidylcholine by a mechanism not related to vesicular transport.

Carbon Radioisotopes

CD44 expression and modulation on human neuroblastoma tumours and cell lines.

The human CD44 cell surface glycoprotein has been involved in a variety of functions including lymphocyte homing, extracellular cell matrix attachment and tumour metastasis. A large family of variants or isoforms, generated by alternative splicing of a single gene, has been reported to be involved in the malignant process, by conferring metastatic potential to non-metastatic cells. Neuroblastoma is a tumour characterised by an aggressive and metastatic behaviour in advanced stages, with amplification of the MYCN protooncogene. In this report, we show that the CD44 standard molecule was highly expressed in the majority of tumours of stages 1-3, in all stage 4s and ganglioneuromas, but only in a subset of stage 4 tumours. A lack of CD44 expression was observed in all MYCN amplified stage 4 tumours, thus demonstrating a highly significant inverse relationship between MYCN amplification and CD44 expression in neuroblastoma. In addition, the expression of 4 different CD44 isoforms was measured on all specimens and was always found to be negative. Using neuroblastoma cell lines and MYCN expressing transfectants, we show that CD44 expression by neuroblastoma cell lines is not directly related to MYCN amplification, but is associated to the stage of differentiation or lineage, and to the tumorigenic properties of the cells. In addition, CD44 expression can be upmodulated parallel to differentiation or maturation as induced by retinoic acid, bromodeoxyuridine or phorbol ester. In contrast, cytokines such as IFN gamma, TNF alpha, or growth factors such as bFGF, SCF and TGF beta were ineffective in modulating CD44 expression.

Biomarkers, Tumor

Effects of stem cell factor and other bone marrow-derived growth factors on the expression of adhesion molecules and proliferation of human neuroblastoma cells.

Metastasis in children with neuroblastoma (NB) is a poor prognostic factor despite intensive therapy. In the near future, stem cell factor (SCF) is likely to be used clinically to accelerate bone marrow (BM) recovery after high-dose chemotherapy in patients with advanced NB. The high frequency of BM metastases in NB could be secondary to BM-derived human growth factors (HGF) modulating the adhesion, secondary growth (or both) of circulating metastatic NB cells. To test this hypothesis, we studied the in vitro effects on NB cell lines grown in chemically defined medium of SCF, interleukin (IL)-1 beta, IL-3, IL-6, basic fibroblast growth factor (bFGF), transforming growth factor-beta (TGF-beta) used alone or in combination. The antigenic expression of NB-associated cell adhesion molecules (CAM) HLA class 1, intercellular CAM-1, neural-CAM and CD44 were assayed by monoclonal antibodies and flow cytometry, and DNA synthesis by 3H-thymidine uptake. The expression of CAM was not modulated by SCF or other HGFs. An increase in thymidine uptake was induced by bFGF alone in IMR-32 cells, while SCF and other HGFs had no notable effect. Our results indicate that SCF and other BM-derived HGFs are unlikely to have a generalised effect on the expression of adhesion molecules by NB cells or proliferation. The clinical administration of recombinant human SCF to children with NB should be safe.

Cell Adhesion Molecules

Comparison of the flow rates of central venous catheters designed for rapid transfusion in infants and small children.

Large-bore Hickman catheters are useful in infants and small children for the rapid transfusion of blood or fluids into the central circulation. Recently high-flow plastic sheaths have been developed for the same purpose. We compared the flow rates of normal saline, 5% albumin and packed red blood cells through two sizes of Hickman catheters that have been recommended for major surgery in infants to five sizes of Arrow plastic sheaths of comparable external diameters, and to 14 and 16 gauge Jelco catheters. The flow rates of all three solutions through the plastic sheaths and the 14 gauge Jelco catheters were superior to both sizes of Hickman catheters. Shortening the Hickman catheters improved their flow. High-flow plastic sheaths can provide a useful alternative to Hickman catheters in patients where permanent, large-bore central venous catheters are not required. Hickman catheters should be shortened as much as safely possible if massive haemorrhage is anticipated.

Catheterization, Central Venous

Altered growth and phenotype in clonal mycN transfectants of the SK-N-SH neuroblastoma cell line.

We have attempted to distinguish in human neuroblastoma between the effects of mycN on differentiation and its potential to promote malignant progression. Others have observed out-growth of autocrine cells with evidence of an advanced malignant phenotype in a mycN-transfected clonal cell line derived from the single-copy mycN neuroblastoma, SK-N-SH. We have now transfected the parental cell line with the same mycN expression vector and selected 5 clones characterized by unique and stable chromosomal integration sites and variable exogenous copy numbers. mycN gene expression was variable in the different clones and correlated roughly with the copy number of transfected mycN genes. Clones with minimal levels of mycN gene expression had a neuroblastic phenotype and low numbers of surface HLA class-I molecules. Clones with high levels of mycN expression had a Schwann/glial-like phenotype with higher surface HLA class-I display without imbalance of expression of specific loci and accelerated growth. Two such clones were capable of anchorage-independent growth in the absence of serum, and acquired tumorigenic properties. Our results show that exogenous mycN expression can be associated with a differentiation of neuroblastoma cells along the Schwann/glial pathway and can induce accelerated and autonomous growth.

Animals

CD44H expression by human neuroblastoma cells: relation to MYCN amplification and lineage differentiation.

The human CD44 cell surface glycoprotein has been involved in a variety of functions including lymphocyte homing, extracellular cell matrix attachment, and tumor metastasis. Due to the alternative splicing of the single gene, a large family of different variants or isoforms is generated. Several reports have indicated an up-regulation of CD44 variant (v) isoforms in malignant process, conferring metastatic potential to non-metastatic cells. Neuroblastoma is a tumor characterized by an aggressive and metastatic behavior in advanced stages with amplification of the MYCN protooncogene. In this report we show that the CD44 standard molecule is highly expressed in 100% of stage I-III, IVs neuroblastomas and ganglioneuromas but only in a subset of stage IV tumors. In contrast, no expression of CD44 was detected on MYCN amplified stage IV tumors, thus demonstrating a highly significant negative relationship between MYCN amplification and CD44 expression in neuroblastoma. The expression of CD44 on neuroblastoma cultured cell lines was not shown to be related to MYCN amplification but rather linked to the S-type, schwann/glial differentiation lineage. Immunochemical analysis of tumor samples with anti-CD44v3 and -v6 antibodies and Northern blot analysis of mRNA from cell lines with probes spanning exons 4-10 did not reveal any expression of splice variants on neuroblastomas of all stages and cell lines, thus ruling out a major role of these isoforms in neuroblastoma progression and metastasis.

Blotting, Northern

Treatment of relapsing acute lymphoblastic leukemia in childhood. III. Experiences with 54 first bone marrow, nine isolated testicular, and eight isolated central nervous system relapses observed 1985-1989.

Of 54 children with acute lymphoblastic leukemia (ALL) and first hematological recurrence observed between 1985 and 1989, 31 relapsed while still on treatment and 23 after cessation of therapy. Of the former, only one survived. Of the latter, 11 children survived after a minimum follow-up of 25 months. During the same period, a first isolated testicular relapse was observed in nine boys, of whom six survived, and an isolated CNS relapse in eight patients, of whom three survived. As a rule, survivors of a bone marrow or testicular relapse were doing well while those surviving a CNS relapse had considerable neuropsychological sequelae. These results, compared with those of two preceding studies, suggest that with intensification of front-line treatments, it becomes more difficult to rescue children who relapse, particularly those with a bone marrow relapse while on therapy.

Adolescent

Studies on the etiology of acute acalculous cholecystitis: the effect of lipopolysaccharide on human gallbladder mucosal cells.

Previous studies in animals have shown that lipopolysaccharide produces experimental cholecystitis possibly through a platelet-activating factor-prostanoid mediated process. In this study it was intended to evaluate the effect of LPS on primary cultures of human gallbladder mucosal cells. Gallbladder mucosal cells were isolated from gallbladders removed during routine cholecystectomies or other operations. The cells were cultured for 24 h before treatment. Unstimulated cells produced low levels of prostanoids and significant basal levels of PAF. LPS produced stimulation of eicosanoid and PAF secretion. The increased prostanoid formation was not enhanced when LPS and PAF were administered together. Prostanoid synthesis was inhibited by the administration of a cyclooxygenase inhibitor while administration of a PAF receptor antagonist significantly increased prostanoid formation, suggesting that increased PAF levels function as a negative control mechanism to decrease prostanoid synthesis. The results suggest that endotoxemia may produce a cascade of inflammatory processes in human gallbladder mucosal cells resulting in the development of acute acalculous cholecystitis.

Acute Disease

[GRIN-endoscopy for laser treatment in ophthalmology].

Endoscopes for use in ophthalmology can be manufactured with diameters smaller than 0.89 mm. Because gradient-index (GRIN) lenses yield a higher resolution than ordered fiber bundles having the same diameter, GRIN lenses with a diameter of either 0.35 mm or 0.50 mm were utilized to manufacture thin high resolution endoscopes. Whatever the working distance, the resolution obtained with a GRIN lens having a diameter of 0.50 mm was higher than that obtained with a GRIN lens of 0.35 mm. Thanks to the miniaturization, an additional working channel could also be integrated within the endoscope. This allowed new combinations of various surgical instruments. This channel can be used as a guide for an optical fiber for laser therapy or as an irrigation/aspiration channel. The use of such an instrument where the observation path is parallel to the surgical instrument was compared to the application of the endoscope as a separate monitoring system. The advantages and inconveniences of both systems are discussed. Preference is given to a system where the diameter of the observation channel is maximum and the optical fiber for laser delivery is integrated but adjustable along its length within the endoscope.

Endoscopes

Genotypic and phenotypic diversity of Rhizobium isolated from chickpea (Cicer arietinum L.).

The diversity of 16 strains of chickpea-infective rhizobia from various geographical origins was analysed using genotypic and phenotypic approaches. Multilocus enzyme electrophoresis was performed, and restriction fragment length polymorphisms of the amplified 16S + IGS (intergenic spacer) rRNA gene, assimilation of 147 carbon sources, antibiotic resistance, and tolerance to NaCl and extreme pH values and temperatures were tested. These approaches had different discriminating powers. Esterase polymorphisms gave a unique pattern for each strain, allowing this method to be used for strain fingerprinting. Genetic distances between strains were estimated. The three approaches used in this study yielded consistent results. They evidenced high heterogeneity among the strains, and made it possible to classify the strains into two clusters. Isozyme patterns for superoxide dismutase were particularly interesting, since they delineated the same two groups. The phenotypic tests clearly confirmed the existence of two genetic groups on the basis of 11 phenotypic characters. Owing to the large phylogenetic distance between the two groups of strains, the taxonomic status of chickpea-infective strains is discussed.

Bacterial Proteins

Comparative metabolism of SC-42867 and SC-51089, two PGE2 antagonists, in rat and human hepatocyte cultures.

1. The metabolism of SC-42867 and SC-51089, two PGE2 antagonists, was studied in cultured rat and human hepatocytes. Both compounds possess an 8-chlorodibenzoxazepine moiety, but differ from each other by the nature of the side chain connected to the nitrogen atom. SC-42867 and SC-51089 and their in vitro metabolites were separated by reversed-phase hplc. The major metabolites of both compounds were identified by mass spectrometry (ms) analysis. 2. SC-42867 was metabolized on the tricyclic moiety only. Oxidative N-dealkylation with opening of the oxazepine ring was the major metabolic pathway obtained in rat hepatocytes. The metabolic profile obtained in cultured human hepatocytes was comparable with that of cultured rat hepatocytes. However, the compound was metabolized to a much lower extent by the human cells. 3. SC-51089 was extensively metabolized by both cultured rat and human hepatocytes. Human cells metabolized this compound quite differently than cultured rat hepatocytes. Aromatic hydroxylation with consequent glucuronidation and sulphation were the main metabolic pathways observed in cultured human hepatocytes. Oxidative N-dealkylation with opening of the oxazepine ring and consequent glucuronidation was the major metabolic pathway observed in rat hepatocytes. Further metabolism occurred, in contrast with the human hepatocytes, mainly on the side chain. 4. The present in vitro results are compared with data of previous in vivo studies performed in rat.

Adult

Transvaginal sonography, color flow imaging, computed tomographic scanning, and CA 125 as a routine follow-up examination in women with pelvic tumor: detection of recurrent disease.

We were trying to determine whether transvaginal sonography combined with color flow imaging should be used routinely for detecting recurrent pelvic tumors in patients treated for pelvic cancer. Eighteen patients who had completed their treatment for pelvic tumor were examined every 6 months by gynecologic examination, a CT scan, serum CA 125 levels, and transvaginal sonography with color flow imaging. Laparotomy or fine-needle biopsy was performed when a recurrent pelvic mass was observed or suspected on CT scans or transvaginal sonography. In nine of 18 patients a recurrent pelvic tumor was observed or suspected on CT scans or on transvaginal sonography. A recurrent pelvic tumor was diagnosed on laparotomy in five patients. In four patients, only a fine-needle biopsy was performed and only two of them had positive results. The highest sensitivity and specificity for detecting recurrent pelvic tumors were achieved by transvaginal sonography (100 and 91%, respectively) and by color flow imaging (86 and 100%, respectively). Intratumoral blood vessels were detected in all cases with recurrent pelvic tumors, and in only one of these cases the PI was high. We recommend a routine follow-up with transvaginal sonography in women completing their treatment for pelvic tumors. When a recurrence of tumor is only suspected on transvaginal sonography, color flow imaging may be used as an additional diagnostic tool.

Aged

[Late sequelae in a group of children and adolescents treated for cancer between 1981 and 1986 in Switzerland].

An ever-growing number of children with malignant diseases can today be cured and will have prolonged survival. Their quality of life, which can be altered by a substantial number of late effects of the disease or its treatment, is a prominent field of long-term assessment. In a cross-sectional study based on clinical and paraclinical parameters and covering 59 children and young adults treated between 1981 and 1986 for a childhood malignant disease, we investigated late effects in the long-term survivors. Late effects were found in 36 (61%) of these patients; their spectrum is wide, but most often affected are the CNS and the musculoskeletal system. Two thirds of the patients have mild or moderate and one third severe problems. The frequency of the late effects and the type of problem vary greatly according to the initial diagnosis; our results show a significant difference of severity according to the number of affected systems (p < 0.05). The results of our study match those of other similar reports and confirm the high frequency of late effects in survivors of pediatric cancer. A prospective study, assessing a large population of patients, is needed to determine the underlying causes and to develop primary and secondary prevention strategies.

Adolescent

Prognostic factors and long-term survival in endometrial adenocarcinoma with cervical involvement.

The clinical, surgical, and histopathologic data from 202 patients with endometrial adenocarcinoma with cervical involvement are presented. One hundred fifty-one (75%) had histopathologically confirmed cervical involvement at the time of their definitive surgery, while in 51 (25%) no cervical involvement was conclusively identified. The 5-year actuarial survival for patients with true surgical stage II endometrial carcinoma (N = 24) was 76%. Extrauterine disease was documented in 32% (27/84) of patients in which the primary treatment modality was surgical. The 5-year actuarial survival was 65% for all patients with clinical surgical stage II disease. There appeared to be a survival advantage for patients treated by radical surgery as compared with more conventional treatments, especially in patients with numerous high-risk factors. The subgroup of patients (N = 53) having tumor grossly involving the cervix had a 5-year survival of 48%. In this subgroup of patients, radical hysterectomy offered improved 5-year survival over more traditional forms of treatment, particularly compared with simple hysterectomy or combined treatment with radiation and surgery. Multivariate analysis positively correlated myometrial invasion, grade, uterine serosal involvement, lower uterine segment involvement, adnexal metastasis, pelvic metastasis, aortic node metastasis, and peritoneal cytology, with disease-free survival. Clinical and surgical findings correlated poorly; therefore, primary surgical evaluation is recommended when possible.

Adenocarcinoma

Culturing of primary hepatocytes as entrapped aggregates in a packed bed bioreactor: a potential bioartificial liver.

Conventional culture systems for hepatocytes generally involve cells cultured as flat, monolayer cells, with limited cell-cell contact, in a static pool of medium, unlike the liver in vivo where the parenchymal cells are cuboidal, with extensive cell-cell contact, and are continuously perfused with blood. We report here a novel bioreactor system for the culturing of primary hepatocytes with cuboidal cell shape, extensive cell-cell contact, and perfusing medium. The hepatocytes were inoculated into the bioreactor and allowed to recirculate at a rate optimal for them to collide and form aggregates. These newly-formed aggregates were subsequently entrapped in a packed bed of glass beads. The bioreactor was perfused with oxygenated nutrient medium, with controlled oxygen tension, pH, and medium perfusion rate. The hepatocytes were viable for up to the longest time point studied of 15 days in culture based on urea synthesis, albumin synthesis and cell morphology. Light microscopy studies of hepatocytes cultured for 15 days in the bioreactor showed interconnecting three-dimensional structures resembling the hepatic cell plate in the liver organ. Electron microscopy studies on the same cells revealed ultrastructure similar to the hepatocytes in vivo, including the presence of plentiful mitochondria, rough and smooth endoplasmic reticulum, glycogen granules, peroxisomes, and desmosomes. We believe that our hepatocyte bioreactor is a major improvement over conventional culture systems, with important industrial applications including toxicology, drug metabolism, and protein/peptide synthesis. The hepatocyte bioreactor concept may also be used as the basis for the development of a bioartificial liver to provide extracorporeal hepatic support to patients with hepatic failure.

Albumins