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J Kieler

Publications and source records attributed to J Kieler.

At least 19 recordsLinked to original sources
PubMed1995

Cell cycle perturbation by sodium butyrate in tumorigenic and non-tumorigenic human urothelial cell lines assessed by flow cytometric bromodeoxyuridine/DNA analysis.

The effect of sodium butyrate on cell proliferation was studied in eight human urothelial cell lines differing in transformation grade (TGr): Hu 1752 (mortal, TGr I); HCV29 (immortal and tumorigenic, TGr II); HCV29T, T24, T24A, T24B, Hu 961A and Hu 1703He (tumorigenic, TGr III). In all cell lines, except Hu 1752, addition of 4 mM sodium butyrate at 18 h after replating resulted in a significantly decreased population of adherent cells after a further 24-48 h. This might partially be explained by detachment of cells, probably mainly S phase cells, from the substrate in the lines HCV29, HCV 29T, Hu 961A and Hu 1703He. Flow cytometric DNA analysis of the adherent cell population showed that all TGr II and III urothelial cell lines were DNA aneuploid, and that butyrate perturbed the cell cycle distribution in these cell lines, mainly by a decrease of the S phase fraction. Flow cytometric bromodeoxyuridine (BrdUrd)/DNA analysis of continuously BrdUrd labelled cultures, using a 'washless' BrdUrd/DNA staining technique, showed that butyrate inhibited the G0/1-S phase transition, indicated by a delayed depletion of BrdUrd negative G0/1 cells in the cell lines HCV29, HCV29T, T24B, Hu 961A and Hu 1703He. BrdUrd/DNA analysis further showed that butyrate inhibited the G2M-G0/1 phase transition, indicated by a pronounced accumulation of BrdUrd positive G2M cells in the cell lines HCV 29T, T24B, Hu 961A and Hu 1703He. Microscopy of HCV29T and Hu 961A cells indicated that this block did not occur in mitosis. The parental cell line T24 and the cell line T24A did not show an accumulation of BrdUrd negative G0/1 cells or BrdUrd positive G2M cells like that occurring in the derived cell line T24B.

Amnion↗
PubMed1993

Two phorbol ester receptor affinities in partially transformed human urothelial cells and decrease of receptor binding in desensitized cells.

The presence of specific binding sites for phorbol esters was studied in a transformed but non-tumorigenic human urothelial cell line HCV-29 by assay of specific binding of 3H-phorbol-12,13-dibutyrate (3H-PDBu) to intact living cells. 3H-PDBu bound specifically to HCV-29 cells in a saturable and competitive manner. Scatchard plot analysis of specific binding yielded a curved plot consistent with two binding sites with Kd of 11 nM and 102 nM, respectively. At saturation the corresponding PDBu binding capacities (Bmax) were 8.8 pmol/10(6) cells (5.2 x 10(6) molecules bound per cell) and 2.8 pmol/10(6) cells (1.7 x 10(6) molecules bound per cell). 3H-PDBu binding was displaced by biologically active phorbol ester tumor promoters such as 12-O-tetradecanoylphorbol-13-acetate (TPA) and mezerein, but not by tumor promoters such as L-tryptophan, anthranilic acid and sodium saccharin. In cells desensitized by pretreatment with 1 microgram/ml (2 microM) TPA or PDBu for 24 h the level of binding was reduced to 28% of the level in non-exposed cells. The ability of desensitized cells to bind 3H-PDBu was gradually restored within 5-6 days. At the same time the cells became sensitive to the morphological alteration induced by PDBu. This suggests that desensitization of HCV-29 cells is due to a decreased receptor-ligand binding capacity probably associated with down regulation of the phorbol ester receptors.

Caenorhabditis elegans Proteins↗
PubMed1993

Identity of tumorigenic human urothelial cell lines and 'spontaneously' transformed sublines.

Restriction fragment length polymorphism (RFLP) analysis, comparative marker chromosome analysis, and polymorphic enzyme analysis was carried out on a total of eight human urothelial cell lines and sublines selected according to our knowledge of their HLA-A,B phenotype. RFLP analysis and cytogenetic analysis showed that the cell lines Hu1703He, Hu1922, and T24 are genuine cell lines of different origin. The identity of Hu1703He could not be confirmed by its isozyme phenotype which was identical to the T24 phenotype. RFLP analysis and isozyme analysis revealed that three cell lines, Hu456, Hu549, and Hu961a, and two transformed sublines, HCV-29Tmv and Hu609Tmv, are sublines of T24. A common origin of Hu456, Hu549, Hu961a, HCV-29Tmv, and Hu609Tmv was confirmed by marker chromosome analysis. However, the T24 origin of these cytogenetically related cell lines was not supported by chromosome analysis of T24. RFLP analysis and HLA phenotyping of two tumorigenic and invasive sublines isolated from a culture of non-tumorigenic Hu609 cells showed that non-tumorigenic Hu609 cells can transform 'spontaneously' in vitro into tumorigenic Hu609T cells. The results emphasise the need for careful monitoring and screening of cell lines for their identity using more than one identification parameter.

Cell Line↗
PubMed1990

Recombinant human interferon gamma exerts an anti-proliferative effect and modulates the expression of human leukocyte antigens A,B,C and DR in human urothelial cell lines.

In this study we have treated three malignant (TGrIII) and two pre-malignant (TGrII) urothelial cell lines with recombinant human interferon gamma (rHu-INF gamma). The malignant cells (HCV29-T112C1, Hu1703He and T24) were inhibited in growth by more than 50% after treatment with 100-1000 units of rHu-INF gamma/ml for 4 days as compared to untreated controls. The growth of the pre-malignant cell lines (HCV29 and Hu609) was not influenced to the same extent in the presence of rHu-INF gamma in the culture medium. Treatment with rHu-INF gamma increased the expression of monomorphic human leukocyte antigens (HLA) A,B,C as well as beta 2-microglobulin in all the cell lines tested, as demonstrated using a quantitative immunofluorescence assay. The tumourigenic cell lines increased their expression of HLA in a dose-dependent way, whereas treatment of the non-tumourigenic cells with higher concentrations of rHu-INF gamma than 10 units/ml, did not increase the HLA-A,B,C expression further. None of the cell lines expressed HLA-DR unless treated with rHu-INF gamma. No correlation between tumourigenicity and the dose of rHu-INF gamma required for "de novo" induction of HLA-DR could be demonstrated. After removal of rHu-INF gamma from the medium, the expression of HLA-DR gradually decreased in less than 14 days, indicating that the expression of HLA-DR is not constitutive but dependent upon the presence of rHu-INF gamma. We conclude that human urothelial cells grown in vitro are sensitive to the anti-proliferative and major-histocompatibility-complex-modulating effects of rHu-INF gamma, and that malignant urothelial cells are more sensitive than pre-malignant cells. Finally, our data indicate a possible role for rHu-INF gamma in the management of human bladder cancer.

Animals↗
PubMed1990

Reduced HLA-A,B,C expression in tumourigenic v-raf transfected human urothelial cells.

Tumourigenic (TGrIII) human urothelial cells grown in vitro have previously been demonstrated to have a markedly decreased expression of beta 2-microglobulin and HLA-A,B,C antigens as compared to non-tumourigenic (TGrII) human urothelial cell lines. Furthermore, during 'spontaneous' in vitro transformation of a non-tumourigenic (TGrII) human urothelial cell line Hu609 into a tumourigenic (TGrIII) subline Hu609T/LLH, changes in morphology and tumourigenicity have been demonstrated to be accompanied by a decreased HLA-A,B,C expression. After malignant transformation of the non-tumourigenic (TGrII) human urothelial cell line HCV29 by DNA transfection with the v-raf oncogene, four sublines could be isolated. In this study we have investigated these sublines for their expression of membrane bound HLA-A,B,C antigens and provide further evidence that an inverse relationship exists between tumourigenicity and monomorphic HLA-A,B,C expression. Treatment of the cells with recombinant human interferon alpha for 3 days increased the expression of HLA-A,B,C antigens by 50-150% indicating that at least some of the reduced HLA-A,B,C expression could be due to decreased synthesis of HLA-A,B,C antigens. All the transfected cell lines overexpress v-raf and c-myc.

Cell Line↗
PubMed1990

Alpha-2-macroglobulin is not synthesized by human urothelial cell lines in vitro.

We have analyzed 14 human urothelial cell lines in two different transformation grades for the production of a tumor-associated-alpha-2-macroglobulin. It has been shown in radioimmunoprecipitation and immunoblotting tests that human urothelial cells in vitro do not synthesize the wide-spectrum proteinase inhibitor alpha-2-macroglobulin. Four of the tested cell lines were additionally tested for the expression of specific mRNA which resulted in negative findings. The significance of the absence of alpha-2-macroglobulin in the human urothelial cells is discussed.

Animals↗
PubMed1989

Spreading of cells on various substrates evaluated by Fourier analysis of shape.

In order to evaluate in mathematical terms the morphological changes occurring in the course of cell spreading, Fourier analysis of shape was applied. Human urothelial Hu 961 b cells plated on type IV collagen, fibronectin, laminin, glass and bovine serum albumin (BSA) were studied. Fourier parameters describing cell shape as well as surface areas covered by the cells on the substrate were subjected to statistical analysis. Using analysis of variance and discriminant analysis it was found that parameters describing cell shape (both gross shape of cells and their fine scale contour foldings) possessed a higher power of discrimination between the cells spread on various substrates than the differences in cell surface areas. In the course of observation (75 and 150 min) the highest number of attached cells and highest degree of spreading were found when cells were plated on type IV collagen. Moderate alterations in cell shape and moderate increase of surface area were seen in the group of cells seeded on fibronectin, whereas the cells plated on laminin, glass and BSA revealed a moderate increase of surface area, but no changes in their shape were observed. The differences in attachment of cells and in the degree of their spreading might be due to the variation in expression of plasma membrane receptors for various substrates. The Fourier analysis of cell shape coupled with measurement of surface area is a good tool for quantitative evaluation of cell spreading and can be used for discrimination between cells spread on different substrates.

Cell Adhesion↗
PubMed1988

Correlation between classification of human urothelial cell lines and HLA-A,B,C expression.

Quantitative changes in major histocompatibility class I antigen expression in tumour cells are believed to affect the host immune response against the tumour. In tumourigenic (TGrIII) human urothelial cell lines the apparent loss of polymorphic HLA-A,B epitopes has previously been demonstrated. In the present study, 3 non-tumourigenic (TGrII) and 6 tumourigenic (TGrIII) human urothelial cell lines have been investigated for their quantitative expression of monomorphic HLA-A,B,C and B2-microglobulin. Evidence is provided that an inverse correlation exists between tumourigenicity and HLA-A,B,C and B2-microglobulin expression. Furthermore, treatment of the cells with neuraminidase partly restored the expression of monomorphic HLA-A,B,C suggesting that at least some of the observed quantitative differences could be due to masking of the membrane bound HLA antigens by sialic acid-containing glycoconjugates.

Antigens, Neoplasm↗
PubMed1988

Defect in lectin-induced interleukin 2 production by peripheral blood lymphocytes of patients with invasive urinary bladder carcinoma.

The production of interleukin 2 (IL-2) by phytohaemagglutinin (PHA)-stimulated peripheral blood mononuclear cells (PBMC) from 21 patients with transitional cell carcinoma of the urinary bladder (BTCC) and 16 control blood donors was measured with a solid phase enzyme immunoassay based on the dual antibody immunometric sandwich principle. PBMC from patients with invasive BTCC (grade III-IV) showed a defect in the production of IL-2. The concentration of IL-2 in the supernatants of PBMC cultures from these patients was substantially lower (0.4 +/- 0.1 U/ml) than that observed in the supernatants of PBMC cultures from patients with non-invasive BTCC, grade II (1.5 +/- 0.7 U/ml), and from tumour-free controls (1.4 +/- 0.8 U/ml). These results suggest an immune dysfunction based on quantitatively impaired IL-2 production in patients with invasive BTCC and indicate that exogenous IL-2 could be used as an immunological response modifier for the treatment of these patients.

Aged↗
PubMed1988

Local administration of cells containing an inserted IL-2 gene and producing IL-2 inhibits growth of human tumours in nu/nu mice.

We have prepared a retroviral expression construct, pPS-IL-2, in which human IL-2 cDNA has been inserted into the polylinker region, and have used the retroviral vector to introduce the functional IL-2 gene into a fibroblast cell line, RAT-1. Peritumoral administration of IL-2-producing RAT-1 cells into congenitally athymic (nu/nu) mice carrying subcutaneous transplants of human carcinoma cells inhibited the growth of the human tumour xenografts.

Animals↗
PubMed1987

Recombinant interleukin-2 inhibits growth of human tumor xenografts in congenitally athymic mice.

Administration of human recombinant interleukin-2 (RIL-2) into congenitally athymic (nu/nu) mice carrying subcutaneous transplants of HeLa, HU 609T and T24B human carcinoma cells partially inhibited growth of the human tumor xenografts. In vitro activation of nu/nu spleen cells with human RIL-2 resulted in generation of killer cells showing in the 51Cr cytotoxicity assay similar levels of cytolysis as RIL-2-activated spleen cells from heterozygous (nu/+) mice. The RIL-2-activated (LAK) cells were cytotoxic for a variety of mouse and human tumors, reaching the peak of their cytotoxic activity after 3 days of cultivation in the RIL-2-containing medium. The cytotoxic activity of activated nu/nu spleen cells was significantly reduced by treatment with antibody against glycolipid asialo GM1, the differentiation antigen of natural killer (NK) cells. This finding suggests that in addition to the conventional, asialo GM1- LAK cells, asialo GM1+ activated NK cells participated in the cytotoxicity displayed by the IL-2-activated nu/nu killer spleen cells.

Animals↗
PubMed1987

Changes in HLA A, B, C expression during "spontaneous" transformation of human urothelial cells in vivo.

The immortalized but non-tumourigenic and non-invasive human urothelial cell line, Hu 609, known to express the appropriate HLA A,B antigens (A2,-; B5,-) has previously been demonstrated to undergo "spontaneous" in vitro transformation into an invasive and tumourigenic subline, Hu 609T/MV. This subline does not express the polymorphic HLA epitopes. In the present investigation we have followed two additional "spontaneous" transformations of the Hu 609 cell line into malignant sublines. Evidence is presented that the development of morphological changes and tumourigenicity were accompanied by a gradual loss of the expression of polymorphic HLA A,B epitopes and a reduction in the expression of monomorphic HLA A,B,C antigens. Antigens could be detected again after neuraminidase treatment. We conclude that the urothelial Hu 609 cell line after "spontaneous" transformation still possesses the HLA A,B epitopes. The observed quantitative differences in HLA expression between Hu 609 and its malignant sublines may be due to masking of the HLA antigens by sialic acid containing tumour-associated highly branched glycoproteins.

Antigen-Antibody Reactions↗
PubMed1987

Localization of a receptor binding site on the IL-2 molecule.

The present study was performed to localize in the IL-2 molecule the active site responsible for interaction with the IL-2 receptor. To predict the receptor binding site on the IL-2 molecule, a computer programme based on the hypothesis that the active site will contain parts of the protein molecule having a high tendency to form a bend was utilized. The tendency to form a bend was evaluated by assessing the probability of beta-turn occurrence; the highest probability was found in the tetrapeptide Asn-Pro-Lys-Leu, occupying the positions 33-36 of the IL-2 molecule. Accordingly, the hexadecapeptide H-Cys-Nle-Gly-Ile-Asn-Asn-Tyr-Lys-Asn-Pro-Lys-Leu-Thr-Arg-Met-Leu-NH2 that spans over the predicted tetrapeptide Asn-Pro-Lys-Leu and comprises the region 27-40 from the IL-2 amino acid sequence was synthesized. This synthetic (I-16) peptide was found to selectively inhibit the IL-2-dependent uptake of 3H-TDR by CTLL cells, apparently by competing with IL-2 for the IL-2 receptor. The synthetic I-16 hexadecapeptide was conjugated to carrier (BSA) protein and used for immunization of rabbits. Resulting I-16 antibodies were capable of binding specifically to the I-16 hexadecapeptide in indirect ELISA test; they reacted substantially with IL-2-producing but not with IL-2-non-producing Jurkat cells in indirect cell membrane immunofluorescence, and inhibited activation of killer spleen cells with human recombinant IL-2 as detected by 51Cr microcytotoxicity assay. Taken together, these results suggest that at least one of the receptor contact sites of the IL-2 is localized within the N-terminal part of the molecule in the region defined by amino acids 27-40 and coded for by the exon 1 and 2.

Antibodies↗
PubMed1987

Unscheduled DNA synthesis related to transformation grade of human urothelial cells.

Unscheduled DNA synthesis (UDS) was studied by quantitative autoradiography in human urothelial cells of three transformation grades (TGr I-III). Cells incubated in arginine-free medium supplemented with hydroxyurea showed dose-dependent UDS after administration of agents injurious to DNA, while the scheduled synthesis of DNA was nearly totally suppressed. UDS was demonstrated after treatment with the ultimate carcinogen N-methyl-N-nitroso-N'-nitroguanidine (CAS: 70-25-7) or with the procarcinogen 4-nitroquinoline-1-oxide (4-NQO; CAS: 56-57-5). In cultures treated with benzo[a]pyrene (CAS: 50-32-8), which requires a different activation system than that for 4-NQO, UDS was less pronounced. In 9 cell lines the average rates of UDS were inversely related to TGr. Two cell lines showed a different pattern.

4-Nitroquinoline-1-oxide↗
PubMed1986

Altered glycosylation of membrane glycoproteins in human uroepithelial cell lines.

Total cellular glycopeptides of 7 human uroepithelial cell lines that differ in the grade of transformation (TGr) were analysed by gel filtration and affinity chromatography on immobilized lectins. The 4 cell lines that are tumorigenic in nude mice and invasive in vitro (TGr III) possess more highly branched, tri- and tetraantennary N-acetyllactosaminic glycans, with less biantennary glycans than the 2 non-tumorigenic, noninvasive (TGr II) cell lines examined. The only exception to this general pattern is the third cell line, which is classified as TGr II. The cellular glycopeptide distribution pattern in this cell line is similar to that of the TGr III cells. The possible relationship between altered glycosylation of membrane glycoproteins and the expression of a malignant phenotype is discussed.

Cell Line↗
PubMed1986

Numerical evaluation of changes in the cytoplasmic microtubule complex of C3H mouse cells by optical diffractometry and of changes in cell shape by Fourier analysis.

MO mouse cells in culture on glass were treated with taxol, or nocodazole, or incubated at 4 degrees C to alter their cytoplasmic microtubule complex (CMTC). From each treated group and from an untreated group, 30 cells stained with an antiserum against tubulin, were photographed under the photomicroscope, and negatives were analysed by optical diffractometry. Differences between groups of cells were tested by variance analysis. Phase-contrast micrographs of the same cells were used for Fourier analysis of cell shape. Both types of analyses provided numerical objective data about changes in the CMTC and in cell shape that were typical for the kind of treatment. We conclude that optical diffractometry of immunostained cells and Fourier analysis of cell shape are complementary to photomicroscopy for the study of the CMTC in cell populations cultured on an artificial substrate.

Alkaloids↗