Experiments were designed to assess age-related changes in the transplantability of human tumours xenografted in congenitally athymic (nu/nu) mice. It has been found that the number of progressively growing human tumour xenografts decreased significantly with increasing age of BALB/c nu/nu recipients. These findings, taken together with a previously recognized increase in the frequency of endogenous interleukin 2 (IL-2)-producing cells with age of nu/nu mice, prompted us to investigate whether administration of exogenous IL-2 to young adult nu/nu mice could change the transplantability of human tumours in the mice. Peritumoral administration of exogenous interleukin 2 to 8-week-old nu/nu mice inhibited the growth of the human tumour xenografts. In vitro activation of nu/nu splenocytes with exogenous Il-2 resulted in the generation of killer cells which have been found to be cytolytic when allowed to react with human tumour targets in 51Cr cytotoxicity assay. In addition, it has been found that the percentage of IL-2-activated Thy 1.2+ and ASGM1+ cells substantially increased with increasing age of nu/nu spleen cell donors. These findings are compatible with the hypothesis that the observed age-related decrease in takes of human tumour xenografts might be determined by the increasing level of IL-2 production and subsequent maturation of IL-2-dependent effector cells.

Aging↗

PubMedDate not supplied

Expression of polymorphic HLA-A,B epitopes in human urothelial cell lines.

Malignant human urothelial cell lines propagated in vitro have previously been demonstrated to express low amounts of monomorphic HLA-A,B,C as compared to premalignant urothelial cells. In this study the expression of polymorphic HLA-A,B epitopes in human urothelial cell lines have been investigated in greater detail. The expression of HLA-B locus coded epitopes in malignant TGrIII cells was demonstrated to be low or absent as compared to pre-malignant TGrII or slightly transformed TGrI cells, suggesting a mechanism by which malignant cells could escape from the host immune response. The extreme polymorphism of HLA-A,B,C antigens suggests that HLA typing could be used as a method to identify the origin of cell lines which is essential in the study of the process of malignant transformation in vitro, or when correlating in vitro data with clinical observations of the patient. Two urothelial cell lines classified as slightly transformed (TGrI) and two as pre-malignant (TGrII) could, according to their expression of polymorphic HLA-A,B epitopes, be identified as genuine independent cell lines. One TGrII cell line previously designated Hu1734 shared the same HLA-A,B phenotype as the genuine HCV29 (TGrII) cell line and is therefore suspected to be a subline of the latter. Out of 18 cell lines and sublines classified as TGrIII the fidelity of four (Hu1922 and three sublines of T24) was proved by their HLA-A,B phenotype. Mistaken identity either by contamination or false labelling of the cultures was suspected in samples of six TGrIII cell lines and sublines. In four of these the HLA-A,B type characteristic for the T24 cell line could be demonstrated, but in general HLA typing of TGrIII cell lines as a method to identify the origin of the individual cell lines failed, primarily due to the decreased expression of HLA-A,B,C antigens and the apparent selective loss of HLA-B locus coded antigens.

Cell Division↗

PubMedDate not supplied

Differences in adhesion of urothelial cells to a panel of substrates are characteristic for established human urothelial cell lines differing in their transformation grade.

Eight human urothelial cell lines established and propagated in the Fibiger Institute, differing in their transformation grade as described by Christensen et al were brought into suspension and contacted with a panel of seven substrates, ie. Con. A, Fibronectin, WGA, Collagen IV, Laminin, PNA and artificial basal membrane. Cell adhesion was measured on big cell populations with the use of flow cytophotometry, measured samples contained nonadhering cells and fluorescent beads, added as an internal standard. It was found that the cell lines differed from each other by the kinetics of cell adhesion to the chosen panel. A higher percentage of cells belonging to established cell lines adhered to PNA, WGA or Con A coated surfaces than to laminin, fibronectin or collagen IV. The difference in adherence of cells to fibronectin, laminin or collagen IV coated surfaces makes differentiation between the single cell lines possible. Discriminant analysis of the total patterns of adhesion of all cell lines was used for comparison. The effect of trypsinization on the structure of the cell surface, accompanied by possible changes in adherence, was evaluated by comparison of the immediate results with those obtained with cells after 3 hours of recovery at 37 degrees C. It was found that all cell lines showed a better adherence to laminin after recovery. This is consistent with the published data on the high sensitivity of laminin receptors to trypsinization. On the other hand, the receptors for PNA, WGA and Con A were less expressed after the recovery period, as evidenced by a decreased adherence of the cells to these three substrates. The effect of recovery of cells on the amount of their receptors for fibronectin and collagen IV was different in different cell lines, but the direction of change for this pair of receptors was identical in a given cell line. In some instances the recovery period had no effect whatsoever on cell adherence. The different pattern of adhesion of cells belonging to established cell lines to the chosen panel of substrates can be used as a parameter for characterization of single cell lines, but it does not correlate with their grade of transformation.

Cell Adhesion↗

PubMedDate not supplied

The Thomsen-Friedenreich antigen on human urothelial cell lines detectable by peanut lectin and monoclonal antibody raised against human glycophorin A.

The Thomsen-Friedenreich (TF) antigen is a cryptic disaccharide structure on human erythrocytes which can be exposed by neuraminidase treatment and which is supposed to be expressed in an unmasked form on some carcinoma cells. For its detection in addition to auto-, allo- and heteroantisera, PNA (peanut lectin) is being applied. In the present studies the mouse monoclonal antibody (MoAb) raised to asialoglycophorin from human erythrocytes was used. The MoAb 22.19 is of mouse IgM isotype and is specifically binding to beta-D Gal-1-3 alpha-D GalNAc. The human urothelial cell lines maintained and characterized earlier were analyzed using indirect immunofluorescence assays. Among the spectrum of cell lines tested, five out of six cell lines belonging to the transformation grade III category (invasive in vitro and tumorigenic in nude mice) expressed TF antigen. The relationship between expression of TF antigen and other earlier defined biological traits related to malignant phenotype is discussed.

Antibodies, Monoclonal↗

PubMedDate not supplied

In vitro studies of human bladder cancer.

Explantation and cultivation in vitro of 55 human biopsies of histologically normal urothelial tissue, and 63 biopsies from transitional cell carcinomas (TCC) revealed a primary success rate of about 75% and a secondary success rate of about 50% with no significant differences, between normal and TCC derived cultures. While only half of the subcultured biopsies of normal origin were able to grow for more than 4 passages all TCC derived cultures were able to do so. Based on morphology, life span, invasive growth into co-cultured fragments of embryonic chick heart, and tumorigenicity in nude mice the following transformation grades (TGr) were defined: Normal (TGr 0), non-malignant with prolonged, but finite life span (TGr I), premalignant with infinite life span but without tumorigenic and invasive potential (TGr II), malignant with infinite life span and tumorigenic and invasive properties (TGr III). Differentiation between TGr I - TGr III was possible by several other parameters, including cytological characteristics, molecular and immunological surface structure, relative oncogene expression, pyruvate kinase isoenzymes, DNA Repair, and growth pattern. However, in addition to these common characteristics, the individual cell lines of the various categories of transformation showed differences that allowed the identification of well defined cell lines and sublines.

Animals↗

PubMedDate not supplied

Growth requirements and growth pattern of human urothelial cell lines of different grades of transformation.

Human urothelial cell lines propagated in culture have been classified into three grades of transformation (TGr I-III). This classification is unrelated to the histological classification of the original biopsy material. Mortal TGr I cell lines differed from immortal and nontumorigenic TGr II cell lines and tumorigenic TGr III cell lines; they have a lower growth rate, growth fraction, and saturation density, and a higher requirement for Ca2+. Except for one TGr II cell line, TGr II cells differed from TGr III cells by having a higher serum requirement and a lower saturation density. The ability to grow in agar was not correlated with the TGr of the cell lines.

Blood Physiological Phenomena↗

PubMedDate not supplied

Chromosomal characterization and isoenzyme pattern of non-malignant and malignant human urothelial cell lines.

The karyotypes and selected isoenzymes of five cell lines, HU 961b, HU 1922, HU 609, HCV-29 and HU 1703 He, were characterized. These cell lines were derived either from histologically normal human urothelial cells or from transitional cell carcinoma (TCC) of the urinary bladder. They displayed different life span in vitro and transplantability in nude mice, as well as a unique isoenzyme and chromosomal pattern. In addition, various chromosomes were involved in the formation of marker chromosomes in these cell lines. Since there was a lack of consistency in chromosomal changes in the cell lines with TCC origin, the cause of malignancy could not be attributed to the abnormality of specific chromosomes.

Carcinoma, Transitional Cell↗

PubMedDate not supplied

A classification of human urothelial cells propagated in vitro.

Human urothelial cell lines of non-malignant and malignant origin were classified into three grades of transformation, i.e. TGr I, TGr II, and TGr III, which differ from primary and early cultures of normal bladder mucosa (TGr 0) by growth in the absence of fibroblasts, prolonged survival, increasing chromosomal abnormalities, and antigenic modifications. Non-malignant pure epithelial cell lines with a prolonged, but finite life span (18-38 passager in vitro) were classified as TGr I. These cells were characterized by chromosomal counts showing a considerable variation but with a near-diploid mode and a smaller size but otherwise normal morphology. TGr II cells differed from TGr I by infinite growth transformation and wide morphological variability. TGr III cells were above all characterized by their ability to produce tumors in nude mice and to invade and destroy fragments of embryonic heart in vitro. These immortalized cells were small with a large nucleus and a uniform polygonal shape. Chromosomal counts showed pronounced aneuploidy with an increased number of chromosomes.

Aged↗