Ha-ras gene codon 12 mutation and DNA ploidy in urinary bladder carcinoma.
Explore the source record for details and available documents.
Codon↗
Biomedical subjects
F Herz
Publications and source records attributed to F Herz.
Quantitation of oncogene products by computer-assisted image analysis and flow cytometry.
The use of antibodies permits the study of oncogene product expression in cells and tissues. However, quantitation of the levels of expression in immunohistochemical preparations is beset by difficulties, and the available scoring system provide semiquantitative data at best. Here we describe the use of computer-assisted image analysis for determination of oncoprotein levels in a model system and compare the results with those generated by flow cytometric analysis. The oncogene products measured are located in the nucleus (c-myc p62 and c-fos p55), the inner surface of the membrane (c-ras p21), and both sides of the membrane (c-erbB-2 p185). In each instance, both analytic modalities yielded concordant results. Our data indicate that computer-assisted image analysis is a useful tool for quantitating cell components in immunohistochemical preparations.
Breast Neoplasms↗
Modulation of alkaline phosphatases in LoVo, a human colon carcinoma cell line.
LoVo, a continuous cell line derived from a human colon carcinoma produces two alkaline phosphatases: the heat-labile, L-homoarginine-insensitive, intestinal form, characteristic of its tissue of origin and the heat-stable, term-placental form, ectopically produced by a variety of tumors. Under basal conditions the activity levels of both enzymes are similar. Hyperosmolality and sodium butyrate induce increased levels of activity of the two alkaline phosphatases in a disparate fashion; whereas hyperosmolality augments the activity of both to the same extent, the effect of butyrate is more pronounced on the activity of the intestinal enzyme. When the two inducers are combined, induction of term-placental alkaline phosphatase is additive and that of the intestinal enzyme is synergistic. The effect of hyperosmolality is blocked by cycloheximide, and induction by sodium butyrate is inhibited by thymidine, cordycepin and cycloheximide. The known alkaline phosphatase inducer, prednisolone, has no effect on the enzymes of LoVo cells. Our results suggest that in these tumor cells the activity levels of the closely homologous term-placental and intestinal alkaline phosphatases appear to be independently controlled.
Alkaline Phosphatase↗
Expression of ras oncogene p21 protein in early gastric carcinoma and adjacent gastric epithelia.
The expression of the ras gene product p21 in normal gastric mucosa, early gastric carcinoma of diffuse (gastric) and intestinal types, and in adjacent mucosal abnormalities is reported. The analysis was performed on paraffin sections by an immunohistochemical assay using the mouse monoclonal antibody RAP-5 and the rat monoclonal antibody Y13-259. Expression of ras p21 was assessed by staining intensity and percentage of positively stained cells. In comparison to normal gastric mucosa of non-cancer patients, p21 was overexpressed in nearly all early carcinomas of both types and in the dysplastic and/or metaplastic mucosal alterations accompanying intestinal type of gastric cancer. Increased p21 expression was also observed in the normal-appearing mucosa adjacent to early carcinomas of diffuse type, but not in the morphologically normal gastric epithelium adjacent to the intestinal type. The results of this investigation suggest that ras p21 overexpression may be related to early events of human gastric carcinogenesis. The study supports the notion of different pathways in the development of diffuse (gastric) and intestinal types of gastric carcinomas.
Adenocarcinoma↗
Predictive value of DNA measurements in bladder washings. Comparison of flow cytometry, image cytophotometry, and cytology in patients with a past history of urothelial tumors.
Comparative DNA ploidy measurements were carried out by flow cytometry and by image analysis on cells in 71 bladder washing specimens from 50 patients with past histories of bladder tumors. Among the specimens classified as diploid or questionable by flow cytometry, 14 showed the presence of aneuploid DNA values documented by image analysis. In 18 of the 50 patients, recurrent tumors were observed during a relatively brief period of follow-up. In 15 of them the DNA pattern was aneuploid and in three it was questionable. In nine of the 15 patients, both methods of DNA analysis disclosed aneuploidy, but in six patients aneuploidy was detected by image analysis only. A combination of DNA aneuploidy, whether observed by flow cytometry, image analysis, or both, and of positive or suspicious urine cytology is highly predictive of recurrence of high grade bladder tumors. Image analysis of DNA content in bladder washings adds information of clinical value above and beyond that obtained by flow cytometry.
Aged↗
Effect of human serum on alkaline phosphatase induction in cultured human tumor cells.
The continuous cell lines T 24 and HT-29, derived from human bladder and colon carcinomas, produce term-placental and intestinal alkaline phosphatase, respectively. Growth in hyperosmolar medium or exposure to prednisolone or sodium butyrate induces increased enzyme levels, and combinations of inducers elicit synergistic activity increases. The effect of the inducing agents is strikingly diminished when cells are grown in the presence of high concentrations of human serum, and the synergistic increases are essentially abolished. Major human serum protein fractions do not affect alkaline phosphatase induction.
Acid Phosphatase↗
Expression of ras oncogene p21 protein in relation to regional spread of human breast carcinomas.
The oncogenes most frequently detected in human tumors belong to the ras gene family (Ha-ras, Ki-ras, and N-ras). These genes encode a group of closely related 21,000 dalton proteins termed p21. An immunohistochemical study of ras p21 expression was carried out on paraffin sections of 54 human breast carcinomas using monoclonal antibodies to p21. The control group consisted of ten cases of benign fibrocystic disease. The p21 expression was significantly higher in cancer cells than in epithelial cells of control specimens. No correlations, however, were observed between oncogene product expression and tumor size, histologic type, or grade. As a group, tumors with axillary lymph node metastases expressed higher levels of ras p21 than nonmetastasizing tumors. However, because of the significant overlap in individual p21 values, it is unlikely that the immunohistochemical assay for p21 could be used to predict the behavior of mammary carcinomas.
Breast Neoplasms↗
The use of 5-fluorocytosine and ketoconazole in the culture of the erythrocytic stages of Plasmodium falciparum and some tumor cell lines.
In vitro culture systems are often contaminated by bacteria and fungi. It is therefore often necessary to supplement culture media with agents such as penicillin/streptomycin, gentamycin or amphotericin B. The latter cannot be used in the in vitro culture of erythrocytic stages of P. falciparum, and thus anti-fungal agents have not been regularly used in this system. We describe the prophylactic use of 5-fluorocytosine (5-FC) and ketoconazole (KTZ) in tissue cultures at concentrations up to 300 and 10 micrograms/ml respectively which have no effect on the growth of P. falciparum (FCR-3 strain). A melanoma cell line (C32) and a line of uterine carcinoma (C41) were also unaffected by similar concentrations of 5-FC and KTZ. When dissolved in complete culture medium (RPMI 1640) with 10% human plasma, the minimum inhibitory concentration of 5-FC for a susceptible strain of Candida remained below 2 micrograms/ml. These experiments suggest that 5-FC (at 50 micrograms/ml) alone or in combination with KTZ (at 1 microgram/ml) is a useful addition to the armamentarium of antimicrobials available to the tissue culture biologist for a variety of cell culture systems.
Animals↗
Caution about assay of alkaline phosphatase to quantitate cell numbers.
Explore the source record for details and available documents.
Alkaline Phosphatase↗
Flow cytometric measurements of DNA and other cell components in human tumors: a critical appraisal.
Fundamental principles of flow cytometry with emphasis on DNA measurements and cell cycle analysis in human cells and tissues are summarized. Some of the pitfalls of cell preparation techniques and histogram interpretation are discussed at length. While consensus has been reached for some organs and tumors that DNA quantitation by flow cytometry (or image cytometry) may be of prognostic value, for most cancers studied to date the information remains incomplete. Thoroughly lacking are well-structured prospective studies because retrospective studies, while suggestive, may not necessarily be of the same value. Potential usefulness of other tumor markers is briefly discussed. Many fundamental questions concerning definitions of "diploid" and "aneuploid" tumors have not been satisfactorily settled. While the goal of "objective measurements" is worthy of further pursuit, the interpretation of results is often highly subjective. The biologic reasons for behavioral differences between diploid and aneuploid tumors are still totally obscure.
Cell Cycle↗
Cell cycle-dependent reactivity with the monoclonal antibody Ki-67 during myeloid cell differentiation.
The specificity and sensitivity of the monoclonal antibody Ki-67 in identifying proliferating cell compartments was tested with the human promyelocytic leukemia cell line HL-60 using multi-parameter flow cytometry. While correlated measurements of DNA content and Ki-67 immunofluorescence indicated that the antigen was present in all phases of the cell cycle, reactivity with the antibody was highest in proliferating S and G2+M cells. The analysis of the BrdU content of cells sorted on the basis of reactivity with Ki-67 showed a correlation between Ki-67 reactivity and BrdU uptake. In HL-60 cells induced to differentiate with dimethyl sulfoxide (DMSO), the loss of reactivity with Ki-67 paralleled the exit of cells from the cell cycle. This was not observed in DMSO-resistant HL-60 cells. These results validate the usefulness of the Ki-67 antibody for determining the proliferative stage of mammalian cells in culture.
Antibodies, Monoclonal↗
ras oncogene p21 as a tumor marker in the cytodiagnosis of gastric and colonic carcinomas.
This study was undertaken to determine whether the expression of ras oncogene product p21 can be used as a tumor cell marker of gastric and colonic carcinoma in brush smears. To detect p21 an immunocytochemical assay with RAP-5 monoclonal antibody was used. Benign epithelial gastric cells obtained from normal gastric mucosa or benign gastric lesions reacted negatively in 12 out of 13 cases. Similarly, benign epithelial colonic cells from normal colon or benign colonic lesions were negative for p21 in nine out of ten cases. Weakly positive reaction, confined to a few cell clusters only, was observed in one smear of a benign gastric ulcer and one smear of chronic ulcerative colitis. All 20 smears from colonic carcinoma and all 20 smears of gastric carcinoma contained cells that stained positively for p21, and the degree of tumor differentiation had no impact on the staining pattern. The results recorded in this study show that the immunocytochemical assay for the ras oncogene product may prove to represent a new tool for the cytodiagnosis of gastric and colonic carcinomas.
Antibodies, Monoclonal↗
Modification of Ha-ras oncogene p21 expression and cell cycle progression in the human colonic cancer cell line HT-29.
Multiparameter flow cytometric measurements of the Ha-ras oncogene product, Ha-p21, versus DNA content were used to study the effect of prednisolone, sodium butyrate, and hyperosmolality on the expression of this gene during the cell cycle of HT-29, a human colonic carcinoma cell line. In control cells the expression of Ha-p21 was cell cycle dependent; it increased during G1 and remained approximately constant as cells traversed the S- and G2 + M phases. Two compartments of G1 cells, one expressing low (G1A) and the other (G1B) high levels of Ha-p21 could be identified. Cells grown with prednisolone (1.4-2.1 microM) expressed higher Ha-p21 levels than controls. Cell cycle analysis revealed that this effect was accompanied by a change in the distribution of cells in G1 phase: whereas the proportion of cells in G1A was reduced, that of cells in G1B was increased. The steroid had no detectable effect on cells in S and G2 + M. By contrast, sodium butyrate and hyperosmolality caused a marked decrease in Ha-p21 content. This reduction was not accompanied by any modification of the proportion of cells in the cell cycle compartments. These results would suggest that Ha-p21 is not likely to be a primary regulator of cell cycle progression in HT-29 cells.
Butyrates↗
DNA distribution patterns in early gastric carcinomas. A Feulgen cytometric study of gastric brush smears.
DNA distribution patterns were studied by cytophotometry in 20 Feulgen-stained brush smears of early gastric carcinomas. Two different DNA distribution patterns, classified as predominantly diploid and aneuploid, were correlated with histologic data. Six of seven carcinomas of diffuse type were predominantly diploid. By contrast, 11 of 13 intestinal type carcinomas were aneuploid and two were diploid. Since the DNA distribution patterns recorded in this study were previously observed in advanced gastric carcinomas, this would suggest that the basic genetic make-up of the tumors does not change with tumor progression.
Adult↗
Expression of Ha-ras oncogene p21 protein in relation to the cell cycle of cultured human tumor cells.
It has been postulated that the expression of the product (p21) encoded by the ras genes may have a role in cell cycle events. Simultaneous multiparameter flow cytometry was used to measure the p21 content in relation to the cell cycle of several cancer cell lines of human origin. These studies revealed that p21 levels rise during the G1 phase of the cycle and remain approximately constant as cells traverse the S and G2 + M phases. The threshold level of p21 expression of S phase cells was used to divide the G1 cell population into cells with low (G1A) and high (G1B) p21 content. The p21 levels of G1B cells were approximately ten times higher than those of G1A cells. The validity of this subdivision was confirmed by synchronous measurements of RNA content and p21. Cells with low RNA content, hence in early part of G1 phase of the cell cycle, expressed low levels of p21, and cells with higher RNA content expressed higher levels of p21. These observations suggest that the levels of p21 are much lower at the onset of the cell cycle than at its end; hence a drop in p21 expression is likely to occur during or immediately after mitotic division.
Cell Cycle↗
Chromosome abnormalities in meningiomas.
Cytogenetic analyses of eight meningiomas grown in culture for 1 week are reported. Normal karyotypes were found in three cases and hypodiploidy in the remaining five. In the five hypodiploid meningiomas, one chromosome #22 was missing in four cases, and one case exhibited a 22q- deletion. In four of these five cases, chromosome #14 was either lost or altered. Chromosome #1 was lost or altered in three, and chromosome #6 in two. These findings lend further support for the association of total or partial loss of chromosome #22 in meningiomas and suggest the involvement of other chromosomes in the clonal evolution of these tumors.
Aged↗
Alkaline phosphatase isozymes in cultured human cancer cells.
Alkaline phosphatase, an ubiquitous enzyme is known to exist in several isozymic forms. At least three different isozymes have now been identified in humans. Alkaline phosphatase isozymes are among the substances synthesized ectopically by a variety of human tumors and many continuous cell lines derived from different cancers have retained the capacity to produce these membrane-located glycoproteins. This paper reviews the identification of alkaline phosphatase isozymes in cultured tumor cells and relates these findings with recent developments concerning these cell membrane located glycoproteins.
Alkaline Phosphatase↗
Flow cytometric identification of cancer cells in effusions with Ca1 monoclonal antibody.
The fluorescence of cells from 42 pleural and peritoneal effusions stained with Ca1 monoclonal antibody (Ca1MA) was studied by flow cytometry. In 14 of 17 malignant effusions a significantly higher intensity of fluorescence was observed in samples exposed to Ca1MA when compared with controls. There was no increase of fluorescence intensity in 25 benign effusions. The method failed in three malignant effusions: one due to endometrial carcinoma and two to malignant lymphoma. The sensitivity of the method was tested in experimental samples with a known percentage of malignant cells. The positive fluorescence with Ca1MA was detected in samples containing 0.1% of carcinoma cells. Flow cytometry with Ca1MA can be a relatively simple method of identification of malignant cells in effusions.
Antibodies, Monoclonal↗