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Biomedical subjects

T R Chen

Publications and source records attributed to T R Chen.

At least 19 recordsLinked to original sources
PubMed1992

Karyotypic derivation of H9 cell line expressing human immunodeficiency virus susceptibility.

BACKGROUND: The T-cell lymphoma HUT78 cell line and the H9 clone of the CD4-positive HT cell line, derived from HUT78, are known to be genetically identical on the basis of allozyme (allelic isozyme) patterns and DNA fingerprinting, but the chromosome compositions of these cell lines have not been determined. The HT cell line and its H9 clone carry susceptibility to the human immunodeficiency virus (HIV). PURPOSE: The purpose of this study was to examine specific chromosome changes linking the expression of the HIV susceptibility that is unique to H9 cells by characterizing the karyotypes of the HUT78 and the H9 cell lines, by comparing chromosome differences between these lines, and by evaluating the relationships between them. METHODS: Air-dried chromosome slides were prepared, and the fast Giemsa-trypsin method was used to delineate G-bands. The numerical distribution of chromosomes from the two cell lines was determined, and complete chromosome analysis in 47 HUT78 metaphases and 21 H9 metaphases with discernible G-banding was used to specify the modal karyotypes of each cell line. RESULTS: Both cell lines had complex karyotypes that contained 47%-65% structurally modified marker chromosomes. The distribution of numbers of chromosomes in HUT78 cultures was broad. There were two distinct modal numbers of chromosomes at 42 and 43 and at 73, resulting from the presence of three actively growing sublines: two hypodiploid (1s) sublines (HUT78/1sA and HUT78/1sB) and a hypertriploid (2s) subline (HUT78/2s), respectively. HUT78/1sA and HUT78/1sB differed by five reciprocal chromosome replacements, and HUT78/2s had double copies of most of the chromosomes in 1s cells. H9 and HUT78/2s had had 21 matching markers, 10 of which were paired; contained 21 chromosomes with the same number of copies; consistently lacked eight homologous normal chromosomes; and showed close modal numbers of chromosomes at 69 and 73, respectively. CONCLUSIONS: Derivation of the 2s subline from the 1s subline by polyploidization is apparent. All lines apparently had the same progenitor cell population, and HUT78/2s represents an intermediate linking HUT78/1s to H9. IMPLICATIONS: These data should be useful for studying specific chromosome changes linking the expression of the HIV susceptibility unique to H9 cells. Karyologic studies such as those presented here can provide data (a) to clearly identify the clonal cell population of a subline, (b) to examine relationships among sublines of the same progenitor, and (c) to provide a clue that may link a subtle chromosome change to a phenotypic expression.

Chromosome Aberrations↗
PubMed1992

Use of the polymerase chain reaction for specific detection of type A, D and E enterotoxigenic Staphylococcus aureus in foods.

By comparison with the DNA sequences coding for Staphylococcus aureus enterotoxins (ents) A, B, C, D and E, oligonucleotides unique to the entA, entD and entE genes were synthesized and used as polymerase-chain-reaction (PCR) primers for the specific detection of type A, D or E enterotoxigenic S. aureus. The relative molecular weights of the PCR products amplified with these primers were 210, 333 and 456 bp, respectively. Despite the high relatedness among these S. aureus enterotoxin genes, each primer pair allows specific detection with total discrimination from other types of enterotoxigenic S. aureus. DNA from non-enterotoxigenic S. aureus or from non-S. aureus would not interfere with the PCR results either. Primers designed for entE detection allow the discrimination of entE strains that when assayed by a serological method might be classified as entA-producing strains. Study of the detection sensitivity showed that by using these primers, DNA from 10(0) cells of enterotoxigenic S. aureus could be detected unambiguously. When these oligonucleotide primers were used for the detection of S. aureus in foods, 10(0)-10(1) cells per gram of food could be detected and the naturally contaminating microflora in the food sample did not interfere with the detection.

Base Sequence↗
PubMed1990

Establishment and characterization of a human adrenocortical carcinoma cell line that expresses multiple pathways of steroid biosynthesis.

We established a continuous cell line, NCI-H295, from an invasive primary adrenocortical carcinoma. The cell line was established in a fully defined medium (HITES) and later could be adapted for growth in a simple medium supplemented only with selenium, insulin, and transferrin and devoid of serum, steroids, fibroblast growth factor, and a source of exogenous cholesterol. NCI-H295 cells had a relatively long population doubling time and were tumorigenic when inoculated s.c. into athymic nude mice. The cultured cells had ultrastructural features of steroid-secreting cells and contained complex cytogenetic abnormalities including the presence of multiple marker chromosomes. Steroid analyses (radioimmunoassays and mass spectrometry), performed 7 to 9 years after culture initiation, demonstrated secretion of more than 30 steroids characteristic of adrenocortical cells. Total unconjugated steroid secretion in serum-supplemented medium was 2.83 micrograms/10(6) cells/24 h and about 4-fold less in serum-free medium. The major pathway of pregnenolone metabolism in NCI-H295 cells is androgen synthesis, with formation of dehydroepiandrosterone, androstenedione, testotesterone, and at least three sulfated androgens, as well as estrogens. In addition, formation of cortisol, corticosterone, aldosterone, and 11 beta-hydroxyandrostenidione indicated the presence of 11 beta-hydroxylase. Thus, multiple pathways of steroidogenesis are expressed by NCI-H295 cells, including formation of corticosteroids, mineralocorticoids, androgens, and estrogens. Our findings indicate the presence in NCI-H295 cells of all of the major adrenocortical enzyme systems, including 11 beta-hydroxylase, desmolase, 21 alpha-hydroxylase, 17 alpha-hydroxylase, 18-hydroxylase, lyase, sulfokinase, and aromatase. The NCI-H295 cell line should prove of value in studying the regulation, metabolic pathways, and enzymes involved in steroid formation and secretion. In addition, it may provide insights into the biology and treatment of adrenocortical carcinoma.

Adrenal Cortex Neoplasms↗
PubMed1990

Characteristics of cell lines established from human gastric carcinoma.

We report the establishment and characterization of four continuous cell lines derived from human primary and metastatic gastric carcinomas, and we compare their properties with a panel of colorectal carcinoma cell lines previously established and reported by us. Our success rate in culturing gastric carcinomas was relatively low, especially from primary tumors, compared to colorectal carcinoma. These observations may reflect the relatively modest number of gastric carcinoma cell lines established (mainly from Japan), compared to the abundance of colorectal carcinoma lines established worldwide. All four gastric lines expressed the surface glycoproteins carcinoembryonic antigen and TAG-72 and three lines expressed CA 19-9. Two of the lines expressed aromatic amino acid decarboxylase but lacked other markers for neuroendocrine differentiation. All four lines were positive for vasoactive intestinal peptide receptors but lacked gastrin receptors. In addition, two lines expressed receptors for muscarinic/cholinergic receptors but not beta-adrenergic receptors. Cytogenetic evidence for gene amplification was present in the cell lines. All four lines contained varying numbers of double-minute chromosomes. One line, SNU-16, was amplified for the c-myc proto-oncogene and contained four homogeneously staining regions. While c-myc and c-erb-B-2 RNA were expressed by all lines, there was no evidence of amplification or overexpression of several other proto-oncogenes and growth factors. The multiple properties we have described in our gastric carcinoma cell lines are remarkably similar to those found in the panel of colorectal carcinoma cell lines. These properties include morphology, growth characteristics, expression of surface glycoproteins, partial expression of neuroendocrine cell markers, frequent chromosomal evidence of gene amplification, and occasional amplification of the c-myc proto-oncogene. Our four well characterized cell lines should provide useful additions to the modest number currently available for in vitro studies of gastric carcinoma.

Antigens, Neoplasm↗
PubMed1989

Mycoplasma infection of cultured cells.

Mycoplasma contamination is tough to detect and even more difficult to eradicate. It is best to start over fresh from clean cell stocks, but several elimination options are available.

Animals↗
PubMed1988

SK-UT-1B, a human tumorigenic diploid cell line.

SK-UT-1B (ATCC HTB 115) produced tumors 2 weeks after subcutaneous injection of 10 million cells into athymic nude mice. Karyotypic analysis at the 500-600 subband level, however, did not reveal any noticeable aberrations. This cell line therefore may represent the first example of experimentally proven human tumorigenic cells of solid tumor origin that retains the diploid karyotype in vitro indefinitely.

Aged↗
PubMed1988

Re-evaluation of HeLa, HeLa S3, and HEp-2 karyotypes.

In contrast to earlier reports, this study on HeLa, HeLa S3, and HEp-2 revealed that karyotypes of each cell line are characterized by a consistent marker chromosome composition and a constant number of copies for both normal and marker chromosomes. Based on these chromosome fingerprints and an analysis of 50 metaphases, the modal karyotype of each cell line was defined. Each modal karyotype had the composite content of the previously reported karyotypes of the same cell line, and, generally, the former had the same or a higher number of copies per chromosome than the latter. This modal karyotype can be used as a standard to identify and further individualize both the cell line itself and a subline within that cell line. We have also found that many cells within each cell line have the same karyotype. Portions of numerical data are compiled in a chart format by which the extent of chromosome differences between cultures can readily be compared. Also discussed in brief are characteristic chromosome changes that may help distinguish clonally derived cell lines from lines derived by cross-contamination.

Cell Line↗
PubMed1987

Characteristics of cell lines established from human colorectal carcinoma.

We have characterized 14 human colorectal carcinoma cell lines established from primary and metastatic sites by us during the years 1982 to 1985. Five lines were established in fully defined ACL-4 medium and 9 in serum supplemented R10 medium. However, after establishment, cultures could be grown interchangeably in either medium. The lines grew as floating cell aggregates in ACL-4 medium, while most demonstrated substrate adherence in R10 medium. The lines had relatively long doubling times and low cloning efficiencies. Twelve were tumorigenic in athymic nude mice when injected s.c., and two grew i.p. as well. Based on culture, xenograft, and ultrastructural morphologies, the 14 lines could be subtyped as follows: 4 were well differentiated; 5 were moderately differentiated; 4 were poorly differentiated; and 1 was a mucinous carcinoma. Membrane associated antigens characteristic for gastrointestinal cells (carcinoembryonic antigen, CA 19-9, and TAG-72 antigens) were expressed by 50-71% of the lines. Lines expressing carcinoembryonic antigen and CA 19-9 actively secreted these antigens into the supernatant fluids while TAG-72 antigen was not secreted. Surprisingly, 5 of 7 of the original tumor samples tested and 13 of 14 cultured lines expressed L-dopa decarboxylase activity, which is a characteristic enzyme marker of neuroendocrine cells and tumors. In addition, one poorly differentiated cell line contained dense core granules, characteristic of endocrine secretion. Preliminary cytogenetic analyses indicated that 9 of 11 lines examined contained double minute chromosomes. In addition, 3 of the 9 lines with double minutes also had homogeneously staining regions. These findings indicate a high incidence of amplification of one or more as yet unidentified genes.

Antigens, Neoplasm↗
PubMed1987

Cytogenetics of somatic cell hybrids. II. Modal karyotypes and mosaicism in mouse mutant cell lines: LM(tk-), A9, and Rag.

Modal karyotypes of mouse mutant cell lines, LM(tk-), A9, and Rag, which are frequently used as a donor line for somatic cell hybridization, are described and compared with karyotypes previously reported by other laboratories. Results are summarized as follows. Many cells (18%-38%) in each line had the same chromosome composition as the modal karyotype. Chromosome differences from the modal karyotype in some cells mainly involved one or a few chromosomes only. These differences are mostly caused by structural changes from well documented mechanisms or simple numerical changes including artifacts arising from the rupture of cells. Karyotypes of coexisting subpopulations within the same cell line can be clearly identified. Assessment of karyologic properties of a heteroploid cell population may vary considerably depending on whether it is estimated on a subpopulation or the total population as a unit. Modal karyotypes of these cell lines contained most marker chromosomes of, and a similar chromosome composition to those previously reported by other laboratories. These cell lines exhibit karyotypic constancy; thus, we believe that chromosome changes associated with clonal growth of somatic cell hybrids can be identified without ambiguity.

Animals↗
PubMed1987

WiDr is a derivative of another colon adenocarcinoma cell line, HT-29.

This paper documents chromosomal and isozymic evidence indicating that WiDr is a derivative of another colon adenocarcinoma cell line, HT-29, which was established in 1964. The phenotypes for seven isozymes studied were identical for all HT-29 and WiDr cell lines. The karyotypes were similar between four HT-29 cultures separated by more than 100 passages and two WiDr cultures that were studied independently in 1979 and 1985. The isozymic and karyotypic data are consistent with that described for the cell line WiDr, originally reported in 1979. Detailed karyotypic comparisons further show that cultures of WiDr possess all nine chromosomal markers found in the initial HT-29 (passage 19) cultures. However, both the higher passaged HT-29 and WiDr differ from the initial HT-29 (passage 19) because it has an additional five to eight constitutive marker chromosomes. An increase or decrease of one copy in several normal chromosomes is also noticed with more normal chromosome gains than losses. The study provides another example demonstrating the remarkable karyotypic stability within a heteroploid cell line with continued passage. It also reemphasizes the critical need for detailed karyotypic and isoenzymic studies on all cell lines in common use.

Adenocarcinoma↗
PubMed1987

Evidence for a multistep pathogenesis in the generation of tumorigenic cell lines from hemopoietic colonies exposed to Abelson virus in vitro.

The present studies were undertaken to investigate the ability of Abelson murine leukemia virus (A-MuLV) to transform cells derived in vitro from pluripotent hemopoietic progenitor cells of high proliferative potential. We now report that continuously growing, autonomous cell lines could be obtained from a high proportion of individually infected multilineage colonies generated in assays of spleen cells from normal adult mice if the infected cells were cocultivated for the first two to three months with irradiated NIH-3T3 cells. No lines were obtained if the 3T3 cell feeders were not initially present. Similar results were obtained when the cells exposed to virus were from multilineage colonies originating from isolated single cells obtained by replating small blast colonies. Characterization of the transformants and a number of derivative cloned sublines revealed the consistent presence of a mast cell phenotype, with some suggestion of macrophage differentiation in a few cases. All cell lines tested produced virus, showed a variable pattern of A-MuLV integration, and gave rise directly to tumors when injected subcutaneously, as shown by both Southern analysis and cytogenetic studies. The early absolute but transient dependence of these A-MuLV mast cell transformants on a fibroblast feeder suggests a multistep process in their evolution, in which the acquisition of autonomy from factors of mesenchymal cell origin may play an important role.

Animals↗
PubMed1985

Modal karyotype of human leukemia cell line, K562 (ATCC CCL 243).

The chromosome composition of the human leukemic cell line, K562, deposited at the American Type Culture Collection (ATCC) (CCL 243), was studied and further compared with those reported by other laboratories. In CCL 243, the modal karyotype had 55 normal and 14 constitutive marker chromosomes and occurred in nearly 50% of the cultured cell population. Other variant karyotypes were observed either at a low frequency in coexisting minor cell types or only once in all other analyzed metaphases. Normal chromosome #9 and the chronic myeloid leukemia (CML) specific Ph/t(9;22) were absent. It appears that karyotypes of most K562 cultures distributed world-wide are similar to each other, with the stable modal chromosome number, and therefore, the modal karyotype presented here should be useful in karyotypic identification of a cell line.

Cell Line↗
PubMed1985

The chinese hamster Don cell line differs from normal diploidy by one chromosome band.

G-banded and C-banded prometaphase and metaphase karyotypes of the Chinese hamster Don cell line from three different laboratories were compared. All the cell lines were pseudodiploid. Each consisted of two distinct cell populations designated S1 and S2. The chromosome complements of each of these populations differed from the diploid by only one additional chromosome band on a subterminal 1p in S1 and on a median 1q in S2. The sites of these extra bands were neither constricted nor heterochromatic. All other different karyotypes, including those from seven distinct subpopulations, had basic patterns of chromosome complements that were best represented by S1 or S2, respectively the major or minor cell type of the cell line. Moreover, nearly 80% of the metaphases analyzed had a modal chromosome number of 22, and about 60% had the same chromosome composition of a cell type or subtype. Our data also suggest that Don cells were probably pseudodiploid by 1964 or before.

Animals↗
PubMed1979

Cytogenetics of somatic cell hybrids. I. Progression of stemlines in continuous uncloned cultures of man-mouse cell hybrids.

Karyotypes of hybrid cells were studied in continuous uncloned cultures by Q- and C-bandings. Cultures were initiated by virus-mediated or spontaneous cell fusions from normal human diploid fibroblasts and mouse heteroploid RAG cells. Heterokaryons containing complete genomes of both parental cells randomly lost chromosomes from both species. The majority of cells in early growth stages, however, still possessed a nearly complete human genome. The rate of human chromosome loss in subsequent growth periods was not uniform, being gradual in some and rapid in others. The initially predominant 2n human-1s mouse (1h:1m) type was soon replaced by a less frequent 2n human-2s mouse (1h:2m) type. Over an increased period of time in mass culture, the number of stemlines decreased. One stemline, often a (1h:2m) type with a greatly reduced human complement, outgrew the others and occupied the entire culture. Therefore, the usual process of clonal isolation may confer a negative selection bias against cell hybrids retaining a large number of human chromosomes. Hybrid stemlines with stable karyotypes were established in the present HAT-agar selection system before 36 days after fusion had elapsed.

Animals↗
PubMed1977

Fluorescent C bands of human chromosomes with 33 258 Hoechst stain.

Air-dried preparations of human metaphase chromosomes normally exhibit a Q band fluorescence pattern with 33 258 Hoechst stain while the C band regions of 1,9,16, and most acrocentric short arms appear dull. If these stained and mounted slides are stored at room temperature in the dark for several days, a spontaneous change from C-negative to C-positive band some times occurs. We postulate that the pH of the buffered saline mounting medium during storage of the slides causes the C band shift since the results can be duplicated experimentally by lowering the pH of the mounting buffer from 5.5 to 4.0.

Buffers↗