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Biomedical subjects

W H Lewis

Publications and source records attributed to W H Lewis.

At least 73 records · Page 4Linked to original sources
PubMed1985

Gene order on the short arm of human chromosome 11: regional assignment of the LDH A gene distal to catalase in two translocations.

Leukemic cells with reciprocal translocations involving 11p13 and 14q13 were obtained from two patients with T-cell acute lymphoblastic leukemia and fused with mouse Ltk- cells. DNA from independent hybrid clones was screened by Southern blot and hybridization to molecular probes for the human catalase and Ha-ras-1 genes. Several clones showed segregation of these two genes, indicating the presence of either the der11 or der14 human chromosomes. When DNA from these hybrid clones was examined for the presence of the human genes for calcitonin and gamma-globin, both genes were found to segregate with the Ha-ras-1 gene and the der14 chromosome indicating that they lie distal to catalase. When the hybrid clones were examined for the presence of human lactate dehydrogenase A (LDH A) activity, only those clones containing the der14 chromosome expressed activity indicating that the LDH A gene is also distal to catalase on the short arm of chromosome 11.

Animals↗
PubMed1985

Establishment of mouse cell lines homozygous for temperature-sensitive mutation in catalase gene.

Contact-inhibited and-transformed fibroblast cell lines have been established from the homozygous C3H/CSb mutant mouse. These cell lines have low levels of a temperature-sensitive catalase enzyme activity. When compared to wild-type C3H cells, the catalase-deficient cells are markedly more sensitive to the toxicity of hydrogen peroxide. Furthermore, this cellular sensitivity to external hydrogen peroxide is temperature sensitive. These cell lines will be of use in the study of the role which catalase plays in the intracellular prevention of transformation induced by oxidative stress.

Acatalasia↗
PubMed1985

Human Ia molecules carrying MT1 determinants: purification with a monoclonal antibody.

A monoclonal antibody 77.34, reactive with polymorphic HLA class II molecules, was produced. The allotype specificity of this IgG2a antibody was analyzed by cytotoxicity, flow cytometry, and cellular radioimmunoassay. Cytotoxic reactivity on a panel of B cells from 88 unrelated individuals was concordant with the MT1 (DC1) allospecificity (r = 0.83). Immunoanalysis by flow cytometry showed that cells from MT1+ homozygous cell lines were reactive, whereas MT2+ and MT3+ homozygous cells were not. A cellular radioimmunoassay performed under saturating conditions indicated that three MT1+ cell lines bound 14-45 X 10(5) molecules of antibody per cell representing 30-40% of the amount detected with monomorphic anti-DR monoclonal antibody 21w4. The subset of molecules bearing the MT1 allospecificity was purified with a 77.34 IgG immunoadsorbent. The purified molecules were antigenically reactive with several antibodies directed at DQw1 molecules but were devoid of reactivity to monomorphic anti-DR antibodies. Two-dimensional gel electrophoresis showed that the alpha subunit is composed of several acidic spots of Mr 32,000 whereas the beta subunit was seen as a single spot of Mr 25,000, corresponding to DQw1 molecules. DR molecules purified by monoclonal antibody affinity were unreactive with 77.34 antibody. All of the 77.34 reactivity was observed with the fractions depleted of DR molecules. Two-dimensional gel analysis showed marked differences between the purified DR and DQw1 molecules. The presence of the MT1 determinant on Ia molecules referred to as the DQw1 molecules and distinct from those bearing the DR epitopes was confirmed on two DR1, MT1 homozygous cell lines. Thus, DQw1 molecules can be purified away from DR1 molecules by affinity chromatography to 77.34 IgG, specifically reactive with the MT1 (DQw1) allospecificity. The binding of 77.34 IgG to MT1+ cells was not inhibited by all monoclonal antibodies reported to be correlated with the MT1 allospecificity suggesting that the latter might be comprised of more than a single epitope.

Animals↗
PubMed1984

Isolation of human fibroblast catalase cDNA clones. Sequence of clones derived from spliced and unspliced mRNA.

We report the isolation and sequence of partial cDNA clones coding for human catalase. These clones were recovered from a human fibroblast cDNA library by screening with mixtures of oligonucleotide probes deduced from the amino acid sequence of human erythrocyte catalase. A comparison of their nucleotide sequence with the known protein sequence and mapping of homologous DNA sequences to the short arm of chromosome 11 in somatic cell hybrids confirmed that they coded for catalase. One of these clones contained a 462-base insertion interrupting the coding sequence with stop codons in all three reading frames. The 5' and 3' ends of the insertion correspond to the donor and acceptor consensus sequences of introns. Inspection of clones lacking the insertion confirm the location of the splice sites. We suggest this clone corresponds to the product of reverse transcription of an unspliced mRNA species. The catalase gene is the closest genetic marker mapped to Wilms tumor, one of the most prevalent of childhood cancers. Catalase cDNA probes will be useful to the examination of mitotic recombination in the etiology of this disease and may provide a useful starting point to the search for the putative Wilms tumor gene.

Amino Acid Sequence↗
PubMed1983

Chromosome-mediated gene transfer with the Chinese hamster ovary cell line.

Using an improved method of chromosome-mediated gene transfer, we have investigated transfer of the codominantly expressed methotrexate-resistant dihydrofolate reductase (MtxRIIIdhfr) gene into Chinese hamster ovary (CHO) cell recipients. The frequency of dhfr gene transfer with CHO cells varied considerably from clone to clone, ranging from 4 X 10(-7) to 5 X 10(-5). Using appropriate cell recipients we were able to test for linkage of several genetic markers available in the CHO cell line. For example, the mutation resulting in the auxotrophic glyB-CHO cell line has been reported by others to be linked to the dhfr gene. However, we could not demonstrate cotransfer of these two markers when glyB- recipient cells were treated with MtxRIII chromosomes and transformant clones were selected for either methotrexate-resistance (MtxR) or glycine prototrophy. We conclude that these two genes are not closely linked in the hamster genome. However, the genes for thymidine kinase (tk) and galactokinase (gk), which are known to be linked in mammalian genomes, were found to cotransfer into CHO recipients with a frequency of about 50%.

Animals↗
PubMed1983

No adaptogen response of mice to ginseng and Eleutherococcus infusions.

There was no significant difference in mortality, weight gain, liquid consumed, or pathogenesis between mice ingesting four different ginseng infusions for up to 96 days, and control mice drinking distilled water. These results suggest no toxicity or tea selectivity of mice subjected to such regimens. Mice that drank concentrated Siberian ginseng wuchaseng extract with sugar showed significantly more aggressive behavior than those drinking other infusions or distilled water. However, there was no significant difference in stamina or longevity between mice drinking infusions of two preparations of Siberian ginseng, Oriental ginseng, or American ginseng, and control mice when subjected to swimming trials in cold water 38, 46 and 96 days after treatment began. Consequently, ingestion of adaptogenic glycosides did not significantly affect the survival of mice under major environmental stress.

Adaptation, Physiological↗
PubMed1983

Chromosome-mediated gene transfer of hydroxyurea resistance and amplification of ribonucleotide reductase activity.

Metaphase chromosomes purified from a hydroxyurea-resistant Chinese hamster cell line were able to transform recipient wild-type cells to hydroxyurea resistance at a frequency of 10(-6). Approximately 60% of the resulting transformant clones gradually lost hydroxyurea resistance when cultivated for prolonged periods in the absence of drug. One transformant was subjected to serial selection in higher concentrations of hydroxyurea. The five cell lines generated exhibited increasing relative plating efficiency in the presence of the drug and a corresponding elevation in their cellular content of ribonucleotide reductase. The most resistant cell line had a 163-fold increase in relative plating efficiency and a 120-fold increase in enzyme activity when compared with the wild-type cell line. The highly hydroxyurea-resistant cell lines had strong electron paramagnetic resonance signals characteristic of an elevated level of the free radical present in the M2 subunit of ribonucleotide reductase. Two-dimensional electrophoresis of cell-free extracts from one of the resistant cell lines indicated that a 53,000-dalton protein was present in greatly elevated quantities when compared with the wild-type cell line. These data suggest that the hydroxyurea-resistant cell lines may contain an amplification of the gene for the M2 subunit of ribonucleotide reductase.

Animals↗
PubMed1981

Group-specific component (Gc) subtypes in the Chinese population of Hong Kong.

The distribution of Gc subtypes in a sample of the Chinese population of Hong Kong was studied using isoelectric focusing followed by immunofixation. A sensitive modification of this technique is described. Nine distinct phenotypes were observed which appear to result from the three common alleles Gc1F, Gc1S, and Gc2, which are found in most populations. The respective gene frequencies were 0.494, 0.258, and 0.247. In addition, two rare phenotypes were observed which appear to be due to a rare allele tentatively identified as Gc2C2.

Adult↗
PubMed1980

Parameters governing the transfer of the genes for thymidine kinase and dihydrofolate reductase into mouse cells using metaphase chromosomes or DNA.

The conditions necessary to achieve high frequency transfer of the thymidine kinase and dihydrofolate reductase genes from hamster cells into mouse cells were investigated. Of the parameters examined, the length of adsorption time, input gene dosage, and treatment with dimethylsulfoxide (DMSO) were found to significantly alter the transfer frequency using either metaphase chromosomes or purified DNA as the transfer vehicle. With the mouse cell line as a recipient, the optimal adsorption period for DNA or chromosomes from MtxRIII cells was found to vary from 8 to 16 h in those experiments where the recipient cells were subsequently treated with DMSO. Without DMSO, similar frequencies could be obtained by extending the period of adsorption. Increasing the dosage of DNA or chromosomes resulted in an almost linear increase in the number of transformants. The optimal conditions for transfer did not significantly differ for the two genes studied. On the average, the optimal conditions yielded 1.5 x 10(3) transformants per 10(7) recipient cells with chromosomes; with DNA an average of only 60 transformants were observed. In general, DNA transformants grown in the absence of methotrexate were unstable; whereas, under the same conditions about 20% of the transformants from the chromosome experiments were stable.

Animals↗
PubMed1980

Somatic cell genetics: a review of drug resistance, lectin resistance and gene transfer in mammalian cells in culture.

During the past decade development of selection procedures for isolating a large variety of altered stable phenotypes in mammalian cell culture has been extremely rapid. The list now includes temperature sensitive, auxotrophic, drug resistant and lectin resistant cell lines, as well as cultures containing altered products of differentiation. As a general class, the drug resistant mutants have been studied more often than any of the others because of the wide variety of very interesting and specifically altered mutants that can be isolated, and the ease with which they can usually be obtained. In this review we discuss the essential features of the best studied drug and lectin resistant systems. Also a section on gene transfer has been included since the new technology being developed in this area is certain to have great influence on our understanding of fundamental aspects of somatic cell variation.

Amanitins↗