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D Housman

Publications and source records attributed to D Housman.

126 records · Page 7Linked to original sources
PubMed1976

Induction of erythroid differentiation in vitro by purines and purine analogues.

The effectiveness of purines and purine analogues as inducers of erythroid differentiation in cultured murine erythroleukemia cells has been investigated. These cell lines have previously been shown to differentiate in vitro in response to dimethylsulfoxide (DMSO) and a number of other polar solvents. Two purine analogues, 6-thioguanine and 6-mercaptopurine, as well as the naturally occuring purine, purine, hypoxanthine, are shown to be extremely potent inducers. 6-Thioguanine is effective at a concentration of 0.06 mM, 750 fold lower than the DMSO concentration required for equivalent induction. 6-Mercaptopurine and hypoxanthine are effective inducers at a concentration of approximately 2 mM. Accumulation of globin mRNA was monitored during induction with purine inducers and shown to be similar in amount to globin mRNA levels reached in DMSO-induced cultures. Induction of differentiation by all three compounds follows a similar time course to induction with DMSO. All three compounds are potent inducers of HGPRT (hypoxanthine-guanine phosphoribosyltransferase)-negative cell lines; hence incorporation of purines into DNA is not required for induction of differentiation. Comparison of these compounds with other purines and purine analogues suggests a high degree of specificity in their interaction with a cellular target.

Cell Line↗
PubMed1976

Commitment to erythroid differentiation by friend erythroleukemia cells: a stochastic analysis.

A method for the clonal analysis of murine erythroleukemia cells has been developed which allows the precise characterization of the number of progeny produced by each cell and the degree of differentiation of each progeny cell. The potential of almost every cell in the culture can be monitored because a plating efficiency close to 100% has been achieved. The effects of treatment with an inducer of differentiation (DMSO) on the proliferative capacity of the treated cells have been studied with this technique. Cells from a mass culture treated with inducer give rise to colonies of differentiated progeny when subsequently cloned in the absence of inducer. Colonies exhibiting this phenotype represent the progeny of cells committed to the differentiation pathway by treatment with inducer. We observe that the commitment decision limits the subsequent proliferative capacity of the cell to four additional cell divisions. A quantitative analysis suggests that the commitment decision for each cell is made in a stochastic manner. Irreversible commitment to the expression of differentiated functions occurs with discrete probability per cell generation for many cell generations. The value for this probability is a function of the concentration of inducer (DMSO). A correlative biochemical study suggests that an irreversible commitment decision by a significant proportion of the population precedes or accompanies increases in cytoplasmic globin mRNA levels, one of the earliest detectable biochemical markers for erythroid differentiation in this system. A specific kinetic model based on these considerations has been developed to predict clonal phenotypes as a function of time and probability of commitment. Quantitative predictions based on this model are in excellent agreement with experimental observations. The effectiveness of a stochastic model in predicting the behavior of this system is discussed in relation to the stochastic behavior of normal hematopoiesis and the biochemical mechanisms which control these differentiation programs.

Cell Division↗
PubMed1976

Absence of messenger RNA and gene DNA for beta-globin chains in hereditary persistence of fetal hemoglobin.

The relative amounts of alpha-amd beta-globin mRNA and globin gene DNA were measured in reticulocyte RNA and lymphocyte DNA of an individual with homozygous hereditary persistence of fetal hemoglobin whose red blood cells contain 100% fetal hemoglobin (hb F: alpha2gamma2.) Molecular hybridization assays used as probes full-length DNA copies of human alpha- and beta-globin messenger RNA. The results of these hybridization assays demonstrated the expected amounts of alpha-globin mRNA and gene DNA, but absence of beta-globin mRNA and absence of beta-globin gene DNA. In the individual studied, hereditary persistence of fetal hemoglobin is associated with total deletion of the beta-globin structural gene.

Child↗
PubMed1976

Friend spleen focus-forming virus production in vitro by a nonerythroid cell line.

We developed a cell line (IS) that continuously produced both Friend spleen focus-forming virus (SFFV) and XC plaque-forming virus in vitro. The line was derived from the enlarged spleen of an inbred SIM mouse previously infected with a polycythemic strain of Friend leukemia virus in vivo. Friend SFFV titers of 10(4) spleen focus-forming units/ml of culture medium were maintained for more than 200 cell generations. The virus from IS cells induced the splenomegaly and high hematocrit typical of Friend virus-induced erythroid disease in SIM mice. Cells of the IS line were adherent and phagocytic and had a low saturation density. They produced no tumors after being injected sc into normal syngeneic hosts and no spleen colonies after being injected iv into supralethally irradiated hosts. IS cells did not have the character of erythroid cells: They did not contain detectable heme as measured by benzidine-peroxide reagent, did not contain globin mRNA in detectable amounts, and did not produce erythroid colonies in plasma culture in the absence or presence of erythropoietin. According to these criteria, growth in the presence of dimethyl sulfoxide did not stimulate erythroid differentiation of IS cells. We concluded that Friend SFFV can infect nonerythroid spleen cells but that replication of the virus was not obligatorily coupled to expression of genes associated with the erythroid phenotype.

Animals↗
PubMed1975

Viral-related information in oncornavirus-lik particles isolated from cultures of marrow cells from leukemic patients in relapse and remission.

Characterization of ribonucleic acid content of particles released from cultures of marrow cells of leukemic patients indicates the presence of RNA molecules of size and base sequence characteristic of oncornarviruses. Seventeen marrow samples obtained from leukemic patients in relapse or in a chronic phase of the disease yielded particles containing high-molecular-weight RNA with a sedimentation velocity (about 70 S) similar to that obtained for murine oncornavirus RNA. Eight of nine marrow samples from non-leukemic patients did not yield detectable high-molecular weight RNA. Among patients in firm hematological remission, three of three samples from patients with acute lymphoblastic leukemia and three of nine samples from patients with acute myeloblastic leukemia were positive for high-molecular-weight RNA. The base sequence of the RNA in particles was characterized by synthesizing complementary (3-H)DNA in an endogenous reaction and hybridizing to excess RNA from known oncornaviruses. Hybridization of 40-60% of input complementary DNA to simian sarcoma virus RNA was detected. No monology was detected with an avian oncornavirus (Rous sarcoma virus) while an intermediate level of homology (10-30%) was detected in hybridization to murine sarcoma virus (Kirsten) and murine leukemia viruses (Rauscher, Moloney, and Gross).

Avian Sarcoma Viruses↗
PubMed1975

Synthesis of DNA complementary to separated human alpha and beta globin messenger RNAs.

Human globulin messenger RNA, purified by oligo(dT)-cellulose column chromatography, is reproducibly separated into two bands by polyacrylamide gel electrophoresis in the presence of 99% formamide. The more rapidly migrating (fast) band is somewhat more abundant than the slow band in normal (nonthalassemic) total reticulocyte globin messenger RNA. In alpha-thalassemic (Hb H disease) messenger RNA, the slow band is 6.5 times more abundant than the fast band, whereas in beta-thalassemic messenger RNA the fast band is three times more abundant than a second band, which has a slightly greater mobility than the slow band of normal and alpha-thalassemic RNA. The RNA bands of nonthalassemic globin messenger RNA were eluted from the gel and efficiently transcribed into DNA copies by use of the RNA-dependent DAN polymerase of avian myeloblastosis virus. Hybridization of these copy DNAs to fast and slow band RANs and to nonfractionated normal, alpha-thalassemic, and geta-thalassemic messenger RNAs revealed that the eluted fast band RNA contains predominantly alpha-chain specific sequences, whereas the eluted slow band RNA contains predominantly beta-chain specific sequences. Nucleotide sequence analysis of 32-P-labeled RNA transcribed from the slow band copy DNA also indicated that the slow band RNA is beta messenger RNA.

Avian Myeloblastosis Virus↗
PubMed1973

Effects of 5-bromo-2' -deoxyuridine on production of globin messenger RNA in dimethyl sulfoxide-stimulated Friend leukemia cells.

Friend leukemia cells grown in the presence of dimethyl sulfoxide show enhanced erythroid differentiation and hemoglobin synthesis. The effects of dimethyl sulfoxide stimulation are inhibited by BrdU. To determine the effect of BrdU on the amount of globin mRNA present in cells treated and not treated with dimethyl sulfoxide, molecular hybridization between total cell RNA and [(3)H]DNA complementary to mouse globin mRNA was used. Cells treated with BrdU and dimethyl sulfoxide had 70% less globin mRNA than cells treated with dimethyl sulfoxide alone. The size and base sequence of the residual globin mRNA in the cultures treated with BrdU and dimethyl sulfoxide were unaltered. Cells treated with BrdU alone contained slightly more globin mRNA than did the untreated controls, suggesting that BrdU may have a dual effect in transcription of messenger RNA.

Animals↗
PubMed1973

Quantitative deficiency of chain-specific globin messenger ribonucleic acids in the thalassemia syndromes.

A hybridization assay procedure was devised that makes possible quantitation of the ratio of mRNA of alpha to mRNA of beta globin chains in an RNA sample. The assay uses the radioactive synthetic DNA copies obtained by incubation of RNA-dependent DNA polymerase of avian myeloblastosis virus with rabbit globin mRNA that is 80-90% enriched in mRNA specific for synthesis of alpha or beta globin chains. The rabbit alpha-chain mRNA is obtained from the postribosomal supernatant of rabbit reticulocyte lysates; the rabbit beta-chain mRNA is obtained from the largest polysomes of rabbit reticulocytes treated with L-O-methylthreonine. Sufficient homology exists between rabbit and human globin chains and globin mRNAs that the synthetic DNA copies of chain-specific rabbit globin mRNA hybridize with human globin mRNA. Applied to the study of globin mRNA isolated from reticulocytes of humans with alpha and beta thalassemia, the technique revealed marked quantitative deficiency of alpha-chain mRNA relative to beta-chain mRNA in alpha thalassemia and similar deficiency of beta-chain mRNA relative to alpha-chain mRNA in beta thalassemia. The thalassemia syndromes are therefore characterized by true quantitative deficiency of the mRNA specific for the affected globin chain.

Animals↗
PubMed1971

Beta thalassemia and translation of globin messenger RNA.

To define the quality and relative quantity of beta and alpha messenger RNA in human nonthalassemic and thalassemic reticulocytes, intact cells were incubated with [(35)S]methionine. The relative amounts of beta- and alpha-nascent chains on polysomes of different sizes were measured by tryptic digestion of pooled polysomes and by determination of the specific activities of beta and alpha peptides that contain methionine. Betachain synthesis predominated on heavy polysomes in nonthalassemic, as well as in thalassemic cells. Since beta chains in thalassemia are made on normal-size polyribosomes, we conclude that the defect in thalassemia does not involve reduction in the rate of initiation of translation due to the production of an abnormal beta message. Such would lead to beta-chain synthesis on very small polysomes. We therefore suggest that the decreased production of beta-globin chains results from a decreased amount of functional beta-globin messenger RNA.

Anemia, Hemolytic↗