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Biomedical subjects

M E Balis

Publications and source records attributed to M E Balis.

At least 55 records · Page 3Linked to original sources
PubMed1976

A normal level of adenosine deaminase activity in the red cell lysates of carriers and patients with severe combined immunodeficiency disease.

The red cell lysates of two children with severe combined immunodeficiency disease (SCID) exhibited a virtually total absence of adenosine deaminase (adenosine aminohydrolase, EC 3.5.4.4) when standard volumes were assayed. Under these conditions the parents exhibited depressed specific activity except for one mother, whose lysate showed a normal value for activity. Upon storage of the lysate at 4 degrees, a significant amount of activity appeared in one of the SCID children, and the activity of the heterozygous carriers was stimulated. With the use of a sensitive spectrophotometric assay based on conversion of inosine to uric acid, it was shown that the specific enzymatic activity in each of the SCID patients increased progressively as the volume of lysate assayed was lowered. With the smallest amount of lysate this specific activity was in the normal range. Similarly, the specific activity of each of the parents' lysates increased to the level of normal (or, in one case, about twice normal) as smaller volumes were assayed. The activity in the SCID patient was inhibitable by 2-fluoroadenosine and N6-methyladenosine, known competitive inhibitors of human red cell adenosine deaminase. The lysate from the SCID patient was also shown to inhibit adenosine deaminase partially purified from a normal individual. The results are interpreted in terms of a genetically programmed production of an adenosine deaminase inhibitor in at least one variant of the severe combined immunodeficiency disease.

Adenosine Deaminase↗
PubMed1976

Studies on cell communication with enucleated human fibroblasts.

Metabolic cooperation, the correction of the mutant phenotype in cells deficient in hypoxanthine phosphoribosyltransferase (HPRT-) by intimate contact with normal cells (HPRT+), represents a form of cell communication that is easily studied with radioautography. In the present study it was found that the formation of cell junctions needed for communication does not require protein synthesis nor is it under the immediate control of the cell nucleus. Enucleated normal cells efficiently communicate with HPRT- mutant cells. The effectiveness of enucleated cells as donors in metabolic cooperation provides evidence that it is the transfer of small molecules, nucleotide, or nucleotide derivatives that is responsible for correction of the mutant phenotype. Karyoplasts (nuclei with small amounts of cytoplasm surrounded by a plasma membrane) are unable to efficiently communicate with intact cells. The utilization of [3H]hypoxanthine by communicating mixtures of HPRT+ and HPRT- human cells is not significantly different than in the normal cells alone. Metabolic cooperation, as studied involves a redistribution of purine-containing compounds among communicating cells.

Adenine↗
PubMed1976

Hypoxanthine phosphoribosyltransferase activity in intact fibroblasts from patients with X-linked hyperuricemia.

Discordance between clinical phenotype and the level of a mutant enzyme activity may reflect differences between enzyme function in vivo and that measured by the customary enzyme assays on cell extracts. In the present study, the conversion of hypoxanthine to phosphorylated products was measured in intact skin fibroblasts and in cell extracts from seven patients with mutant hypoxanthine-guanine phosphoribosyltransferase (HPRT) and six control subjects. The patient's phenotypes ranged from asymptomatic hyperuricemia to the Lesch-Nyhan syndrome. Although there was a general correlation between the HPRT activity in cell extracts assayed by the usual methods and the function of the purine salvage pathway in patients, as reflected by urinary oxypurine excretion, there were notable exceptions. A more accurate appraisal of the functioning of the pathway at the cellular level is achieved by measuring the conversion of substrate to product in the intact cell at physiological concentrations of substrates, activators, and product and metabolite inhibitors, and in a physiological ionic environment. In one of the seven patients, the standard enzyme assay indicated normal function, whereas measurements in the intact cell exposed severe dysfunction of the salvage system. In another, the standard assay suggested a severe deficiency not evident in the intact cell or in the patient.

Adolescent↗
PubMed1976

Biochemical changes in preneoplastic rodent intestines.

Two enzymes were examined as potential indicators of early precancerous changes. Ornithine decarboxylase, an enzyme normally associated with rapid cell division, is low in the rapidly dividing, cancer-susceptible colon. The level of this enzyme was also very high in the nondividing cells of the small intestines. Administration of an intestinal carcinogen, dimethylhydrazine, led to a large increase in colonic ornithine decarboxylase but did not affect the enzyme in liver. A liver carcinogen, acetylaminofluorene, induced manyfold increases in ornithine decarboxylase of the liver but not of the colon. Studies of thymidine kinase of the gut showed that this enzyme changed quantitatively and qualitatively throughout the life of the animal, from fetal rat to newborn and adult. The tumor enzyme has many fetal-like properties. Long-term treatment with dimethylhydrazine led to changes in thymidine kinase reminiscent of the fetal enzyme. Short-term treatment caused sharp increases in the thymidine kinase of nondividing cells of the jejunum and the proximal end of the colon; similar changes in the distal end of the colon were slower in appearing and less pronounced.

2-Acetylaminofluorene↗
PubMed1976

Changes in ornithine decarboxylase activity in rat intestines during aging.

The ornithine decarboxylase (ODC) activity of rat intestines, liver, and brain was found to vary dramatically as animals develop and age. Unusually high activity was present in the small intestines of adult animals. The ODC activity of the small intestines approximated that of fetal tissues and of regenerating rat liver. Putrescine, spermine, and spermidine levels of fetal and adult animal tissues were determined. In all tissues but the stomach mucosa, high putrescine levels correlated with high IDC activity. However, the total polyamine concentrations of the stomach and colon could not be correlated with ODC levels, and no simple relationship between polyamine levels of ODC levels and cellular proliferation in the gut was found.

Aging↗
PubMed1976

Biochemical changes in premalignant intestines.

The immunological properties of thymidine kinase from a variety of human tumors suggest that the form of the tumor enzyme resembles that found in the placenta and in the nondividing colonic flat mucosa. To examine the placenta-like characteristics of tumor thymidine kinase, the jejunum and colon from rats ranging in age from fetal to old and from animals treated with dimethylhydrazine (DMH), an intestinal carcinogen, have been studied. In normal jejunum, thymidine kinase activity decreased rapidly with age. Both the activity and the response to phospholipase C and to mercaptans in DMH-induced tumors resembled that of fetal gut, while those in abnormal appearing DMH-treated jejunum were intermediate between normal control of the same age and tumor. Similar but less pronounced changes were seen in the colon. In the jejunum, the level of another enzyme normally associated with rapid cell division, ornithine decarboxylase, was found to be over 100 times that of the liver, colon, and stomach. Treatment of the animals with acetylaminofluorene and with DMH resulted in elevated levels of the enzyme in liver and in colon, respectively, but had little effect on this enzyme in other tissues. The data presented indicate that there were premalignant changes in the levels of both of these enzymes in target tissues of animals treated with carcinogens.

Adenocarcinoma↗
PubMed1975

Hysteretic characteristic of adenine phosphoribosyltransferase.

Preassay-incubation of the highly purified human erythrocyte adenine phosphoribosyltransferase (EC 2.4.2.7) (AMP pyrophosphorylase) with one of its substrates, 5-phosphoribosyl 1-pyrophosphate (PRibPP), changes the apparent V max value of the enzyme reaction. The extent of inhibition by preassay-incubation with an inhibitor, fructose 1,6-diphosphate (FDP), or a destabilizer, hypoxanthine (Hx), is found not to be proportional to the amount of the inhibitor present. The maximum inhibition achieved by preassay-incubation was about 40%. The PRibPP, FDP, and Hx induced changes in AMP pyrophosphorylase do not require the presence of divalent ions. The inhibtion of AMP pyrophosphorylase produced by preincubation with Hx was prevented when PRibPP was added to the preassay-incubation system. However, the preassay-incubation effect of FDP was only partially diminished under the same conditions. Contrary to the PRibPP-bound AMP pyrophosphorylase, the adenine-bound enzyme was found to be more heat labile than the unbound enzyme. Similar thermal instability was also observed with FDP- and Hx-bound enzyme. Our experimental results indicate that a conformational change of AMP pyrophosphorylase induced by the binding of metabolites is a slow process as compared to the overall catalytic reaction. This hysteretic characteristic of AMP pyrophosphorylase may be one of the regulatory mechanisms in purine intermediary metabolism.

Adenine Phosphoribosyltransferase↗
PubMed1975

Relationships between non-extractable DNA and the bacterial growth cycle.

Escherichia coli DNA has been fractionated into extractable and non-extractable DNA after deproteinization of detergent-lysed cell preparations with chloroform-isoamyl alcohol. The former was extracted with dilut buffered saline whereas the latter remained in the interphase layer associated with residual cellular debris from which almost 40 percent could be released by incubating with pronase. About 20-25 percent more amino acid residues were bound to the pronase-released DNA than to the extractable DNA, but the relative distribution of the residues in the two DNA samples was virtually identical. The specific activities and the relative amounts of denser (1.709g-cm-3) and lighter (on the surface of CsCl gradients) DNA fractions from E. coli, grown in the presence of labeled thymidine, indicated that these two corresponded to extractable and non-extractable DNA, respectively. The relative amounts of the two fractions varied with the growth phase primarily as a function of the growth rate. Age and metabolic state of cells in the culture or those used as inocula could modify this relative distribution. When growth rate was maximal, the ratio of the two remained at about 1. During lag phase when no appreciable net synthesis of DNA could be detected, there was a rapid and preferential incorporation of labeled thymidine into non-extractable DNA. A disproportionate increase in the fraction of the total DNA which was extractable, was also abserved but only when stationary phase cultures were used as inocula. Complete equilibration of the label in the two DNA fractions was attained only after cultures had reached mid-log phase of growth. Similar results were obtained when prelabeled cells were used. These data have been interpreted as suggesting that the rate of cell growth and DNA synthesis are related to the number or size of sites of attachment of DNA to some cellular structure. Newly synthesized DNA would be attached to different sites on this structure and initiation of DNA replication in lag phase would require reorientation of the two kinds of DNA. Small peptides which are firmly bound to the DNA and which vary quantitatively with the rate of DNA synthesis could perhaps be involved in the attachment to the sites.

Amino Acids↗
PubMed1975

In vivo repair of rat intestinal DNA damage by alkylating agents.

In an effort to evaluate the possible correlation of the transforming ability of the known colon carcinogens dimethylhydrazine, 3-2'-dimethyl-4-aminobiphenyl, and methylazoxymethanolacetate to damage and repair of DNA, a series of compounds known to react with DNA-nitrogen mustard, methylmethanesulfonate, and mitomycin C--were administered to rats that had been prelabeled with 3H-thymidine. The DNA of crypt and villus of the jejunum and crypt and surface cells of the large bowel were analyzed by ultracentrifugation on an alkaline sucrose gradient. All fractions suffered degradation to such an extent that essentially no undamaged DNA was detectable. This was followed by repair and an increase in size. However, in the surface cells of the colon of animals that had received a carcinogenic insult there was far less rapid repair. Since this is the site where tumors would ultimately arise these data are supportive of the hypothesis that there is a relationship between decreased repair and carcinogenicity. In view of the age related incidence of colon cancer, repair in older animals was evaluated and was found to be less than that seen in the young. Since multiple treatment with the carcinogen dimethylhydrazine is required and there is a long latent period, the effect of this treatment on repair potential was evaluated. Similar to what was seen in the older animals, these treated rats had greatly reduced capacity to repair DNA. All these observations are consistent with the hypothesis that decreased repair of DNA alterations is a concomitant of carcinogenesis.

Alkylating Agents↗
PubMed1975

Lack of effect of portacaval shunt on ornithine decarboxylase activity in regenerating rat liver.

The influence of the end to side portacaval shunt on the level of activity of the enzyme ornithine decarboxylase following partial hepatectomy was investigated. A sharp rise in the enzyme level was found in both the shunted animals and in the normal controls. The chronology of the increase in enzymic activity was somewhat altered in the portacaval shunted animals, but both groups had similar peak values. The rate of cell division was also found to be unaffected. This report finds no evidence of support for the reported inhibition by the portacaval shunt of the increase in ornithine decarboxylase following partial hepatectomy.

Animals↗
PubMed1975

Ethionine-induced changes in rat liver transfer RNA methylation.

We have confirmed the finding by Rajalakshmi that transfer RNA (tRNA) from livers of ethionine-treated rats can act as a substrate for homologous tRNA-methylating enzymes in vitro. This methyl-deficient tRNA from liver can be methylated in vitro by enzymes from normal or ethionine-treated rats. The in vitro inhibition of tRNA methylation that follows ethionine treatment can be at least partially relieved in vitro. The liver extracts from ethionine-treated animals contained a low-molecular-weight inhibitor of tRNA methylation. Dialysis of enzyme preparations from ethionine-treated, but not control, rats resulted in large increases in tRNA methylase activity, with either Escherichia coli or homologous tRNA's as substrate. Furthermore, the tRNA methylase activity of control rat liver enzyme extracts was greatly depressed by dialysate from liver homogenates of ethionine-treated rats. After 5 days of ethionine administration the liver tRNA methylase activities were significantly higher than those of control preparations despite the continued presence of the dialyzable inhibitor(s). The liver tRNA's from these animals were poorer methyl acceptors than those from 3-day-treated rats, although still better than tRNA's from untreated rats. These observations have been interpreted to indicate that ethionine causes the accumulation in the liver of inhibitors of tRNA methylation. Early in the course of ethionine administration, methyl-deficient tRNA can be isolated. When the period of ethionine treatment is extended, the organism attempts to maintain homeostasis by production of increased amounts of tRNA-methylating enzymes. The increased quantities of these enzymes are able to overcome, at least partially, the effects of the inhibitors and to decrease the extent to which methyl-deficient tRNA is produced.

Animals↗