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Biomedical subjects

M Levine

Publications and source records attributed to M Levine.

At least 343 records · Page 19Linked to original sources
PubMed1990

[Visceral candidiasis in 2 children treated for acute leukemia].

The authors report two cases of splenic and hepatosplenic candidiasis occurring during a protracted neutropenia induced by chemotherapy for acute leukemia (lymphoblastic and myeloblastic respectively). Fungal infection was revealed by persistent or recurrent fever after correction of neutropenia. Diagnosis was suggested by findings at abdominal ultrasonography. It was confirmed by histological analysis which, in the first case, required open liver biopsy. In both cases, lack of improvement of splenic lesions despite treatment with Amphotericin B followed by fluconazole led to splenectomy. Both patients received postoperative anti fungal therapy with Ampho B and at one year's follow up, the patient who had the hepatosplenic candidiasis seems to have recovered.

Adolescent↗
PubMed1989

The graded distribution of the dorsal morphogen is initiated by selective nuclear transport in Drosophila.

The maternal morphogen dorsal (dl) plays a key role in the establishment of dorsal-ventral polarity in Drosophila. We present evidence that the graded distribution of dl protein is initiated by selective nuclear transport. The dl protein is uniformly distributed throughout the cytoplasm of early embryos, but approximately 90 min after fertilization, dl protein present in ventral but not dorsal regions is selectively transported to the nucleus. Mutations in maternally active genes that regulate dl disrupt this transport process, resulting in an inactive, cytoplasmically localized form of the dl protein. Selective nuclear transport of dl protein was reproduced in tissue culture cells. The wild-type dl protein is largely restricted to the cytoplasm, while truncated proteins are predominantly localized within the nucleus. Transient cotransfection assays suggest that dl activates expression from several promoters in an apparently sequence-independent manner. We discuss the role of nuclear transport as a regulated process in gene expression and development.

Amino Acid Sequence↗
PubMed1989

Conserved repetitive epitope recognized by CD4+ clones from a malaria-immunized volunteer.

T cell clones obtained from a human volunteer immunized with Plasmodium falciparum sporozoites specifically recognized the native circumsporozoite (CS) antigen expressed on P. falciparum sporozoites, as well as bacteria- and yeast-derived recombinant falciparum CS proteins. The response of these CD4+ CD8- cells was species-specific, since the clones did not proliferate or secrete gamma interferon when challenged with sporozoites or recombinant CS proteins of other human, simian, or rodent malarias. The epitope recognized by the sporozoite-specific human T cell clones mapped to the 5' repeat region of the CS protein and was contained in the NANPNVDPNANP sequence.

Amino Acid Sequence↗
PubMed1989

Ascorbic acid transport and accumulation in human neutrophils.

The transport, accumulation, and distribution of ascorbic acid were investigated in isolated human neutrophils utilizing a new ascorbic acid assay, which combined the techniques of high performance liquid chromatography and coulometric electrochemical detection. Freshly isolated human neutrophils contained 1.0-1.4 mM ascorbic acid, which was localized greater than or equal to 94% to the cytosol, was not protein bound, and was present only as ascorbic acid and not as dehydroascorbic acid. Upon addition of ascorbic acid to the extracellular medium in physiologic amounts, ascorbic acid was accumulated in neutrophils in millimolar concentrations. Accumulation was mediated by a high affinity and a low affinity transporter; both transporters were responsible for maintenance of concentration gradients as large as 50-fold. The high affinity transporter had an apparent Km of 2-5 microns by Lineweaver-Burk and Eadie-Hofstee analyses, and the low affinity transporter had an apparent Km of 6-7 mM by similar analyses. Each transporter was saturable and temperature dependent. In normal human blood the high affinity transporter should be saturated, whereas the low affinity transporter should be in its linear phase of uptake.

Ascorbic Acid↗
PubMed1989

Characterization of a homologue of bithorax-complex genes in the leech Hirudo medicinalis.

We report the isolation and characterization of the Hirudo medicinalis homoeobox gene Lox2. Sequence analysis shows that it contains a region that has homology to Drosophila and vertebrate homoeodomains of the Antennapedia class. In addition, Lox2 shares homology with sequences in the bithorax complex Ultra-bithorax (Ubx) and abdominal A (abdA) genes in a region adjacent to the C-terminus of the homoeodomain. Whole mount in situ hybridization of embryos of various ages demonstrates that during early development this gene has temporally and spatially restricted patterns of expression that resemble those of the homoeotic genes of the Drosophila bithorax complex and of many vertebrate homoeobox genes. The largest accumulation of transcripts was seen in the posterior two-thirds of the developing leech central nervous system in 7-14-day-old embryos. Adult leeches also express Lox2. We propose that in Hirudo, Lox2 represents the ancestral gene of the Ubx and abdA genes of the bithorax complex of Drosophila.

Amino Acid Sequence↗
PubMed1989

Sequence-specific DNA-binding activities of the gap proteins encoded by hunchback and Krüppel in Drosophila.

The segmentation of the Drosophila body plan depends on a hierarchy of interactions among approximately 20-25 regulatory genes that are active in the early embryo (refs 1-4; for a review see ref. 5). The gap genes have a key role in this process and are responsible for the periodic expression of certain pair-rule genes and the localized expression of several homoeotic genes. The two best characterized gap genes, hunchback (hb) and Krüppel (Kr), contain homologies with the zinc-finger DNA-binding motif, although their mode of action in the early embryo is unknown. Here we report that both of the proteins encoded by these genes possess sequence-specific DNA-binding activities, which indicates that they might regulate gene expression at the level of transcription. The binding sites of the hb gene product are related by a 10-base pair (bp) consensus sequence, G/A C/C ATAAAAAA, whereas the binding sites of the Kr gene product share a distinct 10-bp motif, AACGGGTTAA. It is possible that the hb and Kr proteins cooperatively regulate gene expression, because they are expressed in broad, overlapping gradients in the early embryo. We also provide evidence that the on/off periodicity of the pair-rule gene even-skipped (eve) involves the interaction of the hb and Kr proteins with defined eve promoter elements.

Animals↗
PubMed1989

Ascorbic acid within chromaffin granules. In situ kinetics of norepinephrine biosynthesis.

Ascorbic acid requirements for norepinephrine biosynthesis were investigated in intact bovine chromaffin granules using the physiologic substrate dopamine and a novel coulometric electrochemical detection high pressure liquid chromatography system for ascorbic acid. 10 mM external dopamine, 1 mM Mg-ATP, and 1 mM ascorbic acid produced maximal norepinephrine biosynthesis without granule lysis. When external ascorbic acid was omitted, intragranular ascorbic acid was consumed in a 1:1 ratio with respect to norepinephrine biosynthesis. The initial concentration of intragranular ascorbic acid was 10.5 mM, which was depleted in stepwise fashion to 15 lower concentrations over the range of 9.2-0.2 mM. Chromaffin granules containing these varying concentrations of intragranular ascorbic acid were then incubated with 1 mM exogenous ascorbic acid, and norepinephrine biosynthesis from dopamine was determined. The apparent Km of norepinephrine biosynthesis for intragranular ascorbic acid was 0.57 mM by Eadie-Hofstee analysis and 0.68 mM by Lineweaver-Burk analysis. These data indicate that intragranular ascorbic acid is available and required for norepinephrine biosynthesis, that ascorbic acid is a true co-substrate for dopamine beta-monooxygenase, and that intragranular ascorbic acid is maintained by extragranular ascorbic acid. Continued norepinephrine biosynthesis in granules is dependent on both intragranular and extragranular concentrations of the vitamin. Furthermore, in situ kinetics of dopamine beta-monooxygenase for ascorbic acid may be most accurately determined using intact granules and the true physiologic substrate.

Adrenal Medulla↗
PubMed1989

Attenuation of low dose phenobarbital induction of hepatic microsomal aminopyrine N-demethylase activity in rats by cimetidine.

Cimetidine, a substituted imidazole, is an inhibitor of hepatic cytochrome P-450-mediated drug metabolism in rats and humans. We investigated the effect of cimetidine on phenobarbital induction of hepatic microsomal aminopyrine N-demethylase activity in the rat. Phenobarbital induction of aminopyrine N-demethylase was log-linear in the range of 1-6 mg/kg/day and the ED50 was approximately 3 mg/kg/day. Cimetidine 75 mg/kg (four times a day) attenuated the induction of aminopyrine N-demethylase activity by 58% in low dose (3 mg/kg/day) but not in high dose (40 mg/kg/day) phenobarbital treated rats. This result could not be explained by residual inhibition of enzyme activity by cimetidine and suggests that cimetidine affects the induction of hepatic cytochrome P-450 by low dose phenobarbital.

Aminopyrine N-Demethylase↗
PubMed1989

Expression of human HPRT mRNA in brains of mice infected with a recombinant herpes simplex virus-1 vector.

Complete deficiency of the purine salvage enzyme hypoxanthine-guanine phosphoribosyltransferase (HPRT) results in a devastating neurological disease, the Lesch-Nyhan syndrome. This disorder has been identified as a candidate for initial attempts at somatic cell gene therapy. We have previously reported the construction of a recombinant herpes simplex virus type 1 (HSV-1) vector containing human hprt cDNA sequences under the regulatory control of the viral thymidine kinase gene (tk) [Palella et al., Mol. Cell. Biol. 8 (1988) 457-460]. Infection of HPRT- cultured rat neuronal cells with these vectors resulted in transient expression of human hprt. In this paper, we report the expression of human hprt mRNA transcripts in the brains of mice infected in vivo with this vector by direct intracranial inoculation. Human hprt transcripts were distinguished from endogenous mouse transcripts by RNase A mapping using riboprobes transcribed from human hprt cDNA. These initial studies demonstrate the transfer and transcription of a human gene in brain cells by direct in vivo infection with recombinant HSV-1 vectors.

Animals↗
PubMed1989

Barium ions enter chromaffin cells via voltage-dependent calcium channels and induce secretion by a mechanism independent of calcium.

Barium ions enter chromaffin cells via voltage-sensitive calcium channels, although the intracellular site of barium action is distinct from that of calcium. The entry of barium primarily through voltage-sensitive channels was indicated by experiments showing inhibition of 133Ba2+ uptake by blockers of voltage-dependent calcium channels. In addition, 133Ba2+ uptake was stimulated by 50 mM KCl but not by nicotine. Furthermore, 133Ba2+ uptake was inhibited by hyperosmolarity, which specifically blocks the voltage-sensitive calcium channel but not the receptor-associated calcium channel. These conclusions from studies on barium uptake were also borne out by experiments measuring catecholamine secretion. Thus, blockers of voltage-dependent calcium channels which inhibited barium uptake also inhibited barium-induced catecholamine secretion. In other experiments, simultaneous stimulation with nicotine and barium in the presence of calcium evoked coincident and additive catecholamine secretion. By contrast, when 50 mM KCl was substituted for nicotine in the same experimental design, barium ions inhibited potassium-induced catecholamine secretion at low calcium concentrations. Only at high calcium concentrations were barium-induced and potassium-induced secretion additive. These data also indicate that barium and calcium compete at the voltage-sensitive pathway. Furthermore, these additivity data suggest that once inside the cell, barium and calcium have two distinct mechanisms of action. As predicted by this hypothesis, in digitonin-permeabilized chromaffin cells either calcium or barium stimulated catecholamine release, and in the presence of both cations catecholamine secretion was equivalent to the sum of secretion with either cation alone. Additional support of this concept comes from experiments showing that while calcium-mediated catecholamine secretion is sensitive to trifluoperazine and imipramine, barium-mediated secretion is not. Taken together, all these data indicate that there are two distinct intracellular sites of action for barium and calcium. In contrast to catecholamine secretion, non-exocytotic ascorbic acid secretion was induced by nicotine and potassium in the presence of calcium, but not by barium alone. These data provide additional evidence that barium acts by a different mechanism than calcium, in still another secretory system in chromaffin cells.

Animals↗
PubMed1989

Fast ELISA for measuring serum antibody responses.

A method which speeds up the enzyme-linked immunosorbent assay (ELISA) is described. The procedure uses a modified Falcon fast assay screening system (Becton Dickinson Labware, Lincoln Park, NJ) and Falcon round-bottom 96-well plates. Antigen is adsorbed onto beads which extend from a lid and fit into 96-well plates. The beads are washed in a trough and reacted to antibody in the round-bottom plate. The labor required to wash the plates after coating with antigen, antibody or conjugate is thereby reduced. Greater flexibility and accuracy result, especially with the use of more than one 96-well plate. In this study, naturally occurring human IgG antibody responses to two isolated bacterial antigens were measured in over 200 subjects. It was found that numerical taxonomy could be used to split out the high IgG responders. The IgM response to one of the antigens was less variable and not significantly related to the IgG response. The fast ELISA is as useful to operate as the standard ELISA, but less stressful on the operator and more rapid.

Actinomyces↗
PubMed1989

Autoregulatory and gap gene response elements of the even-skipped promoter of Drosophila.

The pair-rule gene even-skipped (eve) plays a key role in the regulatory hierarchy governing segmentation in Drosophila. Here we describe the use of P-transformation and eve promoter fusions to identify cis elements that regulate the periodic seven-stripe eve pattern. A distal region of the eve promoter, located between -5.9 and -5.2 kb, controls autoregulation. Sequences from this region will induce striped expression of a heterologous hsp70 basal promoter in the presence, but not absence, of endogenous eve+ products. Autoregulatory activity was localized to a 200-bp region of the distal eve promoter. We also provide evidence that individual eve expression stripes are regulated by separate cis sequences. eve promoter sequences located between -4.7 and -3 kb upstream of the transcription start site are important for the initiation of stripe 3, whereas sequences between -1.7 and -0.4 kb are needed for stripes 2 and 7. It is possible that these latter regions are directly regulated by the products of gap genes.

Animals↗
PubMed1989

Ascorbic acid analysis using high-performance liquid chromatography with coulometric electrochemical detection.

A method for the detection of ascorbic acid using high-performance liquid chromatography with coulometric electrochemical detection and a technique for stabilization of the vitamin are described. Since less than 1 pmol of ascorbic acid can be detected, this assay provides significantly greater sensitivity than nearly all of the currently available procedures. Stabilization of 10 pmol or less of ascorbic acid at room temperature for up to 4 h and for several weeks at -70 degrees C facilitates storage of a large number of samples and measurement of ascorbic acid using an automated sampling device. This method was used to quantitate the amounts of ascorbic acid in human polymorphonuclear leukocytes and bovine adrenomedullary chromaffin granules. The calculated concentrations found for human neutrophils (1.35 mM) and bovine chromaffin granules (10.0 mM) are in agreement with previously published data. The assay is suitable for the determination of ascorbic acid in biological samples where only a small amount of tissue is available or very low amounts of ascorbic acid are found. This method is the first application of coulometric electrochemical detection to ascorbic acid HPLC analysis.

Animals↗