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F Rosen

Publications and source records attributed to F Rosen.

At least 19 recordsLinked to original sources
PubMed1991

Luminescent immobilized enzyme test systems for inorganic pyrophosphate: assays using firefly luciferase and nicotinamide-mononucleotide adenylyl transferase or adenosine-5'-triphosphate sulfurylase.

Inorganic pyrophosphate was measured by luminescence produced by a pyrophosphatase (NAD adenylyl-transferase or ATP sulfurylase) coimmobilized with firefly luciferase on Sepharose beads, with continuous flow of saturating concentrations of substrates (NAD plus luciferin or adenylophosphosulfate plus luciferin, respectively) and intermittent injections of samples containing pyrophosphate. In this scheme, the limiting substrate (pyrophosphate) is regenerated, a situation that is well suited to a bioluminescent assay. The instrumentation allowed for automation with a through-put of approximately one sample every 4 min. With standard solutions or samples that do not contain ATP, the sensitivity of the assay permits detection of less than 1 pmol pyrophosphate in a volume of 20 microliters (50 nmol/liter) with a coefficient of variation approximately equal to 4%. To assay biological samples, it was shown that endogenous ATP can be inactivated by oxidation with sodium periodate. Periodate treatment and quenching engenders dilution that limits the sensitivity to approximately 600 nmol/liter pyrophosphate in the starting material. The assay has been applied to the determination of intracellular pyrophosphate in human lymphocytes and to the measurement of nucleoside-triphosphate pyrophosphohydrolase in human fibroblasts. The variability of the assay was greater with biological samples than with standard samples, with a coefficient of variation of 15.3% in a series of determinations of intracellular pyrophosphate in a series of replicate lymphocyte lysates. Bioluminescent systems of coupled coimmobilized enzymes offer great promise for sensitive, safe, automated assaying of metabolites.

Adenosine Triphosphate↗
PubMed1990

Signal transduction via leukocyte antigen CD43 (sialophorin). Feedback regulation by protein kinase C.

CD43 is a constitutively phosphorylated 115-kDa sialoglycoprotein expressed on a variety of blood cells including lymphocytes and monocytes. L10, a mAb directed against CD43, triggers T cell activation and enhances hydrogen peroxide production in monocytes. Activation of mononuclear cells by L10 initiates phosphoinositides hydrolysis, C2+ mobilization, and protein kinase C (PKC) activation. In turn, activated PKC hyperphosphorylates CD43, suggesting a potential role for PKC in the regulation of signaling via CD43. To address this issue, we have analyzed the effect of PKC activation by the tumor promoter PMA on L10-triggered rise in intracellular free Ca2+ concentrations ([Ca2+]i). Treatment of mononuclear cells with PMA profoundly inhibited the increase in [Ca2+]i induced by L10. The inhibition of CD43-mediated signaling by PMA was due, in part, to uncoupling of CD43 from the signal-transducing G protein. This was evidenced by the comparatively modest inhibition by PMA of the increase in [Ca2+]i induced by the direct G protein activator AlF4-. PMA treatment did not affect the surface expression of CD43. However, it induced the hyperphosphorylation of CD43, the extent of which correlated with the inhibition of CD43-mediated increase in [Ca2+]i. Staurosporine, a potent inhibitor of PKC, abrogated the hyperphosphorylation of CD43 and normalized CD43-mediated signaling in PMA-treated cells. Significantly, in the absence of PMA, staurosporine enhanced the rise in [Ca2+]i triggered by L10, suggesting that engagement of CD43 by activating ligands results in feedback inhibition by PKC. It is concluded that activation of PKC inhibits signaling via CD43 by mechanisms involving phosphorylation and uncoupling of CD43 from the signal-transducing apparatus and by distal, post-receptor events.

Alkaloids↗
PubMed1990

Identification of a closely linked DNA marker, DXS178, to further refine the X-linked agammaglobulinemia locus.

X-linked agammaglobulinemia (XLA) is an inherited recessive disorder in which the primary defect is not known and the gene product has yet to be identified. Utilizing genetic linkage analysis, we previously localized the XLA gene to the map region of Xq21.3-Xq22 with DNA markers DXS3 and DXS17. In this study, further mapping was performed with two additional DNA probes, DXS94 and DXS178, by means of multipoint analysis of 20 families in which XLA is segregating. Thirteen of these families had been previously analyzed with DXS3 and DXS17. Three crossovers were detected with DXS94 and no recombinations were found between DXS178 and the XLA locus in 9 informative families. Our results show that XLA is closely linked to DXS178 with a two-point lod score of 4.82 and a multipoint lod score of 10.24. Thus, the most likely gene order is DXS3-(XLA,DXS178)-DXS94-DXS17, with the confidence interval for location of XLA lying entirely between DXS3 and DXS94. In 2 of these families, we identified recombinants with DXS17, a locus with which recombination had not previously been detected by others in as many as 40 meiotic events. Furthermore, DXS178 is informative in both of these families and does not show recombination with the disease locus. Therefore, our results indicate that DXS178 is linked tightly to the XLA gene.

Agammaglobulinemia↗
PubMed1989

Mechanism of mononuclear cell activation by an anti-CD43 (sialophorin) agonistic antibody.

CD43 (sialophorin, gpL115) is a sialoglycoprotein expressed on a wide variety of blood cells including lymphocytes, monocytes, neutrophils, and platelets. L10, an anti-CD43 mAb, has been shown to induce monocyte-dependent activation and proliferation of human T lymphocytes. We have studied the signaling mechanism involved in this activation process. Treatment of PBMC and purified populations of T cells and monocytes with L10 induced the hydrolysis of phosphoinositides with the resultant generation of the phosphoinositide-derived second messengers diacylglycerol and inositol phosphates. This was associated with the translocation of protein kinase C from cytosol to membrane fractions and an increase in free intracellular Ca2+ in treated cells. In human leukemic T cell lines, the magnitude of signaling via CD43 did not correlate with the density of the TCR/CD3 surface expression nor with the intensity of signaling via the TCR/CD3. Moreover, a mutant derived from the leukemic T cell line HPB-ALL that was severely defective in TCR/CD3 surface expression and signaling nevertheless had normal CD43 surface expression and signaling compared with the parent cell line. It is concluded that CD43 is functionally coupled to the phospholipase C/phosphoinositides signaling pathway. In human T cells, signaling via CD43 proceeds independently of TCR/CD3. The widespread expression of CD43 suggests a potentially important role for this molecule in orchestrating the activation of multiple cell types.

Adult↗
PubMed1987

Use of nude mouse xenografts as preclinical drug screens: in vivo activity of established chemotherapeutic agents against melanoma and ovarian carcinoma xenografts.

To evaluate the utility of nude mouse xenografts as preclinical drug screens, the activity of ten established chemotherapeutic agents was evaluated against seven melanoma and three ovarian carcinoma xenografts. Xenografts were established using primary explants from patients who had not received chemotherapy and serially passaged as sc tumors in nude mice. In vivo drug activities for dactinomycin, carmustine, vinblastine, melphalan, amsacrine, cisplatin, bleomycin, mitomycin, doxorubicin, and etoposide were evaluated by 4 weekly ip injections of 10% less than LD10 doses. Plots of relative tumor growth versus time were nearly log-linear. Analysis of in vivo activity was performed using percent control growth (treated/control tumor volume) and by calculation of a novel growth delay index obtained by fitting growth curves to a quadratic regression model. Both modes of data analysis identified alkylating agents (melphalan, carmustine, and mitomycin) as the most active drugs against human melanomas. Melphalan, mitomycin, and cisplatin showed the greatest activity against ovarian xenografts. However, complete tumor regressions were noted only with melphalan, mitomycin, and cisplatin against a single ovarian tumor xenograft. Correlation analysis suggested xenograft tumor growth rate was an important determinant of drug response. These results suggest that preclinical, new-drug screening with melanoma xenografts would identify drugs such as alkylating agents as active, and may not provide an advantage over murine leukemia screens. However, screening with ovarian xenografts may more closely reflect clinical drug activity. Criteria for detecting active drugs in such systems are discussed.

Animals↗
PubMed1986

Use of nude mouse xenografts as preclinical drug screens. Further studies on in vitro growth of xenograft tumor colony-forming cells.

Previous studies suggested tumor colony-forming cells (CFC) grown from xenografts might be useful as a preclinical, in vitro drug screen. To further evaluate this possibility, eight melanoma and six ovarian carcinoma xenografts were established from untreated patients and tested for in vitro CFC growth. For each tumor, linear relationships between cells plated and colony (30 cells or greater than 75 micron diameter) and cluster (10-30 cells or 50-75 micron) growth were observed. All eight melanomas grew sufficient colonies (greater than or equal to 30) for in vitro drug assessment, although four required hypoxic (pO2 = 40) incubation to reliably attain this level of growth. Only one in six of the ovary xenografts consistently grew enough colonies, and growth was not significantly improved by hypoxic incubation, or addition of luteinizing hormone, follicle-stimulating hormone, or steroid hormones. Cloning efficiencies (colonies + clusters/cells plated) for tumors demonstrating adequate growth ranged from 0.01% to 0.3%. For most tumors, no direct relationship was observed between characteristics of xenograft tumors (size) or their resulting cell suspensions (viabilities, cell yield) and CFC growth. Cell suspensions were incubated with a 3 log concentration of nine established chemotherapeutic agents. Resulting dose-effect curves were linear and showed no plateaus of drug effect. Analyzing 447 in vitro drug trials on six melanomas and one ovarian carcinoma, interexperiment variability was high. Cell lines were established from three xenografts using a low concentration of fetal bovine serum (1%), and also examined for in vitro drug sensitivity. Using both liquid culture isotope incorporation and a colony-forming assay, drug sensitivity profiles for the cell lines were nearly identical to those for parent xenograft CFC. However, assays performed using the cell lines were more reproducible than those using xenograft tissue. The authors conclude that tumor CFC can be reliably grown from melanoma xenografts, but in vitro drug assays using these xenografts are poorly reproducible. The xenografts are a resource for establishing cell lines, and drug assays performed using these lines are highly reproducible. Similarities in drug sensitivity profiles for parent xenograft CFC and derived cell lines suggest that, despite poor reproducibility, repetitive assays using melanoma CFC accurately reflect some properties of cells which sustain tumor cell growth.

Animals↗
PubMed1986

Evidence for a defect in "switch" T cells in patients with immunodeficiency and hyperimmunoglobulinemia M.

Immunodeficiency with hyperimmunoglobulinemia M is a syndrome characterized by normal to elevated serum levels of IgM and low levels or absence of IgG and IgA. The defect in this syndrome is thought to reside within the B lymphocyte, which may be unable to undergo a "switch" in immunoglobulin class from IgM to IgG or IgA. To address this question more directly, we cultured B cells from nine patients with this syndrome with pokeweed mitogen and either "switch" T cells or normal control T cells. In cultures with normal T cells, only IgM was secreted, whereas in cultures with switch T cells, IgG as well as IgM, or IgM, IgG, and IgA were secreted. Furthermore, analysis of the immunoglobulin heavy-chain genes in these B cells by means of genetic probes of constant and switch regions revealed normal gene patterns. These data suggest that B cells from patients with hyperimmunoglobulinemia M may not be abnormal, as previously proposed, and that, at least in some patients with this syndrome, a defect in switch T cells may be pathogenic.

Adolescent↗
PubMed1979

Mammary cancer: selective action of the estrogen receptor complex.

Progesterone receptors in the autonomous rat mammary tumor MTW-9B are reduced 80 to 90 percent after ovariectomy, but are not reduced if ovariectomized animals are given estrogen. Tumor growth, however, is independent of estrogen status and insensitive to pharmacological doses of estradiol. This represents an unusual system characterized by a selective action of an inducing agent on the genome.

Animals↗
PubMed1979

Immunoglobulin subclasses in normal children.

The range, mean, and normal bounds of serum IgG 1--4 subclass concentrations in children 0--16 years old are given. Levels increase with age, plateauing at different ages. Definition of the normal range will permit better identification of individuals with either low or high levels.

Adolescent↗
PubMed1979

Sex differences in the concentrations of glucocorticoid receptors in rat liver and thymus.

The effects in rats of adrenalectomy, hypophysectomy, ovariectomy or combinations of these operations on the concentrations of glucocorticoid receptors in the cytosol of liver and thymus were measured. The concentrations of glucocorticoid receptors were lower in cytosols from liver and thymus of female than of male rats. After adrenalectomy, there was a significant increase in the concentrations of receptors measured in the cytoplasm from the liver and thymus of female rats and from the liver of male rats. After adrenalectomy or hypophysectomy, there was no sex difference in the concentration of glucocorticoid receptors in cytosols of liver or thymus. After ovariectomy, the concentration of receptors in cytosols from the thymus, but not from the liver, increased. Ovariectomized rats responded to adrenalectomy in the same way as intact male rats. The different responses shown by male and female rats to endocrine manipulation probably depend upon associated changes in plasma corticosterone concentrations which are influenced by the ovary. Differences in response between the liver and thymus probably reflect a preferential distribution of corticosterone to the liver rather than to the thymus.

Adrenalectomy↗
PubMed1979

Perceptual reactance and task performance.

57 female subjects participated in a study in which perceptual reactance was assessed by the kinesthetic figural aftereffects test. Subjects identified as augmenters were quicker than reducers on a disjunctive reaction time task and a significant interactive effect was found on a paired-associate task. Augmenters performed better on a similar list while reducers were more effective on a dissimilar list.

Female↗
PubMed1979

Successful bone marrow transplantation in sensitized recipients.

Fourteen patients with aplastic anemia and one with the Wiskott-Aldrich syndrome who were specifically sensitized against their donors were successfully engrafted with bone marrow from those donors. Sensitivity was detected in antibody-independent and antibody-dependent cell-mediated lysis assays. In order to erase this immunity to non-MHR familial transplantation antigens, multiagent immunosuppression with cyclophosphamide, procarbazine, and whole rabbit antithymocyte serum (ATS) was used. The data suggest that ATS was largely responsible for abrogation of this sensitivity and indicate that immunity does not represent a barrier to successful transplantation.

Anemia, Aplastic↗
PubMed1978

Comparison of the antitumor activity and toxicity of 2,4-diamino-5-(1-adamantyl)-6-methylpyrimidine and 2,4-diamino-5-(1-adamantyl)-6-ethylpyrimidine.

The effects of two new compounds, 2,4-diamino-5-(1-adamantyl)-6-methylpyrimidine (DAMP) and 2,4-diamino-5-(1-adamantyl)-6-ethylpyrimidine (DAEP), on the growth of Walker carcinoma 256 were studied. In Sprague-Dawley rats, both compounds inhibited the growth of the Walker tumor, which is naturally resistant to methotrexate. Murphy-Sturm lymphosarcoma, which is extremely sensitive to methotrexate, was not inhibited by DAMP. In contrast, 2,4-diamino-5-(3',4'-dichlorophenyl)-6-methylpyrimidine (DDMP) inhibited the growth of the Murphy-Sturm and Walker tumors. The toxicity of DAMP and DAEP was charaxterized by convulsions followed by death; DDMP did not show such toxicity at median lethal doses. DAEP and DAMP reversibly reduced the number of polymorphonuclear leukocytes in the peripheral blood of rats, and DAMP also decreased the lymphocytes and platelets; DDMP markedly reduced all these cellular elements. Consistently, bone marrow was also markedly depressed by DDMP.

Adamantane↗