PubMed Health⌕ Search

Biomedical subjects

N Lin

Publications and source records attributed to N Lin.

At least 19 recordsLinked to original sources
PubMed1995

Thrombopoietin, the Mp1 ligand, is essential for full megakaryocyte development.

The development of megakaryocytes (MKs) from their marrow precursors is one of the least understood aspects of hematopoiesis. Current models suggest that early-acting MK colony-stimulating factors, such as interleukin (IL) 3 or c-kit ligand, are required for expansion of hematopoietic progenitors into cells capable of responding to late-acting MK potentiators, including IL-6 and IL-11. Recently, the Mp1 ligand, or thrombopoietin (Tpo), has been shown to display both MK colony-stimulating factor and potentiator activities, at potencies far greater than that of other cytokines. In light of these findings, we tested the hypothesis that Tpo is absolutely necessary for MK development. In this report we demonstrate that neutralizing the biological activity of Tpo eliminates MK formation in response to c-kit ligand, IL-6, and IL-11, alone and in combination, but that these reagents only partially reduce MK formation in the presence of combinations of cytokines including IL-3. However, despite the capacity of IL-3 to support the proliferation and initial stages of MK differentiation, elimination of Tpo prevents the full maturation of IL-3-induced MK. These data indicate that two populations of MK progenitors can be identified: one that is responsive to IL-3 but can fully develop only in the presence of Tpo and a second that is dependent on Tpo for both proliferation and differentiation. Thus, our results strongly suggest that Tpo is the primary regulator of MK development and platelet production.

Animals↗
PubMed1995

Stem cell factor modulates avidity of alpha 4 beta 1 and alpha 5 beta 1 integrins expressed on hematopoietic cell lines.

Interactions between hematopoietic cells and bone marrow (BM) stroma, composed of extracellular matrix and stromal cells, are crucial for hematopoiesis. Integrins facilitate these interactions by mediating adherence of hematopoiesis. Integrins facilitate these interactions by mediating adherence of hematopoietic cells to both the extracellular matrix and stromal cells. Marrow stromal cells secrete a variety of growth factors, including stem cell factor (SCF). Because treatment with SCF in vivo mobilizes primitive hematopoietic cells from the BM, we investigated the effect of the growth factor SCF of hematopoietic cell adhesion. These studies show that SCF modulates adhesive function in a dose- and time-dependent manner, but does not modulate expression of the integrins alpha 4 beta 1 and alpha 5 beta 1 in the SCF-responsive cell line MO7E. Treatment of MO7E cells with SCF (200 ng/mL) produced a transient increase in adherence to cytokine-activated human umbilical vein endothelial cells (HUVECs) or to vascular cell adhesion molecule 1 (VCAM-1)-transfected Chinese hamster ovary (CHO) cells with peak adhesion at 30 minutes and return to baseline by 60 to 90 minutes. This increase in adhesion was paralleled by increased binding of the beta 1 activation-dependent monoclonal antibody (MoAb) 15/7, as determined by flow cytometry. However, prolonged incubation of MO7E with SCF induced a marked decrease in integrin-mediated adherence, with maximal inhibition by 24 hours. No change in expression of integrins, as determined by flow cytometry, was observed with short- or long-term incubation with SCF. SCF-treated cells were still able to respond to phorbol esters and to the activating beta 1 MoAb 8A2 with increased adherence, but not to the level seen in control cells. This suggests that a subpopulation of expressed alpha 4 beta 1 and alpha 5 beta 1 integrins is disengaged by prolonged incubation with SCF.

Animals↗
PubMed1995

An NMDA receptor antagonist reduces ethanol preference in untrained but not trained rats.

Rats were exposed to a potent antagonist of NMDA receptors, 2-amino-5 phosphonovalerate (AP5) by implanting osmotic pumps delivering AP5 (test group) or the vehicle for AP5 (control group) into the cerebrospinal fluid through previously implanted cannulas. A week later they were given the choice of 6% ethanol or water for 1 h. The control group had a significantly greater preference for ethanol than the test group after the first 5 days. Another group of rats was trained almost to a 100% criterion in preferring ethanol to water in the hour the choice was available. Then they were implanted with osmotic pumps and cannulas and administered AP5. These trained rats showed no significant difference between control and test groups in preference for ethanol after the first 5 days. The results support the hypothesis that activation of NMDA receptors is involved in learning to drink ethanol.

2-Amino-5-phosphonovalerate↗
PubMed1995

Subcapsular intratesticular assay: a preliminary screening method for putative male antifertility drugs.

The subcapsular intratesticular assay was developed for the preliminary screening of male antifertility agents. In the assay, small amounts of the test sample are injected twice, 1 week apart just beneath the tunica albuginea of the testes of rats. The rats were killed within 3 weeks of treatment to examine epididymal spermatozoa and the histology of the testis and epididymis. There is an excellent agreement between this method and the modified MB-50 assay developed by the World Health Organization. To date, more than 300 samples have been assayed, of which the results of 10 plant-derived compounds are reported here. Not one active sample has been missed as confirmed by the modified MB-50 method, although one compound which was positive in the subcapsular assay did not show antifertility action in the in-vivo assay. Compared with the latter assay, the advantages of this method are: simplicity, time-saving, rat-saving and the need for far smaller amounts of test sample. The absence of false-negative results makes the subcapsular intratesticular assay applicable as a preliminary screening method to test potential male fertility regulating agents.

Administration, Oral↗
PubMed1995

Thrombopoietin expands erythroid progenitors, increases red cell production, and enhances erythroid recovery after myelosuppressive therapy.

Thrombopoietin (TPO), the ligand for the receptor protooncogene c-mpl, has been cloned and shown to be the critical regulator of platelet production. Several features of c-Mpl expression, including its presence on erythroid cell lines, and the panmyeloid transformation characteristic of myeloproliferative leukemia (MPL) viral disease led us to investigate whether this receptor-ligand system may play a role in erythropoiesis. We report that although TPO alone did not support the growth of either early or late erythroid progenitors, it acted in synergy with erythropoietin to expand these populations. Moreover, while the effects on erythropoiesis in normal animals were modest, TPO greatly expanded the number of erythroid progenitors and blood reticulocytes and was associated with accelerated red cell recovery in myelosuppressed mice. Together, these data strongly suggest that erythroid progenitors respond to TOP and that this newly cloned cytokine, critical for platelet production, can augment erythropoiesis in states of marrow failure.

Animals↗
PubMed1995

Kinetics and products of OH radical addition to baicalin.

The reaction of OH radicals with baicalin was studied by pulse and gamma radiolysis in N2O-saturated aqueous solutions. The addition of an OH radical to baicalin produced an intermediate species that shows a maximum UV absorbance band at 420 nm and decays with a unimolecular rate constant of 1.25 x 10(4) s-1 and a bimolecular rate constant of 6.1 x 10(8) mol-1 dm3 s-1. Four major final products were obtained by gamma irradiation, and their structures were identified. Two of them (P1 and P2) are suggested to originate from the unimolecular reaction and the other two (P3 and P4) from the bimolecular reaction of the radicals.

Flavonoids↗
PubMed1994

Promotion of megakaryocyte progenitor expansion and differentiation by the c-Mpl ligand thrombopoietin.

The development of blood cells including expansion of megakaryocyte progenitor cells requires the interplay of marrow stromal cells and polypeptide cytokines. Recently, characterization of c-Mpl, the receptor encoded by the proto-oncogene c-mpl, revealed structural homology with the haematopoietic cytokine receptor family, and its involvement in megakaryocyte development. We report here that the ligand for c-Mpl is relatively lineage specific, works both alone and synergistically with early acting cytokines to support megakaryocyte colony formation, and acts at a late stage of development to increase megakaryocyte size, polyploidization and expression of differentiation markers. In vivo, c-Mpl ligand stimulates platelet production by greatly expanding marrow and splenic megakaryocytes and their progenitors, and by shifting the distribution of megakaryocyte ploidy to higher values. Thus, as c-Mpl ligand has the expected characteristics of the major regulator of megakaryocyte development, we propose that it be termed thrombopoietin.

Acetylcholinesterase↗
PubMed1994

Human umbilical vein endothelial cells display high-affinity c-kit receptors and produce a soluble form of the c-kit receptor.

Stem cell factor (SCF) is a hematopoietic growth factor produced by fibroblasts and endothelial cells that stimulates the growth of primitive hematopoietic cells. SCF triggers cell growth by binding to the c-kit receptor. Because endothelial cells can respond to certain hematopoietic growth factors, we tested human umbilical vein endothelial cells for display of the c-kit receptor and examined the effect of SCF on endothelial cell proliferation, adhesion molecule expression, and production of tissue factor. Quantitative binding experiments with 125I-SCF showed both high-affinity (Kd = 42 pmol/L) and low-affinity (Kd = 1.7 nmol/L) c-kit receptors. There were approximately 1,100 high-affinity c-kit receptors, and 5,400 low-affinity c-kit receptors per endothelial cell. Enzyme immunoassays showed that endothelial cells released soluble c-kit receptor and SCF. The transmembrane form of SCF was detected by indirect immunofluorescence analysis using monoclonal or polyclonal anti-SCF receptor antibodies. The addition of SCF (100 ng/mL) did not alter endothelial cell proliferation over a 7-day period. Similarly, there was no change in the release of tissue factor or expression of inducible endothelial adhesion molecules (intercellular adhesion molecule-1, endothelial-leukocyte adhesion molecule-1, and vascular cell adhesion molecule-1) measured by enzyme-linked immunosorbant assay at 4 and 24 hours after SCF addition. The neutralizing anti-c-kit receptor monoclonal antibody SR-1 blocked binding of 125I-SCF to the c-kit receptor by 98% but did not alter endothelial cell proliferation or adhesion-molecule expression. c-kit receptors were also detected on adult endothelial cells lining small blood vessels in normal human lymph nodes. These data indicate that normal human endothelial cells produce SCF and show high-affinity c-kit receptors that have the capacity to dimerize. The lack of response to exogenous SCF may be because of intracellular activation of the c-kit receptor via autocrine production of SCF. Alternatively, SCF and c-kit may play a role other than stimulation of proliferation, adhesion-molecule display, or tissue factor production by endothelial cells. The production of soluble c-kit receptors by normal human endothelial cells may serve to regulate the bioactivity of SCF within the bone marrow microenvironment.

Cell Adhesion Molecules↗
PubMed1994

The increased ethanol preference in rats induced by choice, darkness, or drugs is reduced by ritanserin.

We tested the hypothesis that ritanserin, a serotonin S2 antagonist, reduces voluntary and induced forms of ethanol drinking. We gave 10 mg/kg ritanserin IP or SC to groups of rats given either a) a free choice between 3% ethanol and water, or b) kept in the dark for 5 weeks and given a choice between a range of ethanol concentrations (3-25%) and water, or c) implanted with osmotic pumps filled with tetrahydro-beta carboline and given a choice between a range of ethanol concentrations and water. In each case, ritanserin significantly reduced ethanol consumption and ethanol preference for 8-10 days after the last injection.

Animals↗
PubMed1994

Epigenetic lesions at the H19 locus in Wilms' tumour patients.

To test the potential role of H19 as a tumour suppressor gene we have examined its expression and DNA methylation in Wilms' tumours (WTs). In most WTs (18/25), H19 RNA was reduced at least 20-fold from fetal kidney levels. Of the expression-negative tumours ten retained 11p15.5 heterozygosity: in nine of these, H19 DNA was biallelically hypermethylated and in two cases hypermethylation locally restricted to H19 sequences was also present in the non-neoplastic kidney parenchyma. IGF2 mRNA was expressed in most but not all WTs and expression patterns were consistent with IGF2/H19 enhancer competition without obligate inverse coupling. These observations implicate genetic and epigenetic inactivation of H19 in Wilms' tumorigenesis.

Alleles↗
PubMed1993

Stem cell factor influences the proliferation and erythroid differentiation of the MB-02 human erythroleukemia cell line by binding to a high-affinity c-kit receptor.

Stem cell factor (SCF) acts in synergy with other growth factors such as erythropoietin (Epo), granulocyte-macrophage colony-stimulating factor (GM-CSF), or interleukin-3 (IL-3), to stimulate the growth of primitive hematopoietic cells. Because of the prominent role of CSF in the maintenance of normal erythropoiesis in vivo, we examined the effects of SCF on the Epo-inducible human erythroleukemia cell line MB-02, and characterized the c-kit receptor in these cells. MB-02 cells cultured in serum-containing media do not survive in the absence of exogenous growth factors, but the addition of SCF, Epo, or IL-3 as a single factor enhanced MB-02 survival. Furthermore, in the presence of Epo, SCF (5 to 25 ng/mL) enhanced MB-02 proliferation in a dose-dependent manner, and increased the relative and absolute number of benzidine-positive cells generated. SCF also enhanced cell proliferation in the presence of either IL-3 or low concentrations of GM-CSF. A neutralizing anti-c-kit receptor monoclonal antibody (SR-1) blocked binding of 125I-SCF to MB-02 cells by 98%, and the effect of SCF on MB-02 growth, c-kit receptor-binding parameters were quantitated by equilibrium-binding experiments with 125I-SCF. MB-02 cells display a single class of high-affinity (50 pmol/L) c-kit receptors, with approximately 8,000 receptors per cell. The molecular weight of the c-kit receptor was determined by affinity cross-linking 125I-SCF to MB-02 cells. 125I-SCF-c-kit receptor complexes of approximately 155,000 and approximately 310,000 daltons were found, likely representing the monomeric and dimeric forms of the c-kit receptor. The binding affinity and molecular weight of the c-kit receptor on MB-02 cells are similar to those of normal human marrow cells. These results suggest that SCF synergizes with Epo to influence not only the proliferation but the erythroid differentiation of MB-02 cells. Thus, the MB-02 cell line may be a useful model in which to investigate the molecular mechanisms of SCF action.

Antibodies, Monoclonal↗
PubMed1993

[Intratesticular subcapsular assay, a simple method for the preliminary screening of male contraceptives].

The MB-50 protocol worked out for long by the World Health Organization has been widely employed for the screening of male contraceptives. With this method, 40-80 rats will be used for 1 sample and the course of study will be about 3 months. In view of the high expenses and long duration of time needed, it seems to be not suitable for a preliminary screening. We have exploited a much simpler method, the intratesticular subcapsular assay. The method is as follows: Puncture the scrotal skin and the tunica albuginea at the equatorial plane of the testis just opposite to the epididymis with an intradermal needle fixed at a tuberculin syringe. Then push on the needle for 3-4 mm along the equator parallel to the tunica albuginea. Inject 100 microliters of solution containing 0.1-5 mg of the sample to each testis. To the controls, only the vehicle is injected. One week later the same procedure is repeated and 2 more weeks later, the animals are sacrificed and the data are observed (see Table 1 and 2). In this paper, 7 samples screened with this method were compared with results of the MB-50 method, no positive sample was missed except one false positive sample. These results indicate the coincidence between these two methods.

Animals↗
PubMed1992

Blasts from patients with acute myelogenous leukemia express functional receptors for stem cell factor.

Stem cell factor (SCF) acts in concert with lineage-specific growth factors to stimulate the growth of hematopoietic colonies. To determine if neoplastic human hematopoietic cells would also respond to SCF, we cultured marrow mononuclear cells from 20 patients with newly diagnosed acute myelogenous leukemia (AML) and two normal donors with SCF, interleukin 3 (IL-3), granulocyte-macrophage colony-stimulating factor (GM-CSF), or combinations of growth factors in semisolid medium, and assessed colony growth. SCF receptors (c-kit receptors) were quantitated by equilibrium binding studies with 125I-SCF, and binding parameters were estimated using the ligand program. The cellular distribution of c-kit receptors was determined by autoradiography. Our results show that SCF alone or in combination with IL-3 or GM-CSF increased both the number and size of colonies in 10 of the patients. Receptors for SCF were identified on the blasts from all 20 AML patients. The number of receptors ranged from 600 to 29,000 per cell. In the majority of patients, both high- and low-affinity binding sites were identified. Neither the number of receptors per cell nor the finding of one or two classes of receptors correlated with growth response to SCF. Autoradiographic analysis of 125I-SCF binding to normal marrow mononuclear cells revealed grains associated with blasts and megakaryocytes. Grain counts on blasts from 10 AML patients and on normal marrow blasts suggested that high-affinity c-kit receptor expression on AML blasts is lower than or similar to that of normal blasts. These results identify c-kit receptors on human AML blasts, and indicate that SCF acts synergistically with IL-3 or GM-CSF to stimulate colony growth from the marrow cells of a portion of patients with AML.

Autoradiography↗
PubMed1992

Isolation and characterization of a monoclonal antibody that recognizes the human c-kit receptor.

Stem cell factor (SCF) stimulates the growth of burst-forming unit-erythroid (BFU-E) and colony-forming unit granulocyte-macrophage (CFU-GM) by binding to a specific cell surface receptor. The receptor for SCF is encoded by the protooncogene c-kit. After immunizing mice with the human erythroleukemia cell line OCIM1, we obtained a monoclonal antibody (MoAb) that recognizes the human c-kit receptor. This MoAb, designated SR-1, blocks binding of 125I-human SCF to the c-kit receptor, and neutralizes the biologic effects of SCF in hematopoietic colony assays. With few exceptions, c-kit expression was identified on all hematopoietic and lymphoid cell lines tested by indirect immunofluorescent analysis using SR-1 and by binding studies with 125I-SCF. SR-1 recognizes a small fraction of normal bone marrow mononuclear cells, and these cells have the morphologic appearance of blasts. Colony assays show that BFU-E and CFU-GM display the c-kit receptor. SR-1 does not cross-react with murine c-kit protein, indicating that the binding epitopes of the human and murine c-kit receptors are antigenically distinct. This MoAb may be useful to characterize the spectrum of cells that display the c-kit receptor and to further define the role of SCF in hematopoiesis.

Animals↗
PubMed1992

ANP and naloxone reduce postdeprivation drinking after subfornical organ lesions.

We tested a report that atrial natriuretic peptide (ANP) injected into, or near, the subfornical organ (SFO) will reduce the water consumption of previously water deprived rats and that suggested ANP acts on neurons in the SFO to bring about this action. We tested this suggestion and the hypothesis that the SFO is involved in the facilitation of drinking produced by opioids. ANP (5 nmol in 4 microliters, IVT) or naloxone (2 mg/ml/kg, SC, or 200 micrograms in 2 microliters, IVT) when given to rats deprived of water for 16 h (SC treatment) or 23 h (IVT treatment) significantly depressed postdeprivation drinking measured at 15 and 60 min. Rats with complete, partial, or control lesions of the SFO, after the same treatment, also showed a significant depression of postdeprivation drinking and, after 23-h deprivation, a significant hyperdipsia. There was no interaction between drug effects and lesion effects (two-factor analysis of variance, Tukey's post-hoc tests). The hyperdipsia declined exponentially and was lost 45-50 days after lesioning. Our results do not support the hypothesis that the SFO is involved in the actions of ANP or of opioids on postdeprivation drinking.

Animals↗
PubMed1992

DnaK-mediated alterations in human growth hormone protein inclusion bodies.

Protein overproduction in microbes frequently results in protein misfolding and aggregation though the molecular basis for this process is unclear. The HSP70 chaperonin, DnaK, was identified as an important factor controlling heterologous protein aggregation in Escherichia coli. Co-overproduction of DnaK significantly reduced human growth hormone (HGH) protein inclusion body formation and the extent of HGH aggregation.

Bacterial Proteins↗
PubMed1992

Protein kinase C subspecies in rabbit corneal epithelium: increased activity of alpha subspecies during wound healing.

Protein kinase C (PKC) has been implicated in cell proliferation and differentiation. Multiple forms of PKC have been isolated, principally from the brain where PKC is most abundant. In rabbit corneal epithelium, two distinct major peaks of PKC activity were resolved by hydroxyapatite column chromatography. Peak 2, with 65% of the total PKC activity, corresponds to alpha-PKC, based on its mobility in the column and Western blot analysis using specific monoclonal antibodies. Peak 1 did not react with either polyclonal or monoclonal antibodies to PKC alpha-, beta-, and gamma-isoforms suggesting the presence of isoforms specific to the corneal epithelium, or of another member of the PKC family. To investigate possible changes in the amounts of the various PKC subspecies during wound healing, the enzyme activities of the isolated subspecies were assayed 2, 5, and 7 days after corneal de-epithelialization. Two days after wounding, by which time the migratory limbal epithelium had covered the denuded area, total PKC activity was unchanged but alpha-PKC activity had increased to 77% of the total activity, compared with 65% in non-wounded epithelium. An increased proportion of alpha-PKC activity was also observed 5 and 7 days after wounding, during which time proliferation of epithelium continued. We hypothesize that alpha-PKC plays a role in long-term responses after injury such as gene expression and corneal epithelial proliferation. Moreover, these studies indicate that the cornea provides a good model of in vivo wound healing for PKC studies.

Animals↗