PubMed Health⌕ Search

Biomedical subjects

U Testa

Publications and source records attributed to U Testa.

At least 145 records · Page 8Linked to original sources
PubMed1989

Constitutive expression and abnormal glycosylation of transferrin receptor in acute T-cell leukemia.

The expression of transferrin receptors (TrfRs) was investigated in acute T-cell leukemia (T-ALL) blasts at the molecular, biochemical, immunological, and functional level. TrfRs, although not detected on quiescent T-cells from normal adults, are constitutively expressed at high level on the blasts from all T-ALL patients and bind normally to transferrin. Their number is modulated by the intracellular iron level, but is independent of exogenous interleukin 2. They also exhibit immunological and biochemical abnormalities, in that: (a) they react preferentially with monoclonal antibodies (MAb) that recognize ligand-binding domains of TrfR (42/6 and 43/31), as compared to MAbs (B3/25, OKT9) that interact with the nonligand binding domains; (b) they have a reduced molecular weight, as compared to TrfR on normal thymocytes and activated T-lymphocytes: this phenomenon is apparently related to a defective glycosylation. It is noteworthy that expression of TrfR was not observed in a large series of other types of acute leukemias, i.e., pre-B, B, and myeloid leukemias, excluding erythroleukemias. The constitutive, high level expression of TrfRs on T-ALL blasts may play a key role in the stepwise progression of this malignancy and particularly provide a proliferative advantage to T-ALL blasts as compared to normal T-lymphocytes. Furthermore, indirect evidence suggests that the glycosylation defect of TrfR on T-ALL blasts contributes to their tumorigenic capacity.

Antibodies, Monoclonal↗
PubMed1989

Iron up-modulates the expression of transferrin receptors during monocyte-macrophage maturation.

We have investigated the effect of iron on the expression of transferrin receptors (TrfRs) and ferritin chains in cultures of human peripheral blood monocytes maturing to macrophages. Monocyte-macrophage maturation is associated with a gradual rise of Trf-binding capacity in the absence of cell proliferation. At all culture times, treatment with ferric ammonium citrate induces a dose-dependent rise of the Trf-binding level as compared with nontreated cells. Scatchard analysis revealed that this phenomenon is due to an increase in receptor number rather than an alteration in ligand-receptor affinity. Biosynthesis experiments indicated that the rise in number of TrfRs is due to an increase of receptor synthesis, which is associated with a sustained elevation of the TrfR RNA level. The up-regulation of TrfR synthesis is specific in that expression of other macrophage membrane proteins is not affected by iron addition. Conversely, addition of an iron chelator induced a slight decrease of TrfR synthesis. The expression of heavy and light ferritin chains at RNA and protein levels was markedly more elevated in cultured macrophages than in fresh monocytes, thus suggesting modulation of ferritin genes at transcriptional or post-transcriptional levels. Addition of iron salts to monocyte-macrophage cultures sharply stimulated ferritin synthesis but only slightly enhanced the level of ferritin RNA, thus indicating a modulation at the translational level. These results suggests that in cultured human monocytes-macrophages, iron up-regulates TrfR expression, thus in sharp contrast to the negative feedback reported in a variety of other cell types. These observations may shed light on the mechanism(s) of iron storage in tissue macrophages under normal conditions and possibly on the pathogenesis of diseases characterized by abnormal iron storage.

Adolescent↗
PubMed1989

Three new class I HLA alleles: structure of mRNAs and alternative mechanisms of processing.

Sixteen HLA class I clones have been isolated from a SV40-transformed human fibroblast line (GM637) cDNA library. The clones, characterized by hybridization to ABC locus-specific probes and sequence analysis, correspond to transcripts from four different class I genes: A2, A10, Cw4, and Cw6 (or Cw7), as implied by cell typing. Only the A2 sequence was known. The nucleotide and deduced amino acid sequence of the new alleles are reported here, and their structural features are discussed. Two independent cDNAs of A2 specificity display an unusual polyadenylation site located 100 bp upstream from the canonical one. Moreover, two cDNAs pertaining to the same C allele display two alternative mechanisms of splicing, which cause either presence or absence in mature transcripts of the transmembrane exon 5 sequence. Transcripts missing this region are predicted to synthesize a nonmembrane-bound, secreted antigen. A soluble protein, specifically reacting with class I-specific HLA antibodies, is found in the supernatant of the GM637 cells. The significance of HLA class I transcripts generated by differential processing is discussed.

Alleles↗
PubMed1989

Expression of protein kinase C genes during ontogenic development of the central nervous system.

We have analyzed the RNA expression of three protein kinase C (PKC) genes (alpha, beta, and gamma) in human and murine central nervous systems during embryonic-fetal, perinatal, and adult life. Analysis of human brain poly(A)+ RNA indicates that expression of PKC alpha and beta genes can be detected as early as 6 weeks postconception, undergoes a gradual increase until 9 weeks postconception, and reaches its highest level in the adult stage, and that the PKC gamma gene, although not expressed during embryonic and early fetal development, is abundantly expressed in the adult period. Similar developmental patterns were observed in human spinal cord and medulla oblongata. A detailed analysis of PKC gene expression during mammalian ontogeny was performed on poly(A)+ RNA from the brain cells of murine embryos at different stages of development and the brain cells of neonatal and adult mice. The ontogenetic patterns were similar to those observed for human brain. Furthermore, we observed that the expression of PKC gamma is induced in the peri- and postnatal phases. These results suggest that expression of PKC alpha, beta, and gamma genes possibly mediates the development of central neuronal functions, and expression of PKC gamma in particular may be involved in the development of peri- and postnatal functions.

Animals↗
PubMed1989

Granulocyte-macrophage colony-stimulating factor reactivates fetal hemoglobin synthesis in erythroblast clones from normal adults.

Reactivation of fetal hemoglobin (HbF, alpha 2 gamma 2) synthesis was previously reported in normal human adult erythroblast colonies ("bursts") generated by erythroid progenitors (BFU-E) in fetal calf serum-supplemented (FCS+) semisolid cultures stimulated with erythropoietin (Ep). Our studies focused on the reactivation of HbF synthesis in normal adult erythroid bursts generated by peripheral blood mononuclear cells (PBMCs) seeded in FCS+ methylcellulose culture. Reactivation is almost totally suppressed when (a) PBMCs are grown in optimized FCS- culture, or (b) PBMCs are first stringently depleted of monocytes and then plated in FCS+ medium (ie, BFU-E growth in FCS+ Mo- culture). In both experimental conditions, the proliferation of lymphocytes and macrophages interspersed among colonies is drastically reduced, and the cloning efficiency of granulocyte-macrophage (GM) progenitors is sharply diminished. In either case, addition of biosynthetic GM colony-stimulating factor (GM-CSF) induces a dose-related increase of HbF synthesis up to the level in FCS+ culture, with even more elevated values on delayed addition of Ep. A dose-related increase was also observed in erythroblast clones generated by highly purified BFU-E. These results suggest that reactivation of HbF synthesis in normal adults is at least in part mediated by GM-CSF. Furthermore, they imply intriguing hypotheses on the mechanism(s) of perinatal Hb switching. Finally, they raise the possibility of reactivation of HbF synthesis in beta-thalassemia and sickle cell anemia by GM-CSF therapy.

Biological Factors↗
PubMed1989

[Use of colony-stimulating factors as possible therapeutic agents].

The first clinical trials have already indicated that long-term administration of recombinant hemopoietic growth factors can be a clinically effective procedure for correcting abnormalities in blood cell formation. Thus, the colony stimulating factors (CSFs) have proved to be highly effective agents in stimulating the formation of granulocytes, monocytes and eosinophils both in animals and man, and in enhancing resistance to otherwise lethal infections. They offer the bright prospect of use in any clinical situation in which the formation or function of these cells is abnormally low (due to the reduced production of myeloid elements such as occurring in aplastic anemias or to the killing of myeloid progenitors and precursors caused by anti-tumor chemotherapy) and causes a real risk of life-threatening infections. From initial clinical trials the CSFs, particularly GM-CSF and G-CSF, can be anticipated to perform effectively in offering the clinician the possibility of active intervention in sustaining myelopoiesis.

Agranulocytosis↗
PubMed1989

Erythropoietin alone induces erythroid burst formation by human embryonic but not adult BFU-E in unicellular serum-free culture.

Erythroid progenitors (BFU-E) from adult human peripheral blood generate erythroid bursts in semisolid culture supplemented with at least two growth factors, ie, erythropoietin (Ep) and interleukin-3 (IL-3) or granulocyte-macrophage colony-stimulating factor (GM-CSF). We have analyzed the hematopoietin(s) requirement of human embryonic BFU-E, as compared to that of adult peripheral blood progenitors: This was basically evaluated in fetal calf serum-free (FCS-) methylcellulose culture of partially or highly purified progenitors treated with human recombinant hemopoietins. At a low seeding concentration (2 x 10(3) cells/dish) purified embryonic BFU-E generated erythroid bursts when treated only with Ep: Further addition of IL-3 or GM-CSF had no effect on BFU-E cloning efficiency, although the size of bursts was increased in a dose-dependent manner, particularly with IL-3. At a similar seeding concentration (ie, 10(3) cells/dish), purified adult BFU-E efficiently generated erythroid bursts in the presence of Ep and GM-CSF or IL-3, while only few small erythroid colonies were observed in the presence of Ep alone. In a final series of experiments, unicellular FCS-cultures of purified embryonic BFU-E gave rise to erythroid bursts in the presence of Ep alone. Furthermore, the cloning efficiency induced by Ep was unmodified by further addition of GM-CSF or IL-3. Unicellular FCS- cultures of highly purified adult peripheral blood progenitors generated no erythroid bursts in the presence of Ep alone. The addition of GM-CSF or IL-3 was required to generate BFU-E colonies. These studies indicate that in human embryonic life, BFU-E require only Ep for efficient erythroid burst formation, while IL-3 and GM-CSF essentially enhance the proliferation of early erythropoietic precursors.

Cells, Cultured↗
PubMed1989

Zinc modulates the mitogenic activation of human peripheral blood lymphocytes.

In the present study we have investigated the effect of Zn salts on the mitogenetic activation of human peripheral blood lymphocytes (PBL). Our results show that Zn2+ enhances the level of DNA synthesis in human T lymphocytes stimulated by a mitogenic lectin, phytohemagglutinin (PHA); this effect seems to be mediated through an enhanced expression of both interleukin-2 (IL-2) and transferrin (Trf) receptors. We have also analyzed the mitogenic effect of Zn2+ alone on PBL, in the absence of other mitogenic stimuli. In this regard we have identified large light density T lymphocytes as the PBL population which is activated to proliferate by Zn2+. Finally, we showed that Zn2+ stimulates natural killer (NK) activity; this effect is apparently not due to a direct action on NK lymphocytes, but is related to endogenous cytokines released by accessory cells which in turn stimulate the cytolytic activity of NK lymphocytes.

Humans↗
PubMed1989

Altered growth factor sensitivity in EL2 rat fibroblasts: influence of this biological characteristic on cell growth.

Extensive evidence indicate that platelet-derived growth factor (PDGF) and epidermal growth factor (EGF) play a key role in the stimulation of the 3T3 fibroblast replication: in this connection, PDGF and EGF act as a competence and a progression factor, respectively. We have previously demonstrated that EGF alone leads density-arrested EL2 rat fibroblasts to synthesize DNA and proliferate in serum-free cultures. Here, we have analyzed the role of EGF in the control of EL2 cell proliferation. Our data show a dose-related effect of EGF on DNA synthesis and cell growth, with maximal stimulation for both parameters at 20 ng/ml. On the other hand, autocrine production of PDGF or PDGF-like substances by EL2 cells is seemingly excluded by experiments with anti-PDGF serum or medium conditioned by EL2 fibroblasts. EGF binding studies show that EL2 cells possess high affinity EGF receptors, at a density level 3 to 4-fold higher than other fibroblastic lines. In addition, EL2 cells show a normal down-regulation of EGF receptors, following exposure to EGF, but PDGF, fibroblast growth factor (FGF), transforming growth factor beta (TGF beta) and bombesin have not decreased the affinity of EGF receptor for its ligand. Moreover, in EL2 cells, the EGF is able to induce the synthesis of putative intracellular regulatory proteins that govern the PDGF-induced competence in 3T3 cells. Our data indicate that EGF in EL2 cells may act as both a competence and a progression factor, via induction of the mechanisms, regulated in other cell lines by cooperation between different growth factors.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

TGF beta induces a sustained c-fos expression associated with stimulation or inhibition of cell growth in EL2 or NIH 3T3 fibroblasts.

We have previously indicated that epidermal growth factor (EGF) plays a fundamental role in the proliferation control of EL2 rat fibroblast line. It is shown here that transforming growth factor beta (TGF beta) stimulates both DNA synthesis and proliferation of EL2 cells, while exerting an inhibitory effect on the growth of murine NIH-3T3 fibroblasts. We also report the effect of TGF beta and EGF on c-fos expression in EL2 cells, as compared to that of TGF beta in NIH-3T3 fibroblasts. In EL2 cells EGF induces a transient c-fos expression at both mRNA and protein level, as previously observed in NIH-3T3 fibroblasts treated with platelet-derived or fibroblast growth factor (PDGF, FGF). Conversely, TGF beta induces in EL2 cells a sustained expression of fos mRNA and protein, which are still detectable at least 24 and 7 hr after treatment respectively. In NIH-3T3 fibroblasts TGF beta causes a sustained fos RNA expression, which is not associated, however, with detectable fos protein. We conclude that in fibroblasts stimulated by mitogens c-fos expression may be differentially modulated, depending of the growth factor and the cell line. This is seemingly due to differential regulation of fos gene expression, not only at the transcriptional and/or post-transcriptional level (transient or sustained fos RNA induction by EGF or TGF beta in EL2 cells), but also at the translational level (fos protein(s) induction by TGF beta in EL2 but not NIH-3T3 fibroblasts, possibly related to the stimulatory vs inhibitory effect of this factor on the growth of the former vs the latter line).

Animals↗
PubMed1988

Effect of endogenous and exogenous interferons on the differentiation of human monocyte cell line U937.

The effect of interferons (IFNs) on the differentiation of hematopoietic cells was examined with the human monocyte cell line U937. The differentiation of U937 was induced by hydroxyvitamin D3 and was evaluated through the study of specific markers. The induction of the U937 differentiation was associated with a production of IFN and with a marked increase in (2'5') oligoadenylate synthetase. Addition of anti-IFN-alpha/beta antibodies inhibited the enhancement of (2'5') oligoadenylate synthetase and reduced the inhibitory effect of hydroxyvitamin D3 on cell growth. Nevertheless, neutralization of endogenous IFN excreted during U937 cell maturation did not modify the expression of the differentiation markers examined. Exogenous natural IFN-alpha, IFN-beta, or recombinant (r) IFN-gamma, when added to the culture medium, did not promote a "global" U937 differentiation. Most of the differentiation markers, except for reduction of nitroblue-tetrazolium, were not induced by IFN-alpha or -beta. However, rIFN-gamma was able to induce the appearance of several monocytic membrane markers at an extent comparable or slightly inferior to that elicited by hydroxyvitamin D3. Different effects on the expression of HLA antigens were obtained with these IFNs: IFN-alpha or -beta enhanced mainly class I HLA antigen expression, whereas rIFN-gamma increased selectively the expression of class II HLA DC1 but not HLA DR antigens. In contrast, phytohemagglutinin-leukocyte conditioned medium elicited a marked and selective enhancement of the expression of HLA-DR antigens. This induction of HLA DC1 antigens by rIFN-gamma was not observed in two other leukemic cell lines (HL60 and HEL). The present study shows that IFN-alpha or -beta may participate in the antiproliferative effect occurring during cellular differentiation, while IFN-gamma may be involved in the induction of the expression of specific monocytic markers involved in cellular immunoregulation.

2',5'-Oligoadenylate Synthetase↗
PubMed1988

Human platelet alpha granules contain a nonspecific inhibitor of megakaryocyte colony formation: its relationship to type beta transforming growth factor (TGF-beta).

Whole blood serum (WBS) and platelet-poor plasma-derived serum (PDS) from the same normal subject were compared for their abilities to support human megakaryocyte (MK) colony formation. In all cases, PDS promoted the growth of a higher number (20-50%) of MK colonies than did WBS. Increasing amounts of WBS decreased the number of colonies, whereas increasing concentration of PDS had no marked effects. Crude platelet extracts or platelet secretory products from thrombin-activated platelets also elicited an inhibition of MK colony formation in a dose-dependent manner. A complete inhibition was found for a dose equivalent to 1.10(9) platelets/ml and a 50% inhibition in a range of 1.10(7)-1.10(8) platelets/ml. These platelet products were also inhibitory for erythroid progenitor growth. Platelets from two patients with gray platelet syndrome elicited only a minor inhibition of MK growth, suggesting that the platelet alpha granule is the origin of this inhibition. When platelet extracts were acid-treated, the biological activity of the inhibitor on CFU-MK and CFU-E growth was 20-50-fold higher. In addition, a potent stimulatory activity on the growth of day 7 CFU-GM was observed. The enhancement of biological activities by acid treatment suggests that type beta transforming growth factor (TGF-beta) could be involved in this platelet inhibitory activity. The homogeneous native TGF-beta (from 1 pg to 1 ng/ml) produced the same effects previously induced by platelet products. It totally inhibited CFU-MK growth (at a 500 pg/ml), it inhibited CFU-E growth, and it stimulated growth of day 7 CFU-GM in the presence of a colony-stimulating factor. The inhibition of CFU-MK growth was also observed on purified progenitors. In conclusion, these results suggest that TGF-beta may be implicated in negative autocrine regulation of megakaryopoiesis. However, since this molecule has ubiquitous biological activities, its physiologic relevance as a normal regulator of megakaryopoiesis requires further investigation.

Blood Physiological Phenomena↗
PubMed1988

Mechanisms underlying T-lymphocyte activation: mitogen initiates and IL-2 amplifies the expression of transferrin receptors via intracellular iron level.

Peripheral blood mononuclear cells (PBM) pulsed with lectin (PHA or Con A for 0.25-3 hr) show a low expression of interleukin-2 and transferrin receptors (IL-2Rs, TfRs) and a mild decline of intracellular ferritin level, compared to control cultures grown in continuous presence of mitogen. Interestingly, lectin-pulsed PBM do not release detectable amounts of IL-2 in the medium. Furthermore, expression of TfRs in these lymphocytes is not inhibited by addition of excess anti-IL-2 neutralizing monoclonal antibody, but is significantly inhibited by treatment with iron salts. These observations suggest that mitogen triggers an IL-2-independent expression of TfRs, at least in part via a decrease of intracellular iron level. Addition of either recombinant IL-2 (rIL-2) or an iron chelator (picolinic acid) to lectin-pulsed PBM induces both a marked enhancement of TfR synthesis and a sharp decline of intracellular ferritin level, which are comparable to the corresponding pattern observed in control cultures. Conversely, addition of iron salts fully inhibits the increase of TfR expression induced by rIL-2. These observations strongly suggest that the enhanced TfR synthesis elicited by rIL-2 is mediated by depletion of a regulatory intracellular iron pool. In line with these studies, greater than 99% purified T lymphocytes stimulated by lectin show a low expression of TfRs, which is markedly enhanced by addition of exogenous rIL-2. Altogether, we postulate that: (i) in resting T lymphocytes the gene encoding TfR is apparently in a 'closed' configuration; (ii) even in the absence of IL-2 activity, a mitogen pulse is sufficient to initiate the expression of TfRs, at least in part via a decline of intracellular iron level; and (iii) TfR synthesis is then largely amplified by IL-2, again via a decrease of the size of a regulatory intracellular iron pool.

Cells, Cultured↗
PubMed1988

Activation and proliferation of normal resting human T lymphocytes in serum-free culture: role of IL-4 and IL-6.

Purified human T lymphocytes, completely depleted of accessory cells [i.e. monocytes, large granular lymphocytes (LGL) and B lymphocytes], have been grown in serum-free culture in presence of a mitogenic lectin (phytohaemagglutinin, PHA) and different recombinant cytokines. Only IL-2 and IL-4 induced a marked stimulation of [3H] thymidine ([3H]TdR) uptake, cell proliferation and expression of activation markers [transferrin receptor (TrfR), IL-2R]. The other cytokines (IL-1 alpha, IL-1 beta, IFN-gamma, GM-CSF, TNF-alpha) had no significant effect, except for a moderate, but significant, stimulation of [3H]TdR uptake induced by IL-3. Simultaneous addition of IL-4 and anti-IL-2 neutralizing monoclonal antibodies (mAb) did not modify the effects induced by IL-4 alone. Furthermore, IL-2 was not detected in the supernatant of T cells grown in the presence of PHA and IL-4. Thus, our results indicate that IL-4 acts on T lymphocytes independently of IL-2. We also observed that IL-6 moderately activates DNA synthesis in PHA-stimulated T lymphocytes, but markedly potentiates the proliferative effect of suboptimal amounts of IL-2. In conclusion, the present study suggests that B-cell growth factors, in addition to IL-2, control the proliferation of normal circulating T lymphocytes.

Cells, Cultured↗
PubMed1987

Specific modulation of surface receptors in J.774 macrophages by anchorage.

The J.774 murine macrophage cells were cultured in suspension in Teflon flasks. When allowed to attach on culture plastic dishes, a 2-3-fold increase in transferrin binding was observed. This occurred in 10 min, reached a steady state at 60 min, remained stable for several hours and was reversible after resuspension of the cells at 37 degrees C. The phenomenon was not dependent on the synthesis of new protein. An opposite change of acetyl LDL receptors was observed, with a threefold decrease of the binding 1 h after the attachment of the cells. The increase of transferrin binding affected almost equally the cell surface and the intracellular sites; therefore it could not be related to a simple shift between these two compartments. It is suggested that the attachment of the cells induced a recruitment of binding sites from a 'silent' pool of receptors. Serum factors, as well as phorbol esters and db-cAMP potentiated the effect of anchorage.

Animals↗
PubMed1987

Neonatal jaundice and severity of glucose-6-phosphate dehydrogenase deficiency in Sardinian babies.

We have investigated the association of neonatal jaundice (NNJ) and G6PD deficiency in consecutive births in a Northern Sardinian hospital. After excluding known causes for NNJ, and after correcting for the incidence of NNJ from unknown causes, we estimated that 20% of G6PD deficient male newborns develop NNJ resulting from their enzyme deficiency. By analyzing in detail 100 G6PD deficient babies we found no differences in birth weight or haemoglobin level between those without and those with NNJ, four of whom required exchange transfusion. We further showed by an accurate quantitative method that the residual G6PD enzyme activity was not significantly lower in G6PD deficient babies with NNJ compared to G6PD deficient babies without NNJ.

Bilirubin↗