Search PubMed⌕ Search

Biomedical subjects

R Gambari

Publications and source records attributed to R Gambari.

At least 163 records · Page 9Linked to original sources
PubMed1978

Terminal stages of erythroid differentiation: persistent production of globin messenger RNA is necessary to sustain globin synthesis.

The kinetic relationship between the globin mRNA accumulation and the rate of synthesis of globin chains was studied during the terminal stages of differentiation in erythroid cells derived from the yolk sac of mouse fetuses. RNA derived from the whole cells and from different cell compartments were hybridized to DNA complementary to embryonic globin mRNA. The relative proportion of embryonic globin RNA molecules and their absolute number per cell were estimated on the 11th, 12th, and 13th days of mouse fetal development. During erythroid terminal differentiation globin mRNA became progressively predominant on polyribosomes along with the progressive specialization of cell functions. The number of embryonic globin RNA molecules per cell remained constant while yolk sac erythroid cells underwent two rounds of cell division. These data indicate that the transcription of globin genes is operative throughout the last stages of terminal differentiation and that there is no detectable storage of globin RNA sequences in these cells. The rates of accumulation of mRNA molecules and of globin synthesis both seem correlated to the length of the cell cycle.

Animals↗
PubMedDate not supplied

Hypomethylation of the human HLA-DR alpha gene in breast carcinomas and autologous metastases.

The methylation pattern of the human HLA-DR alpha gene was analyzed in normal breast tissues, breast primary tumors and lymphonodal metastases isolated from patients carrying breast carcinomas. In breast adenomas and also in normal tissues (including breast, muscle, brain, sperm and T- and B-lymphocytes), the HLA-DR alpha gene is hypermethylated at the CCGG and GCGC sites. In all tissues studied, the only constantly unmethylated region is located in the 5' portion of the gene, near the promoter sequence. Further, the results indicate that the HLA-DR alpha gene is hypomethylated in carcinomas and in the relative metastatic lymph nodes. It is suggested that hypomethylation of the human HLA-DR alpha gene could be proposed as a molecular marker of malignant breast tumors.

Adenoma↗
PubMedDate not supplied

Inhibition of 'in vitro' tumor cell growth by aromatic polyamidines exhibiting antiproteinase activity.

Aromatic polyamidines containing two, three or four benzamidine residues inhibit proteinase activity and proliferation of different human tumor cell lines, including leukemic (K562, HEL), melanoma (Colo 38) and B-lymphoid (WI-L2) cell lines. In addition, the benzamidine derivatives analysed in the present study inhibit cell growth of the Chinese hamster FHO6T1-1 cell line, obtained after transfection of primary lung cells with the activated human T24-Ha-ras-1 oncogene. After treatment of FHO6T1-1 cells with benzamidine derivatives, a sharp decrease of the content of Ha-ras-1 mRNA was found, but not of transferrin receptor mRNA. We found that inhibition of cell proliferation by tetra-benzamidine derivatives is not restricted to tumor cells, but concerns also non-tumorigenic cell lines as well as normal primary fibroblasts. Therefore, our analysis was extended to di- and tri-benzamidine derivatives, which could be proposed as useful substrates in the synthesis of drug-conjugated monoclonal antibodies or growth factors. The data obtained demonstrate that these latter compounds and their halo-derivatives also exhibit strong antiproliferative effects on in vitro cultured cells.

Amidines↗
PubMedDate not supplied

Methylation pattern of the HLA-DR alpha gene in human tissues.

The CCGG and GCGC sites of the human HLA-DR alpha gene are hypermethylated in human tissues (including B-lymphocytes, T-lymphocytes, muscle, brain, sperm, skin, kidney, suprarenal and mammary glands) and three B-lymphoid cell lines. Therefore, the HLA-DR alpha gene can be transcribed even though extensively methylated. The only exception to the hypermethylated state of the HLA-DR alpha gene is represented by one or both of the two HhaI sites (H1 and H2) localized in the 5' portion of the gene. Analysis of the computer-generated secondary structure of the HLA-DR alpha mRNA suggests that the H1 and H2 sites belong to a region (5'-GAGCGCCCA-3'/5'-UGAGCGCUC-3') exhibiting extensive base pairing. Therefore, unmethylation of these CG sites can contribute in preventing mCG----TG/CA changes in this region, which would lead to extensive alterations of the secondary structure of the 5' portion of the HLA-DR alpha MRNA. On the other hand, the selective pressure to maintain unaltered the methylated CG dinucleotides in the coding regions of the HLA-DR alpha gene could be due to codon restrictions, since the majority of the methylation-related CG----TG or CG----CA variations would generate aminoacid changes. Accordingly, the analysis of different HLA-DR alpha genomic sequences indicates that variations of the CpG dinucleotides occur only in the non-coding portions of the HLA-DR alpha gene.

Base Sequence↗
PubMedDate not supplied

Kinetics of bromocriptine release from microspheres: comparative analysis between different in vitro models.

This paper describes how the use of different in vitro experimental systems can influence the determination of (a) the drug release profile from microparticles and (b) the interpretation of the release mechanism(s). We employed, as model dosage form, the Parlodel LA, a recently marketed microsphere system especially designed for bromocriptine-controlled delivery. The release kinetics of bromocriptine from microspheres were determined by using two different experimental approaches: a dialysis method and a flow-through cell method. From the comparison of the obtained data it clearly appears that different in vitro experimental models lead to distinct results in terms of drug availability. On the contrary both series of data can be convincingly fitted with the same mathematical equation, giving almost identical results in terms of postulated release mechanism. Taken together these results indicate that different experimental approaches should always be employed to determine drug release kinetics from microparticles in order to obtain more reliable information on the therapeutic dose (bioavailable drug, for in vivo experiments) and on the uniformity of different batches of microspheres.

Bromocriptine↗
PubMedDate not supplied

Production and in vitro evaluation of gelatin microspheres containing an antitumour tetra-amidine.

The preparation and characterization of gelatin microspheres (GMs) containing the antitumour aromatic tetra-amidine TAPP-Br and its release characteristics from microspheres are described. Spherical gelatin microspheres, showing a high percentage of drug encapsulation (over 95 per cent) and an average diameter of 16 microns were obtained by a coacervation method. In addition, the effects of various reaction parameters on the production and chemical hardening of GMs have been investigated. In order to compare the TAPP-Br release profiles from GMs prepared with different procedures, in vitro studies were carried out. The biological effect of TAPP-Br released from gelatin microspheres was determined on 'in vitro' cultured tumour cell lines, showing that, after encapsulation, TAPP-Br still retains antiproliferative activity comparable with that exhibited by the free drug. Taken together, the results obtained suggest that gelatin-based microspheres offer excellent potential as carriers for the controlled release of polyamidines in anticancer chemotherapy.

Antineoplastic Agents↗
PubMedDate not supplied

Mouse foetal development: translational inactivation of globin genes during the terminal differentiation of yolk sac erythroid cells.

The molecular mechanism of globin genes inactivation has been explored at the translational level in yolk sac erythroid cells from mouse foetuses. Ageing of the messenger molecules is paralleled by a progressive decrease of the poly(A) tail. Poly(A) less messengers are less active when tested in a wheat germ cell free system and become uncapable to terminate globin chain elongation.

Age Factors↗
PubMedDate not supplied

DNA elements target of transcriptional factors are not restricted to long terminal repeat of human immunodeficiency virus.

In this report we show that signals for transcriptional factors are not restricted to the HIV-1 LTR, but are present throughout the HIV-1 genome. Furthermore, we identified a sequence, AGAACAGATG, highly homologous to the X-box of class II MHC genes and located within the tat-IVS/env region of HIV-1. Double stranded oligonucleotides mimicking the HIV-1 region containing AGAACAGATG were synthesized and band shift experiments were performed demonstrating that this HIV-1 genomic region binds nuclear proteins. We further demonstrate that the binding of nuclear factors to this tat-IVS/env HIV-1 sequence is competed for, in the band-shift assay, by the highly homologous X-box of the promoter of the human HLA-DR alpha gene. The presence in the HIV-1 genome of DNA sequences homologous or identical to regulatory sequences of cellular genes represents a potential mechanism of predation of DNA elements recognized by DNA binding proteins.

Base Sequence↗
PubMedDate not supplied

Recombinant immune interferon down-regulates Ha-ras-1 proto-oncogene products in a human melanoma cell line.

The antiproliferative and antineoplastic effects of the interferons may result, at least in part, from changes in the expression and quantity of specific oncogene products. To explore this hypothesis we have determined the effect of interferons, including recombinant leukocyte (IFN-alpha), fibroblast (IFN-beta) and immune (IFN-gamma), on expression of the Ha-ras proto-oncogene in the human melanoma cell line Colo 38. While concentrations of up to 1000 U/ml of either IFN-alpha or IFN-beta did not affect the total amounts of Ha-ras products, IFN-gamma at concentrations ranging from 20 to 200 U/ml caused a dose- and time-dependent (48-96 hr) reduction (approximately 40%) in the accumulation of Ha-ras-1 mRNA and in the synthesis of the specific protein products. Downregulation of this proto-oncogene occurs prior to the antiproliferative effects of IFN-gamma and parallels similar IFN-gamma mediated changes in the expression of certain melanoma associated antigens. The present findings indicate that this experimental model may prove valuable in determining whether a direct relationship exists between the antiproliferative activity of specific interferons and the downregulation of oncogene expression.

Antibodies, Monoclonal↗
PubMedDate not supplied

Binding of nuclear factors to the IFN-gamma consensus sequence of the human HLA-DR alpha gene: effects of IFN-gamma on tumor cell lines.

The binding of nuclear proteins from human tumor cells to synthetic double stranded oligonucleotides mimicking upstream regions of the HLA-DR alpha gene was studied. As the HLA-DR alpha gene is inducible by interferon(IFN)-gamma, nuclear extracts were also purified from IFN-gamma treated cells. Our data indicate that a) nuclear binding proteins (named IFN-gamma-B3 and Z-B1/B2) are detectable, specific for the IFN-gamma and Z boxes of the human HLA-DR alpha gene; b) both IFN-gamma-B3 and Z-B1/B2 are present in HLA-D negative cell lines and c) the content of IFN-gamma-B3 and Z-B1/B2 is not modulated by IFN-gamma treatment. These data suggest that the presence in the nuclear compartment of these factors, presumably necessary for the correct regulation of the HLA-DR alpha gene, is not sufficient to explain its differential constitutive and induced expression in a variety of in vitro cultured cell lines of different lineage.

Base Sequence↗
PubMedDate not supplied

Effects of the proteinase inhibitor tetra-p-amidinophenoxyneopentane on in vitro adhesion and invasiveness of tumor cells.

Some of the biological processes which enhance the metastatic capacity of tumor cells are associated with the activation of proteinases. In this paper we examined the effects of aromatic polyamidines on "in vitro" invasiveness of tumor cell lines. In addition to their antiproteinase activity, aromatic polyamidines exhibit antiproliferative activity toward tumor cell lines and inhibit expression of specific oncogenes. The results obtained show that the aromatic polyamidine TAPP-Br does not affect attachment, but inhibits the "in vitro" invasiveness of tumor cells cultured on a reconstituted basement membrane composed by laminin, type IV collagen, entactin, heparan sulphate proteoglycans. Our data suggest that this and related compounds should be further studied as experimental antitumor agents.

Amidines↗
PubMedDate not supplied

Regulation of the antigenic phenotype of human melanoma cells by recombinant interferons.

The ability of interferons to modulate the antigenic phenotype of tumor cells may involve alterations in the transcription, translation, membrane expression and shedding of Major Histocompatibility Complex (MHC) and Tumor Associated Antigens (TAAs). In the present study we have investigated possible mechanisms by which recombinant human interferons, IFN-alpha, -beta and -gamma, alter the antigenic profile of long- and short-term human melanoma cultures. IFN-alpha and -beta induced similar changes in the synthesis, expression and shedding of two TAAs, a HMW-MAA and a Cyt-MAA, in the established melanoma cell line Colo 38, whereas IFN-gamma exerted a differential effect on these melanoma associated antigens. Moreover, IFN-gamma was relatively more effective than IFN-alpha or -beta in upregulating the synthesis, expression and shedding of class I MHC antigens. At the same time a dramatic differential effect of the interferons was observed with class II MHC antigens. IFN-alpha or -beta induced a modest increase in the synthesis and expression of these antigens, whereas IFN-gamma was greater than 3-fold more active in inducing the synthesis and expression of DR/DP antigens and greater than 4- and greater than 10-fold more effective in increasing the synthesis and expression, respectively, of DQ antigens. Analysis of the levels of cytoplasmic mRNA for the DR-alpha and DQ-beta genes indicated no significant difference between IFN-alpha, beta or -gamma treated cells suggesting that IFN-gamma enhancement of the synthesis of DR and DQ antigens may occur at a posttranscriptional level. In the case of a newly established human melanoma cell line (MG-3) IFN-gamma enhanced the synthesis but not the expression of DR antigens and did not alter either the synthesis or expression of DQ antigens. Our studies indicate that the effects of interferons on the antigenic phenotype of melanoma cells will vary depending on the type of interferon employed, the antigen monitored and the target cell studied. In addition, it is also apparent that some of the biosynthetic steps involved in regulating the synthesis, expression and shedding of antigens may be coordinately regulated in some melanoma cells, whereas these processes may be under independent control in other melanoma populations.

Antigens, Neoplasm↗
PubMedDate not supplied

Regulation of class II MHC gene expression by interferons: insights into the mechanism of action of interferon (review).

Modulation of class I and II MHC antigen expression by interferons has been the focus of considerable attention because the regulation of these molecules serves as a useful model system to study factors exerting transcriptional control of gene expression and because of the relevance of these molecules to expression of the neoplastic phenotype. While our knowledge of the molecular mechanisms regulating the ability of interferon to mediate enhancement of MHC genes has increased, this information is primarily based on studies employing established cell lines, and it remains to be determined whether similar controls are also exerted in short term cultured cell lines. In this short review, we have discussed the structural organization of the 5' flanking regions of the MHC genes, with special emphasis on class II genes, and the implications of these data for the transcriptional regulation of these and of other interferon inducible genes. Present evidence indicates the existence of at least four conserved upstream sequences which are shared by interferon responsive genes and which appear to be involved in the transcriptional control of these genes. The pattern of metabolic requirements for IFN-alpha and IFN-beta versus IFN-gamma upregulation of the class I and II MHC genes suggests that the regulation of gene expression by IFN-gamma requires unique regulatory molecules, i.e. newly synthesized proteins, which are not required by IFN-alpha and IFN-beta treated cells. These putative mediators of gene expression are likely to be shared by many of the biochemical induction pathways involved in the regulation of genes which exhibit interferon responsive sequences. However, in addition to common regulatory signals, other specific pathways and possibly additional regulatory sequences, are also required to account for locus--or subunit-specific patterns of antigenic modulation. Future studies are required, including those employing short term tumor cell cultures, to precisely define the molecular details of gene regulation of class I and II MHC genes, as well as other interferon--responsive--genes, by interferons. These investigations will not only prove valuable on a fundamental scientific level, but they will also be crucial for the more effective application of interferons in clinical oncology.

Gene Expression Regulation↗
PubMedDate not supplied

Antitumor activity of the proteinase inhibitor tetra-p-amidinophenoxyneopentane in a nude mouse model of human melanoma.

An aromatic poly-amidine (tetra-p-amidinophenoxyneopentane, TAPP-Br) exhibiting anti-proteinase activity and known to exert antitumor activity in vitro was analysed for its ability to inhibit the in vivo growth of a human melanoma cell line transplanted in nude mice. 5 X 10(6) melanoma cells were injected subcutaneously in groups of nude mice and treatment with TAPP-Br was performed (0.125-1 mg/0.2 ml injections, repeated three times at the beginning of the experiment and after 20 days). After 25 days tumors displaying a volume of 0.9-1.8 cm3 were detectable in control untreated mice. Mice treated with TAPP-Br on the other hand did not develop sizable tumors or consistently developed tumors of significantly smaller sizes. Despite these therapeutic effects, significant chronic toxicity of the compound was observed when administered at higher dosage (500-1000 micrograms). These side effects, which may hamper the therapeutic use of TAPP-Br, are likely to be circumvented by alternative routes of administration or by vehiculation into liposomes. These alternative strategies of treatment are currently investigated.

Animals↗
PubMedDate not supplied

New synthetic retinoids: effects on proliferation and differentiation.

Nine retinoid analogues were synthesized and their effects on cell proliferation and differentiation of tumor cell lines were analysed and compared with those of natural retinoids (retinoic acid, retinol, retinal). Our results demonstrate differential inhibitory effects of the synthetic retinoids on cell growth. This inhibitory activity of the synthetic retinoids was, in some cases, higher than that of retinoic acid and retinol. Natural and synthetic retinoids were found to be able to induce to a different extent erythroid differentiation of murine erythroleukemia cells and adipogenic conversion of Chinese Hamster FHO6N1-1 cells. Our data suggest that studies on the relationship between structure and biological activity could be approached by using the analysed synthetic retinoids.

Animals↗
PubMedDate not supplied

DNA methylation of the Ha-ras-1 oncogene in neoplastic cells.

The DNA methylation pattern of the human Ha-ras-1 oncogene and the levels of specific mRNA have been analysed in established tumor cell lines and in surgical biopsies. In long-term cultures, the Ha-ras-1 gene is hypomethylated at its 5' end and highly methylated at its 3' portion. The GCGC sites at the 5' end have been shown to become methylated only when E. Coli HhaI methylase is added to isolated DNA, while they remain unmethylated when the enzyme is added to chromatin. These data indicate that the 5' portion of this protooncogene is maintained physiologically unmethylated, possibly because it is necessary for gene transcription. Along the same line of evidence, when levels of mRNA and methylation of the gene were comparatively analysed in breast tumors and autologous normal mammary tissue, no appreciable differences in the degree of methylation were found despite significant differences in mRNA content. These findings show that the levels of Ha-ras- mRNA are independent of the DNA methylation state.

Breast Neoplasms↗
PubMedDate not supplied

Effects of benzamidine derivatives on Ha-ras-1 mRNA accumulation in a Chinese hamster cell line transformed with the activated human T24 Ha-ras-1 oncogene.

Tetra benzamidine derivatives were found to inhibit proteinase activity, cell proliferation and accumulation of the Ha-ras-1 mRNA in the FHO6T1-1 Chinese Hamster cell line, transformed with the activated human T24-Ha-ras-1 oncogene. Di- and Tri-benzamidine derivatives were also found to be potent inhibitors of proliferation of FHO6T1-1 cells. These latter compounds could be proposed as useful substrates in the synthesis of drug-conjugated monoclonal antibodies or growth factors.

Amidines↗