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Biomedical subjects

U Consoli

Publications and source records attributed to U Consoli.

32 records · Page 2Linked to original sources
PubMed1996

Flt3 ligand stimulates proliferation and inhibits apoptosis of acute myeloid leukemia cells: regulation of Bcl-2 and Bax.

Flt3/flk-2 ligand (flt3-L) is a potent costimulator of normal bone marrow (BM) myeloid progenitors. Flt3-L is produced by BM stromal cells and its receptor is expressed in the majority of acute myeloid leukemia (AML) cases. Therefore, flt3-L may play a role in the paracrine and/or autocrine loops sustaining leukemic cell growth. We evaluated the effects of recombinant human flt3-L on proliferation, apoptosis, and Bcl-2 and Bax expression in primary AML cells and compared them with those of stem cell factor (SCF). Mononuclear BM cells from patients with newly diagnosed AML were cultured in serum-free conditions with flt3-L, SCF, granulocyte colony-stimulating factor (G-CSF) and granulocyte macrophage-colony-stimulating factor (GM-CSF) alone and in combination. In 9 of 10 samples, flt3-L significantly increased [3H]thymidine uptake (geometric mean stimulation index, 7.5; range, 2.4 to 41.5). Flt3-L also increased the number of AML blast colonies by 126% (range, 61% to 181%). In these 9 samples, flt3-L significantly enhanced the proliferative response triggered by G-CSF or GM-CSF. Flt3-L prevented apoptosis in AML blasts. It reduced the number of apoptotic cells by 36% +/- 3.9% compared with control cultures. Combining flt3-L with G-CSF or GM-CSF doubled the antiapoptotic effect. Cellular Bcl-2 and Bax levels were determined separately for apoptotic and nonapoptotic cells by flow cytometry. Cells undergoing spontaneous apoptosis had low Bcl-2 and high Bax levels, whereas nonapoptotic cells had high Bcl-2 and low Bax levels. Flt3-L alone or in combination with G-CSF or GM-CSF did not upregulate Bcl-2. However, Bax expression decreased in viable cells in the presence of these cytokines and the lowest level was achieved when a combination of flt3 and GM-CSF was used. Proliferative and viability effects of flt3-L were similar to those of SCF. Our results demonstrate that flt3-L acts as a stimulatory factor for primary AML cells. The antiapoptotic effects of flt3-L or its combinations with G-CSF or GM-CSF correlate with their ability to prevent upregulation of Bax.

Acute Disease↗
PubMed1996

The novel anthracycline annamycin is not affected by P-glycoprotein-related multidrug resistance: comparison with idarubicin and doxorubicin in HL-60 leukemia cell lines.

A major factor in limiting the efficacy of anthracyclines is overexpression of the MDR1-encoded p-glycoprotein (p-gp). A new analogue less affected by p-gp is annamycin (ANN), an anthracycline antibiotic with high affinity for lipid membranes and significantly more activity than doxorubicin (DOX). We investigated whether ANN was affected by p-gp-mediated multidrug resistance (MDR) by comparing the cellular accumulation and retention of ANN, idarubicin (IDR), and DOX in the p-gp-negative human leukemia cell lines (HL-60S) and its DOX-selected p-gp-positive subline (HL-60/DOX) with and without verapamil (VER). As expected, HL-60/DOX cells showed lower DOX uptake than HL-60S cells; coincubation with VER (10 mmol/L) increased uptake 2.6-fold restoring it to 100% of uptake in HL-60S cells. IDR uptake increased 1.5-fold in the presence of VER, but ANN was not affected. Coincubation with VER increased DOX retention in HL-60/DOX cells 2.8-fold and IDR retention 1.4-fold; unchanged ANN retention indicated that ANN may overcome p-gp. In the cytotoxicity assay to correlate intracellular anthracycline content with antitumor activity, we found ANN to be less potent than DOX and IDR In sensitive cells, ID 50 being the drug concentration that inhibits cell growth by 50% but its resistance index (RI; ID50 resistant cells divided by ID50 sensitive cells) was lower than that of IDR and DOX (2.6 v 40 and 117.5). Coincubation in the presence of VER resulted in 4.5-fold and 2-fold RI decreases of DOX and IDR, respectively, whereas ANN did not change, further confirming ANN's ability to circumvent p-gp-mediated MDR. Confocal microscopy studies of IDR, ANN, and DOX showed higher intracellular drug compartmentalization for DOX in HL-60/DOX cells incubated in the presence of VER. This study provided evidence that, unlike DOX and IDR, ANN is not affected by p-gp-mediated MDR.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

A pilot study of high-dose interleukin-3 treatment of relapsed follicular small cleaved-cell lymphoma: hematologic, immunologic, and clinical results.

The growth stimulatory effects of interleukin-3 (IL-3) on normal hematopoietic progenitor cells are well established, and clinical trials using IL-3 after bone marrow transplantation for various malignancies including lymphomas are frequently conducted. Although the IL-3 receptor is expressed on the surfaces of follicular small cleaved-cell lymphoma (FSCCL) cells, the in vivo effects of IL-3 on FSCCL have not been studied previously. Because our preclinical data suggested that IL-3 may have dose-dependent inhibitory effects on FSCCL cells in vitro, we treated eight FSCCL patients with high-dose IL-3 in an outpatient setting. Each patient received 1 mg/m2 of IL-3 subcutaneously daily for 14 days followed by 7 days without IL-3. After three courses (9 weeks), the patients were evaluated for clinical responses. One patient had a minor response, and four had no responses. Three patients who had progressive disease before IL-3 treatment continued to have progressive disease. In two patients with bone marrow involvement with lymphoma, IL-3 had no effect on FSCCL cells. One patient with peripheral blood involvement with FSCCL cells that expressed IL-3 receptors had temporary growth arrest of the circulating malignant cells. IL-3 significantly increased the absolute neutrophil count in seven patients (87%) but had little effect on the number of normal circulating B cells. There was an increase in the number of circulating natural killer cells and CD8+ cells in four patients. Treatment was very well tolerated; no life-threatening toxicities were observed. The most common toxicities were injected conjunctivae (100%), fever (100%), fatigue (87%), and skin rash (75%). Most of the side effects subsided with the continued use of IL-3. These preliminary results suggest that high-dose IL-3 does not stimulate the growth of FSCCL cells in vivo and, in some instances, may cause growth inhibition.

Aged↗
PubMed1996

CD30 ligand is expressed on resting normal and malignant human B lymphocytes.

CD30, a member of the tumour necrosis receptor superfamily, is physiologically expressed on a subpopulation of T helper cells in normal individuals but is also expressed on several malignant and virally transformed cells. Its ligand (CD30L) is a pleiotropic cellular transmembrane protein that can induce cell death in several CD30+ cell lines. CD30L expression has been reported on activated human peripheral blood T lymphocytes and macrophages but not on B cells. Here we show that the CD30L is expressed on resting normal and on malignant B cells in addition to both CD4+ and CD8+ subsets of activated T cells, making it the second tumour necrosis family member, in addition to the CD27 ligand, that can be expressed on both T and B cells. These findings raise the possibility that the CD30L has a role in B-cell/T-cell communication and that B and T cells are likely to be involved in the growth regulation of CD30+ tumours.

B-Lymphocytes↗
PubMed1996

Semi-automated PCR method for quantitating MDR1 expression.

The MDR1 gene is involved in drug resistance in many hematopoietic and solid tumors. The Quantitative PCR System 5000 (QPCR-5000; Perkin-Elmer) is a new instrument system that uses electrochemiluminescence to automatically quantitate polymerase chain reaction (PCR) products. A comparative study between radioactively labeled PCR (32P-PCR) and QPCR was performed to analyze the MDR1 gene expression in the drug-resistant (Doxorubicin) cell lines Dox40, Dox6, the parental cell line 8226/S, CEM Dox1 and three acute myeloid leukemia (AML) patient samples. Using the Dox40 and Dox6 resistant cell lines, we compared the sensitivities of QPCR and 32P-PCR. A strong signal was obtained from QPCR at 20 to 25 cycles (which is in the linear range for quantitation), while a weak signal was obtained using 32P-PCR at the same cycle number. Dilution experiments gave better precision with the QPCR than with the radioactive method. AML samples were studied with the MDR1-specific MAbs MRK16 and 4E3, and the efflux function was analyzed using Rh-123 retention in the absence or presence of verapamil. The three samples showed high (D = 0.79), medium (D = 0.52) and negative (D = 0.08) p-glycoprotein (P-gp) levels and correlated with efflux function. The MDR1/beta 2-M mRNA ratios for 32P-PCR were 0.41, 0.40 and 0.12, respectively, and were 0.127, 0.097 and 0.028, respectively, for QPCR. There were significant differences between the samples with high and medium P-gp levels comparing the two methods. Very low levels of MDR1 in CEM Dox1 cells could be detected only by QPCR. In conclusion, QPCR was found to be more reproducible, accurate and sensitive than 32P-PCR.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

Fludarabine-mediated inhibition of nucleotide excision repair induces apoptosis in quiescent human lymphocytes.

Incorporation of fludarabine, 9-beta-d-arabinofuranosyl-2-fluoroadenine (F-ara-A), into replicating DNA inhibits further chain elongation and is the critical event in F-ara-A-mediated cytotoxicity. We have used the normal cellular process of nucleotide excision repair to create an opportunity for F-ara-A incorporation into the DNA of noncycling cells. Irradiation of quiescent lymphocytes with UV light (254 nm, 0. 5-30 J/m2) in the presence of [3H]F-ara-A produced a dose-dependent increase in F-ara-A monophosphate incorporation into DNA that reflected a 60-70% inhibition of DNA repair at 2 h. Lymphocytes pretreated with 3 micrometer F-ara-A for 2 h before irradiation with 0.5 or 2.0 J/m2 were incubated for 24 h in the presence or absence of F-ara-A. Morphological features of apoptosis and DNA cleavage into high molecular weight fragments were increased in cells treated with UV plus F-ara-A compared to those treated with UV or F-ara-A alone. FACScan analysis confirmed the morphological and biochemical results. A 2-h pulse of F-ara-A produced intracellular F-ara-ATP levels of 40 micrometer, and removal of F-ara-A from the media resulted in a monophasic elimination (r2 = 0.88) in F-ara-ATP levels with a half-life of 5.6 h. Lymphocytes undergoing apoptosis demonstrated a G0 DNA content, indicating that entry into the cell cycle was not required. This study demonstrates that F-ara-AMP is incorporated into DNA during UV-induced repair in quiescent lymphocytes and that this is associated with the inhibition of ongoing DNA repair and an increased incidence of apoptosis. Combinational therapies involving fludarabine with agents and modalities that initiate DNA repair may have clinical relevance in the treatment of human malignancies.

Antineoplastic Agents↗
PubMed1995

Activation of p21WAF1/Cip1 expression by a temperature-sensitive mutant of human p53 does not lead to apoptosis.

The cyclin-dependent kinase inhibitor p21WAF1/Cip1 (p21) inhibits cellular proliferation and has been implicated in p53-dependent apoptosis. In this study, we examined the regulation of p21 in the K562 erythroleukemia cell line using a human temperature-sensitive mutant of p53, 143Ala. We showed that 143Ala at the permissive temperature (32 degrees C) activated the expression of p21. Stable cell lines expressing 143Ala or other p53 mutants were established. We then examined whether elevation of p21 promotes apoptosis and sensitized cells to radiation-induced apoptosis. Our results showed that p21 elevation did not promote or sensitize K562 cells to apoptosis.

Apoptosis↗
PubMed1995

PML/RARalpha, a fusion protein in acute promyelocytic leukemia, prevents growth factor withdrawal-induced apoptosis in TF-1 cells.

A unique mRNA produced by the t(15;17) (q22-24;q11-21) translocation in the leukemic cells of acute promyelocytic leukemia patients encodes a chimeric protein, PML/RARalpha, which is formed by the fusion of the retinoic acid receptor alpha (RARalpha) and the promyelocytic locus gene (PML). This translocation is often the only visible karyotypic aberration present which is detected in almost 100% of acute promyelocytic leukemia patients. As an initial step to study the role of PML/RARalpha in leukemogenesis, we attempted to express the fusion protein in hematopoietic cells through retrovirus-mediated gene transfer of the retroviral vector, pGPRCHT, which contains the PML/RARalpha cDNA. Transduction of the PML/RARalpha cDNA fragment used in this vector, which extends from the position 31 bp to the position 2638 bp in a transcription unit driven by the Moloney murine sarcoma virus LTR, was found to abrogate the growth factor dependence of TF-1 cells. In addition, introduction of PML/RARalpha into TF-1 cells can protect these cells from the apoptosis usually induced in TF-1 cells by growth factor withdrawal, as measured by three assays for apoptosis: morphology, DNA ladder formation, and end labeling of nicked DNA with fluorescent-conjugated nucleotide precursors followed by a fluorescence-activated cell sorting assay. These data suggest that the PML/RARalpha fusion protein may inhibit programmed cell death in myeloid cells.

Apoptosis↗
PubMed1991

Enhanced percentage of Leu M3+DR+ and Leu M3+CD25+ cells in newly diagnosed IDDM patients.

The percentage of Leu M3+DR+ and of Leu M3+CD25+ cells was determined by means of immunofluorescence analysis in a group of patients with insulin dependent diabetes mellitus (IDDM). Our results show that an increased percentage of these cells may occur in the early stage of the disease. These data provide evidence for a "phenotypical" activation of Leu M3+ cells at the onset of the disease and warrant future studies to evaluate the potential role of these cells in the pathogenesis of IDDM.

Adolescent↗
PubMed1990

Enhanced percentage of CD5+ B lymphocytes in newly diagnosed IDDM patients.

The percentage of CD5+ B lymphocytes, the prevalence of islet cell antibodies (ICA) and of anti-insulin autoantibodies (IAA) and HLA-A-B-C and DR antigens were studied in 32 newly diagnosed insulin-dependent diabetes mellitus (IDDM) patients, in 12 non-insulin-dependent diabetes mellitus (NIDDM) patients and in 12 healthy subjects. The percentage of CD5+ B lymphocytes ranged from 18% to 51.2% (mean 40.3 +/- 11%) in IDDM patients, whereas in NIDDM patients and in controls it ranged from 20% to 25.2%, (mean 21.3 +/- 4.1%) and from 16% to 24%, (mean 19.3 +/- 1.9%), respectively (P less than 0.01 vs. NIDDM patients and vs. controls). There was no correlation between a higher percentage of CD5+ B lymphocytes and the presence of ICA and/or IAA, and their titres, and/or of any HLA-A-B-C and DR antigens. Thus, an enhanced percentage of CD5+ B lymphocytes may be present in newly diagnosed IDDM patients; the possible role of this cell type in the pathogenesis of IDDM needs further investigation.

Adolescent↗
PubMedDate not supplied

Inhibition of lung cancer proliferation by antisense cyclin D.

The growth and tumorigenicity of murine lung cancer cells transfected with an antisense cyclin D1 construct were evaluated in studies pertaining to mouse lung carcinogenesis. This antisense construct inhibited the expression of cyclin D in these cells, significantly reducing both their in vitro proliferation and tumorigenicity in nude mice relative to control cells. These data may have implications regarding the treatment of human neoplasms of aerodigestive tract origin that either overexpress the cyclin D oncogene or exhibit mutations that influence cell cycle progression via cyclin D-dependent mechanisms.

Animals↗