Biomedical subjects

M L Larramendy

Publications and source records attributed to M L Larramendy.

At least 55 records · Page 3Linked to original sources

PubMed1994

Cell lineage involvement of recurrent chromosomal abnormalities in hematologic neoplasms.

Analysis of most hematologic neoplasms indicates the involvement of one or more cell lineages in the bone marrow and/or the blood but rules out the involvement of all lineages in any one neoplasm. It is important to detect lineage involvement in order to clarify which stem cells are involved in leukemia, to predict prognosis, and to select appropriate treatment. Our aim was to study the cell lineage involvement of some of the recurrent chromosomal abnormalities seen in hematological neoplasms. The direct morphology-antibody-chromosomes (MAC) method was used. The deletion 20q in myeloproliferative diseases (MPD), the deletion of 5q and t(1;7) in myelodysplastic syndromes (MDS), and t(3;3) in acute myeloid leukemia subtype M7 (AML-M7) were seen in all or at least in two myeloid lineages. These were interpreted as stem cell abnormalities. Deletion 13q in MPD, t(8;21) in AML-M2 and t(15;17) in AML-M3 were seen in granulocytic lineages only; t(14;18) in non-Hodgkin's lymphoma and trisomy 12 as the sole abnormality in chronic lymphocytic leukemia (B-CLL) were seen only in immunoglobulin light chain clonal B cells; inversion 14 in T-CLL was seen only in T cells, whereas t(15;14) in acute lymphocytic leukemia with eosinophilia (ALL-EO) was seen in lymphoid stem cells but not in mature granulocytes or lymphocytes. Additional abnormalities (in addition to the Philadelphia chromosome) in chronic myeloid leukemia (CML) were seen in all myeloid cell lineages and also in mature granulocytes, B cells, and large granular lymphocytes. Abnormalities in Hodgkin's disease were restricted to CD30-positive Reed-Sternberg cells. Trisomy 8 and monosomy 7 are abnormalities that may be present in either stem cells or any of the single cell lineages.

Blast Crisis↗

PubMed1994

Absence of trisomy 7 in nonneoplastic human ascitic and pleural fluid cells. An interphase cytogenetic study.

Trisomy 7 is a frequent aneuploid change in lymphomas, adenocarcinomas, and malignant mesenchymal and neurogenic tumors. Moreover, it has been observed in cultured and uncultured non-neoplastic cells from brain, kidney, liver, lung, and atherosclerotic plaques, among other tissues, opening debate on the role of this change in normal and neoplastic tissue. We used nonradioactive in situ hybridization (ISH) with a biotinylated chromosome 7-specific alpha-satellite DNA probe to seek an extra copy of chromosome 7 in ascitic and pleural fluid interphase cells from 26 donors. The donors comprised 24 patients with nonmalignant clinical history, one patient with non-Hodgkin's malignant lymphoma (positive control), and one patient with chronic myeloid leukemia (CML, negative control). The highest frequency of fluid cells with three hybridization signals in patients without neoplasia was 0.5%, in contrast to the frequency of 40.5% noted in the fluid cells of the patient with non-Hodgkin's malignant lymphoma. The results demonstrate that the frequency of trisomic cells in pleural as well as in ascitic fluid is very low, making possible use of the cells in ascitic or pleural fluids in identification of malignancy.

Adult↗

PubMed1994

Analysis of genotype and phenotype on the same interphase or mitotic cell. A manual of MAC (morphology antibody chromosomes) methodology.

The purpose of this paper is to serve as a MAC (Morphology Antibody Chromosome) manual describing combined methodologies that allow simultaneous and/or sequential analysis of cell morphology, immunophenotype, and banded chromosomes and/or in situ hybridization signals. The MAC techniques used at the Department of Medical Genetics of the University of Helsinki, Finland, are described and modifications or related techniques reported by other authors are discussed. A list of references concerning applications is also given.

Chromosome Banding↗

PubMed1994

Ploidy in bone marrow cells from healthy donors: a MAC (morphology antibody chromosomes) study.

The ploidy of human bone marrow cells belonging to the megakaryocytic, granulocytic-monocytic and erythrocytic lineages was studied by in situ hybridization using the biotin-labelled Y chromosome-specific DNA probe pY431 and the chromosome 1-specific probe pUC1.77 on cells identified morphologically and immunologically by the MAC procedure. Cells of the granulocytic-monocytic and erythrocytic lineages were seen to be 2N in ploidy, whereas the ploidy of the megakaryocytic lineage ranged from 2N to 32N, with the ploidy classes 4N and 8N being predominant. The frequency of megakaryocytes with 2N chromosomes was also high.

Adult↗

PubMed1994

Lineage involvement and karyotype in a patient with myelodysplasia and blood basophilia.

We report a 63-year-old woman with myelodysplastic syndrome (MDS), refractory anaemia with ring sideroblasts (RARS), and blood basophilia with pathological forms. Karyotype analysis revealed a complex rearrangement: 46,XX,del(3)(p13p25),del(5)(q13q33),der(16)t(1;16) (p13;q12)/47,idem,add(20)(?p11)/49,idem, + add(1)(q32),add(20)(?p11), + mar/46,XX. Karyotype, immunophenotype and in situ hybridization studies by the MAC (morphology antibody chromosomes) combination technique revealed the chromosomal abnormality in granulocytic/monocytic and erythrocytic metaphase cells. Also mature basophils and other granulocytes were involved with the abnormality. We suggest that our patient with the MDS has a stem cell disorder affecting all myeloid cell lineages and that basophilia constitutes a part of the malignant process.

Basophils↗

PubMed1994

Detection of minimal residual disease using fluorescence DNA in situ hybridization: a follow-up study in leukemia and lymphoma patients.

We used fluorescence DNA in situ hybridization (FISH) to detect chromosomal abnormalities as an indicator of minimal residual disease in follow-up samples from the bone marrow (BM), or peripheral blood, of 25 patients with leukemia, lymphoma and myelodysplastic syndromes. Trisomies were detected by interphase FISH with repeat-sequence probes (RSP) or by using metaphase FISH with whole-chromosome paint probes (WCP). Specific translocations were detected using WCP probes. Translocations were observed using metaphase FISH in two patients in uncertain or complete remission (CR), who both later suffered relapse. Five patients with no abnormal cells remained in CR. Four patients with trisomies detected during CR suffered relapse; metaphase FISH detected the trisomy in 0.17-16% of metaphase cells. Five patients for whom the trisomy occurred in 0.034% of cells remained in CR. Trisomic nuclei were observed in 0.27-2.3% of interphase cells, by means of RSPs, in four patients who later suffered relapse. Five patients with trisomic nuclei in 0.061% remained in CR. When two probes were used simultaneously in a sample from one patient, 1% of the residual cells were abnormal. The patient later suffered relapse. In one patient with anaplastic large cell lymphoma, CD30-positive interphase cells were shown to have trisomic chromosome 7 by immunophenotyping and FISH. Our results suggest that metaphase FISH using WCP probes is a sensitive and specific method for detecting minimal residual disease especially in patients with translocations.

Acute Disease↗

PubMed1993

Cell lineage involvement in four patients with myelodysplastic syndrome and t(1;7) or trisomy 8 studied by simultaneous immunophenotyping and fluorescence in situ hybridization.

Four patients with myelodysplastic syndrome (MDS), one with t(1;7) and three with trisomy 8, were studied by immunophenotyping and fluorescence in situ hybridization (FISH) to assess cell lineage involvement. The t(1;7) was detected using a biotin-labeled chromosome 1 centromere-specific DNA probe. This aberration was present in CD34-positive stem cells, the erythroid cell lineage (GPA+), and the granulocytic/monocytic (CD13+ and CD64+) cell lineages. We were not able to demonstrate the abnormality in the lymphoid cell lineages. In the patients with trisomy 8, the aberration was detected with chromosome 8 centromere-specific DNA probe or by chromosome in situ suppression hybridization (CISS) with a chromosome 8-specific library probe. The trisomy was detected in stem cells, erythroid precursor cells, megakaryocytes, and granulocytes/monocytes. In these MDS patients, the chromosome aberrations appear to occur only in cells of myeloid lineage.

Aged↗

PubMed1993

Pig plasma modulates cell cycle kinetics but not the baseline frequency of sister-chromatid exchanges in human lymphocytes.

The effect of human and pig plasma on the baseline frequency of sister-chromatid exchanges (SCEs) in human and porcine plasma leukocyte cultures was studied. Human plasma leukocyte cultures, but not porcine plasma leukocyte cultures, showed at least a twofold increase in SCE frequency over whole blood culture values. Addition of pig plasma to human plasma leukocyte cultures and human plasma to pig plasma leukocyte cultures did not modify the baseline SCE frequencies. In both human and porcine cultures, cell proliferation was slower in plasma leukocyte cultures than in whole blood cultures. The addition of pig plasma to human plasma leukocyte cultures, but not the incorporation of human plasma in pig plasma leukocyte cultures, accelerated the cell cycle progression of lymphocytes. With 10% pig plasma in the plasma leukocyte culture medium, lymphocyte proliferation was similar to that in whole blood cultures. Smaller concentrations of pig plasma rendered cell cycle progression intermediate between the basal plasma leukocyte culture and whole blood culture values. Exchanging fetal calf serum for human AB serum in human plasma leukocyte cultures did not affect the cell cycle kinetics of lymphocytes but it did decrease their baseline SCE frequency.

ABO Blood-Group System↗

PubMed1993

Simultaneous detection of high-resolution R-banding and fluorescence in situ hybridization signals after fluorouracil-induced cellular synchronization.

A method for simultaneous detection of fluorescence in situ hybridization of DNA probes and high resolution fluorescent R banding is described. Human lymphocytes were stimulated with phytohemagglutinin and synchronized using a fluorouracil block followed by exposure to bromodeoxyuridine and Hoechst 33258 prior to harvest. Metaphase preparations were treated with Hoechst 33258 and exposed to UV light. Thereafter they were incubated in sodium phosphate buffer and dried prior to in situ hybridization with a biotin-labelled centromere-specific alpha-satellite DNA probe for chromosome 1 (pUCl.77) and two digoxigenin-labelled probes, i.e., a PCR-generated chromosome 8-specific alphoid probe (#8) and a cosmid probe for FLT4 gene on 5q33-qter (class III receptor tyrosine kinase). Hybridization signals were detected by an indirect immunofluorescence method using fluorescein isothiocyanate. The chromosomes were counterstained with propidium iodide and 4',6-diamidino-2-phenylindole dihydrochloride. This simple method allows unambiguous chromosome band identification simultaneously with detection of the hybridized probes.

Cell Division↗

PubMed1992

Chromosomal sensitivity of human lymphocytes to bleomycin. Influence of antioxidant enzyme activities in whole blood and different blood fractions.

The activity of the antioxidant enzymes catalase (CAT), peroxidases (POD), and superoxide dismutases (SOD) in whole blood and different blood fractions was analyzed in 20 normal human beings and correlated with the chromosomal sensitivity of lymphocytes to bleomycin (BLM) (measured as frequency of dicentric chromosomes per BLM dose). Our results demonstrate that both the physiologic activities of the enzymes and the chromosomal sensitivity to BLM exhibit an ample and significant interindividual variability. An inverse and linear correlation between chromosomal sensitivity to BLM and the concentration of 1) CAT and POD in plasma and 2) SOD in whole blood, erythrocytes, and plasma was found. On the other hand, the chromosomal sensitivity to BLM showed a direct correlation with the concentration of SOD and POD in mononuclear leukocytes. It is suggested that a determination of antioxidant enzyme (AOE) activities in a given cell population may serve to predict the chromosomal sensitivity to BLM.

Adult↗

PubMed1991

Increased frequency of micronuclei in B and T8 lymphocytes from smokers.

The frequency of micronuclei was measured in peripheral B lymphocytes and some T lymphocyte subpopulations from 5 medium-tar cigarette smokers, and 5 non-smokers with no regular exposure to tobacco smoke. The peripheral lymphocytes were stimulated in vitro with phytohemagglutinin and B lymphocytes and the various T lymphocyte subsets were classified by a recently developed MAC (Morphology, Antibody, Chromosomes) method which allows the immunologic identification of different cell lineages. An increased frequency of micronuclei was observed in B and especially in the suppressor/cytotoxic T8 lymphocytes from smokers, as compared with non-smoker values. In non-smoker cultures, no differences in the frequency of micronuclei were observed among the different T lymphocyte subsets. In these cultures, B cells tended to have a higher frequency of micronuclei than did T cells. The proportions of B cells and the various T subpopulations among mitotic and interphasic lymphocytes from smokers and non-smokers were also determined. The proportions of B cells and T cell subsets among all mitotic lymphocytes were similar in smokers and non-smokers. Contrarily, a significant decrease in the proportion of T8 lymphocytes among all interphasic lymphocytes was observed in cultures derived from smokers.

Adult↗

PubMed1990

Immunophenotype and sister chromatid differentiation: a combined methodology for analyzing cell proliferation in unfractionated lymphocyte cultures.

Combination of the MAC (morphology, antibody, chromosomes) and harlequin staining procedures offers a method for direct analysis of cell kinetics in cultures of unfractionated hematopoietic cells. In the present study unfractionated human mononuclear leukocyte cultures were stimulated with PHA or PWM mitogens and exposed to bromodeoxyuridine for various periods. For MAC, cytospin preparations were made and cells were classified with monoclonal B and T antibodies by the immunoperoxidase technique. After differentiation of the different lymphocyte subsets, the cells were stained by a fluorescence-plus-Giemsa method to distinguish sister chromatids and to determine the proportions of first, second, third, or subsequent mitoses among the previously identified subsets. The results showed (1) that the relative proportions of mitotic T and B cells are the same regardless of the mitogen used; (2) that T and B lymphocytes proliferate faster in cultures stimulated by PWM than in those stimulated by PHA; and (3) that T cells enter mitosis earlier than B cells when PHA or PWM are used as mitogens.

Adult↗

PubMed1990

Erythrocytes modulate the baseline frequency of sister-chromatid exchanges and the kinetics of lymphocyte division in culture.

The baseline sister-chromatid exchange (SCE) frequencies of human plasma lymphocyte cultures (PLC), but not pig PLC, were nearly twice as high as those of whole-blood cultures (WBC). Addition of human red blood cells (RBCs) to human PLC decreased the SCE frequency in proportion to the RBC-leukocyte co-incubation interval. When the period of RBC-leukocyte co-incubation was equivalent to the total length of the culture period (72 h), the SCE frequency was similar to that observed in WBC. Shorter co-incubation periods yielded SCE frequencies intermediate between those of PLC and WBC. Regardless of the species, cell proliferation was slower in PLC than in WBC. Experiments where RBCs were added to PLC showed that the time sequence of RBC incorporation also affects the cell-cycle progression of human and pig lymphocytes. When either human or pig RBCs were added immediately after PLC stimulation, the cell-cycle kinetics was similar to that of WBC. Shorter co-incubation periods made cell-cycle progression intermediate between PLC and WBC values. Thus, PBCs modulate the baseline frequency of SCEs in human PLC and the cell-cycle progression of both human and pig lymphocytes in a time-dependent manner. Two possible hypotheses for the heightened frequency of SCEs of human lymphocytes in RBC-free cultures were assessed. The loss of RBC-to-lymphocyte cellular contact in PLC did not influence the SCE frequencies of lymphocytes. Finally, the increase of SCEs in human PLC could not be related to differences in the generation time of lymphocytes in culture.

Animals↗

PubMed1990

Chromosomal abnormality limited to T4 lymphocytes in a patient with T-cell chronic lymphocytic leukaemia.

Cytogenetic, immunologic and DNA studies were performed on a patient with the T-cell type of chronic lymphocytic leukaemia. Standard G-banding karyotype analysis revealed clonal chromosome abnormalities with the karyotype 42-44,XY,-11,-12,-13,-16,-21,-22,i(8q),inv(14) (q11q32), t(15;?)(p11;?), +4-6mar/43-44,X,t(Y;14)(p11;q11),-11,-12, -13,-21,-22,i(6p),i(8q),inv(14)(q11q32),t(15;?)(q11;?), +1-3mar. MAC (morphology, antibody, chromosomes) methodology, which allows the immunophenotyping of mitotic cells, showed that the chromosome abnormalities were restricted to CD4-positive helper/inducer T lymphocytes and that CD8 suppressor/cytotoxic T cells and B lymphocytes possessed a normal karyotype. The results also indicate that the proportion of abnormal metaphases and the overall mitotic activity after 3.5 days of stimulation in vitro were highest when PHA and TPA were used as mitogens. When the culture period in the presence of PHA + IL-2 was extended, the proportion of the abnormal cell population decreased in direct relation to the length of the culture period, ranging from 100% at 3.5 d to 0% at 31 d after stimulation. Southern blotting analysis revealed rearrangements of both the immunoglobulin heavy chain gene and the beta T-cell receptor gene.

Aged↗

PubMed1989

Effect of the metal chelating agent o-phenanthroline on the DNA and chromosome damage induced by bleomycin in Chinese hamster ovary cells.

A 30-min pulse treatment with bleomycin to Chinese hamster ovary cells in culture produces DNA degradation and chromosomal aberrations in a dose-dependent manner. Bleomycin also induces a long-lasting effect on the cell cycle producing a lengthening of two or more cycles after the treatment. The presence of o-phenanthroline, which chelates metal ions, totally inhibits DNA cleavage and the appearance of chromosome aberrations while partially correcting the lengthening of the cell cycle. These findings suggest that an important cellular target for bleomycin is the DNA. Chromosomal aberrations are a secondary effect resulting from DNA cleavage. On the other hand, the increase in the duration of the cell cycle is probably induced by DNA degradation and, perhaps, by damage to other cellular structures.

Animals↗

PubMed1989

Ag-NOR staining and in situ hybridization of rDNA in the chromosomes of the South American camelids.

The location and frequency of Ag-stained NORs and sites of rDNA hybridization were studied in the chromosomes of the South American camelids. In the four camelids these regions occur distally on chromosomes 18, 21, and 27 and the smallest biarmed elements. Quantitative analysis of NOR distribution showed variations between both cells and species. In llama, guanaco and alpaca the NORs number averaged 6 per cell, this being higher than in vicuña where the average was 3. Relative frequencies of NOR-bearing chromosomes in the four camelids were similar. Yet, in vicuña virtual absence of NOR sites on one of the smallest biarmed pairs was observed. The rDNA sites assessed in llama and vicuña by in situ hybridization with cloned 18S DNA were coincident with the NOR locations and with the frequencies characteristics for each species. Moreover, varying the exposure time of the autoradiographs, labeling patterns specific for each camelid were observed. Grain counts on individual chromosomes indicated that under our conditions one month exposure is enough to demonstrate all the rDNA sites available in the complement of llama. Conversely, at least two months are necessary to show the total sites existing in vicuña. Most probably this finding reflects the presence of variations in the amount of copies of the ribosomal genes per chromosome.

Animals↗

PubMed1989

Correlation between the anti-oxidant enzyme activities of blood fractions and the yield of bleomycin-induced chromosome damage.

The concentration of the anti-oxidant enzymes catalase (CAT), peroxidases (POD) and superoxide dismutases (SOD) in different blood fractions, and the chromosomal sensitivity of lymphocytes to bleomycin-induced free radicals (expressed as frequency of dicentrics per bleomycin dose) were analyzed in 10 normal human donors. Our results demonstrate that the physiological concentration of the enzymes as well as the chromosomal sensitivity exhibited a wide interindividual variability. An inverse correlation between chromosomal sensitivity to bleomycin and SOD concentration in whole blood, plasma and red cells was found. On the other hand, no correlation between the yield of bleomycin-induced dicentrics and the concentration of CAT or POD was detected in any of the blood fractions analyzed. These findings suggest that the concentration of SOD may play an important role in the cellular susceptibility to DNA damage by free radicals.

Adult↗

PubMed1986

Variation in sister chromatid exchange frequencies between human and pig whole blood, plasma leukocyte, and mononuclear leukocyte cultures.

Sister chromatid exchange (SCE) induction by ultraviolet (UV) light was studied in both human and pig whole blood cultures (WBC) and plasma leukocyte cultures (PLC). No variation in SCE frequency was observed between pig WBC and PLC in control as well as in treated cells. Conversely, SCE frequencies of human PLC were consistently higher than those of WBC in control and UV-exposed cells. Thus, red blood cells (RBCs) do not influence the sensitivity of lymphocytes to UV light exposure, and there must be some different culture condition(s) in the induction of SCEs between human WBC and PLC but not in swine lymphocyte cultures. Since the BrdUrd/lymphocyte ratio of WBC was halved in PLC, the effect of BrdUrd concentration in inducing the SCE baseline frequency of PLC may be ruled out. Also, Ficoll-Hypaque-separated human mononuclear leukocytes in culture (MLC), but not pig MLC, showed a two-fold increase in SCE frequency over WBC values. Thus, neither the cell separation technique nor polymorphonuclear leukocytes had a significant role in the elevated SCE frequency of human PLC or MLC. Experiments where human RBCs were titrated into human PLC showed that the induction of an elevated SCE frequency of PLC was suppressed in a dose-dependent manner by the presence of RBCs in the culture medium. Since the incorporation of pig or human RBCs into human PLC as well as into MLC reduced the SCE frequency to that of WBC, a common component and/or function existing in these cells is suggested. Analysis of different RBC components showed that RBCs, specifically RBC ghosts, release a diffusible but not dialyzable "corrective" factor into culture medium that is able to reduce the SCE frequencies of PLC. Antioxidant enzymes such as catalase and horseradish peroxidase were unable to reduce the SCE frequency of human PLC to WBC values.

Animals↗