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1.
Fertil Steril ; 74(2): 245-50, 2000 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-10927039

RESUMO

OBJECTIVE: [1] To determine whether apoptosis can be measured in ejaculated spermatozoa by flow cytometry using the Annexin V assay, which measures expression of phosphatidylserine on the outer leaflet of the cell membrane, or the TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP [deoxy-uridine triphosphate] nick end labeling) assay, which measures occurrence of DNA strand breaks and [2] to correlate the outcome with routine semen variables and the hypoosmotic swelling (HOS) test. DESIGN: Pilot study and clinical trial. SETTING: Large teaching hospital and fertility center. PATIENT(S): Men whose semen was studied for various reasons. MAIN OUTCOME MEASURE(S): Percentage of apoptotic spermatozoa by two different assays, percentage of necrotic spermatozoa, concentration and motility of spermatozoa, and outcome of the HOS test. RESULT(S): Apoptosis can be measured in spermatozoa by flow cytometry using the Annexin V assay and the TUNEL assay. Twenty percent of spermatozoa were apoptotic according to both assays. A significant inverse correlation was seen between phosphatidylserine expression (Annexin V assay) and sperm concentration (r = -0.389; P<.05) and motility (r = -0.289; P<.05). A highly significant inverse correlation was seen between DNA double-strand breaks (TUNEL assay) and sperm concentration (r = -0.629; P<.0001). CONCLUSION(S): Flow cytometry can easily and reliably detect phosphatidylserine expression on the outer leaflet of the cell membrane and DNA strand breaks, both of which are hallmarks of apoptosis. About 20% of ejaculated spermatozoa are apoptotic, and the concentration of spermatozoa is lower in men with more apoptotic spermatozoa.


Assuntos
Apoptose , Citometria de Fluxo/métodos , Marcação In Situ das Extremidades Cortadas/métodos , Infertilidade Masculina/patologia , Espermatozoides/fisiologia , Anexina A5/análise , Anexina A5/metabolismo , Fragmentação do DNA , Fluoresceína-5-Isotiocianato/análise , Fluoresceína-5-Isotiocianato/metabolismo , Corantes Fluorescentes , Humanos , Masculino , Fosfatidilserinas/análise , Fosfatidilserinas/metabolismo , Projetos Piloto , Propídio , Motilidade dos Espermatozoides
2.
Cytometry ; 21(2): 203-10, 1995 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-8582241

RESUMO

In vivo S-phase cell labeling with iododexoyuridine (IdUrd) was performed in six multiple myeloma (MM) patients. Myeloma cells from four patients were hyperploid. In three out of four patients, DNA/IdUrd flow cytometry revealed that most of the labeled cells, which had divided during the period, elapsed between flash labeling and sampling, had returned to the diploid G0/G1 compartment and not to the hyperdiploid peak. To eliminate contaminating cells belonging to the normal hematopoiesis, plasmocytic and lymphocytic cells were fractionated and analyzed separately. Cell enrichment was performed with use of murine monoclonal antibodies (MoAbs) against plasmocytic and lymphocytic cell markers and subsequent magnetic activated cell sorting with immunobeads, i.e., polysterene magnetic particles coated with sheep anti-mouse IgG. The purified plasmocytic cells were mainly hyperdiploid and largely unlabeled. The lymphocytic cells appeared to belong chiefly to the diploid cell population. The IdUrd-labeled cells were predominantly lymphocytic cells, returning after mitosis to the diploid G0/G1 peak. Although this pattern of S-phase cells in hyperdiploid MM, belonging to the diploid cell compartment, was observed in three out of four hyperploid cases and although the number of observations is small, S-phase cells may demonstrate an aspect of tumor cell kinetics in hyperploid MM, which has been debated for many years and which indicates the existence of a non-plasmocytic stem cell compartment that feeds the plasmocytoma. The behavior of the labeled cells as observed in a few cases of MM provides another, hitherto undescribed, argument that, at least in some MM patients, a part of the proliferating tumor cells may be diploid lymphocytic (precursor) cells. These findings should be considered when targeting and monitoring treatment of MM and also in purging procedures of bone marrow in patients to be treated by ablative cytotoxic therapy and autologous bone marrow transplantation.


Assuntos
Medula Óssea/patologia , Ciclo Celular , DNA de Neoplasias/análise , Mieloma Múltiplo/patologia , Idoso , Diploide , Feminino , Citometria de Fluxo/métodos , Fase G1 , Células-Tronco Hematopoéticas/patologia , Humanos , Idoxuridina , Linfócitos/patologia , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/genética , Estadiamento de Neoplasias , Plasmócitos/patologia , Fase de Repouso do Ciclo Celular , Fase S
3.
Rheumatology (Oxford) ; 41(4): 458-61, 2002 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-11961179

RESUMO

BACKGROUND: Interaction with platelet function by non-steroidal anti-inflammatory drugs (NSAIDs) is related to the inhibition of cyclo-oxygenase-1 (COX-1). In patients with rheumatoid arthritis (RA), only one of the COX-2-selective NSAIDs (nabumetone) has been demonstrated to spare platelet function partially. OBJECTIVE: To compare the effects of the COX-2-selective inhibitor, meloxicam, with those of the non-selective NSAID, naproxen, on platelet function and thromboxane levels in RA patients. METHODS: In this randomized, controlled, cross-over trial, 10 RA patients used meloxicam 7.5 mg bid and naproxen 500 mg bid, each during a 2-week period. Washout periods were applied. Before and after each 2-week period of NSAID intake, laboratory studies were performed. RESULTS: Platelet aggregation was significantly less influenced, thromboxane levels were less inhibited (246 vs 117 pg/ml) and bleeding times were less prolonged with meloxicam than with naproxen (3.2 vs 2.3 min). Moreover, the results of all tests during meloxicam exposure were comparable with baseline values. CONCLUSION: In RA patients, meloxicam, a representative of the selective COX-2 inhibitors, does not interfere with platelet function and thromboxane levels, in contrast with naproxen (a non-selective COX inhibitor).


Assuntos
Anti-Inflamatórios não Esteroides/uso terapêutico , Artrite Reumatoide/tratamento farmacológico , Inibidores de Ciclo-Oxigenase/uso terapêutico , Naproxeno/uso terapêutico , Agregação Plaquetária/efeitos dos fármacos , Tiazinas/uso terapêutico , Tiazóis/uso terapêutico , Adulto , Artrite Reumatoide/sangue , Artrite Reumatoide/enzimologia , Tempo de Sangramento , Plaquetas/efeitos dos fármacos , Plaquetas/enzimologia , Estudos Cross-Over , Relação Dose-Resposta a Droga , Feminino , Humanos , Técnicas In Vitro , Masculino , Meloxicam , Pessoa de Meia-Idade , Distribuição Aleatória , Tromboxano B2/sangue
4.
Acta Haematol ; 98(1): 8-13, 1997.
Artigo em Inglês | MEDLINE | ID: mdl-9210907

RESUMO

It has been reported that cultured peripheral B lymphocytes of chronic lymphocytic leukemia (B-CLL) patients show a high degree of apoptosis (programmed cell death). Till now, no data exist about the occurrence of in vitro apoptosis of normal B and T cells. We measured the amount of apoptosis and secondary necrosis (type 2 necrosis) in B-CLL lymphocytes and in normal peripheral B and T lymphocytes in culture. Observations were made on B-CLL lymphocytes and on normal B and T cells purified by immunomagnetic cell sorting. Apoptosis and secondary necrosis were measured using a recently described sensitive flow-cytometric assay, probing simultaneously for cell surface exposure of phosphatidylserine with the use of FITC-labeled annexin-V and for cell membrane integrity as demonstrated by the exclusion of propidium iodide. The degree of in vitro apoptosis and secondary necrosis of normal B cells appears to be higher than that of normal T cells, and even higher than that of B-CLL cells. The results indicate that cultured mature circulating normal B lymphocytes exhibit a higher cell death rate than normal T cells and B-CLL lymphocytes.


Assuntos
Apoptose , Leucemia Linfocítica Crônica de Células B/patologia , Anexina A5 , Linfócitos B/patologia , Citometria de Fluxo , Fluoresceína-5-Isotiocianato , Corantes Fluorescentes , Humanos , Separação Imunomagnética , Cinética , Necrose , Linfócitos T/patologia , Células Tumorais Cultivadas
5.
Arthritis Rheum ; 46(6): 1498-503, 2002 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-12115179

RESUMO

OBJECTIVE: Platelets are involved in various thrombotic events, often by means of platelet-derived microparticles (PMPs). It is likely that platelets are also involved in inflammation. Because inflammatory processes play a central role in rheumatoid arthritis (RA), we sought to determine whether PMPs are present in this disease. METHODS: This descriptive, cross-sectional study included 19 RA patients and 10 healthy controls. Nine of the patients had active RA (erythrocyte sedimentation rate [ESR] > or =28 mm/hour and/or C-reactive protein [CRP] level > or =28 mg/liter, > or =9 painful joints, and > or =6 swollen joints), and 10 had inactive disease (ESR < or =27 mm/hour, CRP < or =27 mg/liter, no tender joints, and no swollen joints). Platelet counts and PMP numbers were determined using cell counter and flow cytometry, respectively. RESULTS: Platelet counts in the 3 groups were similar. However, levels of PMPs in RA patients were significantly higher than those in healthy controls (median 616 versus 118 x 10(6)/liter; P = 0.005). PMP levels were higher in patients with active RA than in those with inactive RA (median 2,104 versus 504 x 10(6)/liter; P > 0.05). Moreover, PMP levels correlated with disease activity (r = 0.67, P = 0.05). CONCLUSION: PMPs are associated with RA, and PMP levels are correlated with disease activity. Thus, platelets probably play a part in the inflammatory process of RA by means of PMPs. Given the importance of PMPs in cardiovascular diseases, this may be one reason for the enhanced cardiovascular morbidity and mortality in RA.


Assuntos
Artrite Reumatoide/sangue , Artrite Reumatoide/imunologia , Plaquetas/imunologia , Adulto , Idoso , Estudos Transversais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Contagem de Plaquetas , Índice de Gravidade de Doença
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