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Biomedical subjects

A Raza

Publications and source records attributed to A Raza.

At least 91 records · Page 5Linked to original sources
PubMed1996

Indication of an involvement of interleukin-1 beta converting enzyme-like protease in intramedullary apoptotic cell death in the bone marrow of patients with myelodysplastic syndromes.

Our previous studies using in situ end labeling (ISEL) of fragmented DNA revealed extensive apoptotic cell death in the bone marrows (BM) of patients with myelodysplastic syndromes (MDS) involving both stromal and hematopoietic cells. In the present report we show greater synthesis of interleukin-1 beta (IL-1 beta) in 4 hour cultures of density separated BM aspirate mononuclear (BMAM) cells from MDS patients as compared to the cultures of normal BM from healthy donors or lymphoma patients (1.7 +/- 0.37 pg/10(5) cells, n = 29 v 0.42 +/- 0.24 pg/10(5) cells, n = 11, respectively, P = .049). Further, these amounts of IL-1 beta in MDS showed a significant correlation with the extent of apoptosis detected by ISEL in corresponding plastic embedded BM biopsies (r = .480, n = 30, P = .007). In contrast normal BMs did not show any correlation between the two (r = .091, n = 12, P = .779). No significant correlation was found between the amounts of IL-1 beta and % S-phase cells (labeling index; LI%) in MDS determined in BM biopsies using immunohistochemistry following in vivo infusions of iodo- and/or bromodeoxyuridine. Neither anti-IL-1 beta antibody nor IL-1 receptor antagonist blocked the apoptotic death of BMAM cells in 4 hour cultures (n = 5) determined by ISEL (apoptotic index; AI%), although the latter led to a dose-dependent accumulation of active IL-1 beta in the culture supernatants. On the other hand, a specific tetrapetide-aldehyde inhibitor of ICE significantly retarded the apoptotic death of BMAM cells at 1 mumol/L in 5/6 MDS cases studied (AI% = 2.99 +/- 0.30 in controls v 1.58 +/- 0.40 with ICE-inhibitor, P = .05) and also reduced the levels of active IL-1 beta synthesized (5.59 +/- 2.63 v 2.24 +/- 0.93 pg/10(6) cells, respectively). In normal cells, neither IL-1 blockers nor the ICE inhibitor showed any effect on the marginal increase in apoptosis observed in 4 hour cultures. Our data thus suggest a possible involvement of an ICE-like protease in the intramedullary apoptotic cell death in the BMs of MDS patients.

Antibodies, Monoclonal↗
PubMed1996

Novel insights into the biology of myelodysplastic syndromes: excessive apoptosis and the role of cytokines.

The paradox of myelodysplastic syndromes (MDS) which present with pancytopenias despite cellular bone marrows (BM) was investigated by conducting detailed studies of proliferation and apoptosis in 89 MDS patients. Our results demonstrated a rapid rate of both proliferation as well as apoptosis. Levels of three cytokines, tumor necrosis factor-alpha (TNF-alpha), transforming growth factor-beta (TGF-beta) and interleukin-1 beta (IL-1 beta) were measured in the same patients. High levels of TNF-alpha were found to correlate with high levels of apoptosis in 83 MDS patients (P = 0.0045). We propose a dual role for TNF-alpha (or other cytokines) in the pathogenesis of MDS. On the one hand, TNF-alpha induces apoptosis in the maturing cells causing pancytopenia while on the other, it stimulates the proliferation of the primitive progenitors accounting for the hypercellular BM frequently seen in MDS. A new model for MDS is presented. The initial abnormality probably affects a primitive hemopoietic progenitor which acquires a growth advantage leading to monoclonal hemopoiesis, which in turn makes these cells susceptible towards acquiring additional mutations and appearance of cytogenetically marked (or unmarked) clones. Cytokines such as TNF-alpha whose source is presently unknown, then contribute towards the clinical syndrome of pancytopenia and hypercellularity.

Antigens, CD34↗
PubMed1996

Enzymatic programming of apoptotic cell death.

The phenomenon of 'apoptosis' was recognized in the early seventies. However, most of the mechanistic insights into the process of apoptotic cell death came about only in the last few years. It is now clear that a variety of enzymes constitute the major mediators of this process. The present article reviews the possible interrelationships between these enzymes and proposes a model for enzymatic programming of apoptotic cell death.

Animals↗
PubMed1996

A paradigm shift in myelodysplastic syndromes.

A poorly defined transforming event(s) affects the pluripotential bone marrow (BM) stem cell in myelodysplastic syndromes (MDS), conferring a growth advantage upon it which leads eventually to monoclonal hematopoiesis. The progeny of this transformed ancestor undergo recognizable albeit dysplastic maturation. We propose that this picture is further complicated by a variety of cytokines, tumor necrosis factor alpha (TNF-alpha), transforming growth factor beta (TGF-beta) and interleukin 1beta (IL-1beta) which exert a dual effect on the diseased cells. The immature CD34+ cells are stimulated to proliferate, while their later differentiated daughters are induced to undergo apoptosis accounting for the clinical syndrome of pancytopenia despite hypercellular BMs. Studies directed at measuring the rates of proliferation and apoptosis as well as the levels of TNF-alpha, TGF-beta and IL-1beta confirm this hypothesis and are presented in greater detail. A novel approach towards MDS therapy emerges as a result of this paradigm shift based upon the premise that anti-cytokine therapy would prevent excessive intramedullary apoptosis and result in improved cytopenias as well as cause a slowing down of the diseased precursor cell proliferation resulting in resumption of polyclonal hematopoiesis. Because a number of cytokines function through common lipid second messengers, interruption of this pathway should theoretically cause disruption in the signalling of a cascade of cytokines.

Apoptosis↗
PubMed1995

Apoptosis in bone marrow biopsy samples involving stromal and hematopoietic cells in 50 patients with myelodysplastic syndromes.

Cell-cycle kinetics were measured in situ after infusions of iododeoxyuridine and/or bormodeoxyuridine in 50 patients with myelodysplastic syndromes (MDS) and the median labeling index in bone marrow (BM) biopsy samples was 28.6%. Unfortunately, 26 of 50 patients showed that > or = 75% of hematopoietic cells of all three lineages were undergoing programmed cell death (PCD) in their biopsy samples as shown by the in situ end labeling (ISEL) technique. Ten patients had 1/3 and eight had 2/3 ISEL+ cells. Stromal cells were frequently ISEL+ and often S-phase cells were also found to be simultaneously ISEL+. Nucleosomal DNA fragments as a ladder in agarose gel were present in BM aspirates of four patients who showed high ISEL and were absent in two who had no ISEL staining in biopsy samples, but only when DNA was extracted after a 4-hour in vitro incubation in complete medium. Therefore, laddering data confirmed the ISEL findings that the majority of hematopoietic cells in MDS are in early stages of PCD. We conclude that extensive intramedullary cell death may explain the paradox of pancytopenia despite hypercellular marrows in MDS patients. Investigating approaches that protect against PCD in some MDS subsets would be of interest.

Acute Disease↗
PubMed1995

Simultaneous assessment of cell kinetics and programmed cell death in bone marrow biopsies of myelodysplastics reveals extensive apoptosis as the probable basis for ineffective hematopoiesis.

Despite hypercellular bone marrows (BM), peripheral cytopenias are the rule in patients with myelodysplastic syndromes (MDS). This study examined the roles played by cell birth and cell death rates in generating this paradox. Cell kinetics from BM biopsies of 35 MDS patients were measured using intravenous infusions of either iododeoxyuridine or bromodeoxyuridine, or both. Degree of apoptosis or programmed cell death (PCD) was estimated using in situ end-labeling of DNA directly from BM biopsies, which were simultaneously double-labeled from proliferation/PCD. MDS were found to be highly proliferative disorders with large numbers of myeloid, erythroid, and megakaryocytic cells synthesizing DNA. Median cycling time (Tc) of myeloblasts was more rapid than that of patients with acute myeloid leukemia (44.1 hr vs. 56.0 hr). Interestingly, most marrow cells of all three lineages in 32 of 34 evaluable cases were undergoing PCD. In 19 of 32 patients, greater than 75% cells were apoptotic. Surprisingly, large numbers of S-phase cells were found to be simultaneously undergoing PCD, as were stromal cells of the BM microenvironment. We conclude that the extensive apoptosis in hematopoietic cells effectively cancels the high birth rate resulting in ineffective hematopoiesis and accounting for deficient bone marrow function.

Acute Disease↗
PubMed1995

Distribution of cell cycle times amongst the leukemia cells within individual patients with acute myelogenous leukemia.

Seventeen patients with AML received infusions of BrdUrd to permit measurement of the cell parameters of leukemia cells in vivo. The range of S-phase times was measured by using a two color BrdUrd/Pl analysis to determine the range of BrdUrd incorporation into cells which had been in S-phase throughout the entire duration of BrdUrd administration. These data were, in turn, used to calculate the range of cell cycle times amongst the leukemia cells present within individual patients. The range of cell cycle times amongst the leukemia cells present within individual patients differs between patients, with some leukemia cell populations characterized by narrow and others by broad ranges. In general, the longer the mean cell cycle time (between 27 and 112 h) the broader the range of cell cycle times. These differences may help to explain, in part, the differences in response to therapy among patients whose mean leukemia cell kinetic parameters are similar.

Bromodeoxyuridine↗
PubMed1995

Fibroblasts isolated after fibrotic lung injury induce apoptosis of alveolar epithelial cells in vitro.

Primary lung fibroblasts were isolated from patients with idiopathic pulmonary fibrosis (HIPF), from normal human lung tissue (NH), from rats treated with 75% oxygen and paraquat (PA), and from normal adult rats (NR). Serum-free media conditioned by each fibroblast strain were tested on the human A549 cell line (HIPF and NH media) or on primary alveolar epithelial cells (AEC) isolated from normal adult rats (PA or NR media). Over 20-h incubation, HIPF- or PA-conditioned media induced DNA fragmentation and significant decreases in total recoverable DNA and cell number of A549 or AEC, respectively; NH or NR media had no significant effect relative to serum-free unconditioned media. Apoptosis of A549 and AEC was detected by altered nuclear morphology and was confirmed by terminal deoxynucleotidyl transferase-mediated bio-dUTP nick end labeling. The endonuclease inhibitors 10 microM aurintricarboxylic acid and 50 microM zinc inhibited HIPF-induced apoptosis of A549 cells by 68 and 71%, respectively. Both apoptosis and necrosis were induced by HIPF and PA media in a concentration-dependent manner. These results suggest that altered fibroblasts emerging during fibrotic lung injury release a soluble factor(s) capable of inducing cell death and net loss of AEC.

Animals↗
PubMed1995

A multivariate study of non Hodgkin's lymphoma involving proliferation, apoptosis, bcl-2 and the microenvironment.

The study was carried out on 22 patients with non-Hodgkin's lymphoma (NHL) who had received sequential infusions of two thymidine analogues iododeoxyuridine (IUdR) and bromodeoxyuridine (BrdU). Cell cycle kinetic studies seemed to differentiate distinctly between low grade lymphoma (n = 8, LI = 2.6%) compared to that of intermediate grade (n = 9, LI = 13%, p = 0.0001) and high grade NHL (n = 5, LI = 16.3%, p = 0.0062). While the majority of 14 intermediate and high grade lymphomas had a high labeling index there were 3/14 patients with a LI of 5.5%, 5.5% and 4.1% respectively. A decrease in the rate of programmed cell death (PCD) or apoptosis due to the overexpression of bcl-2 has been implicated as the possible pathogenesis for follicular lymphoma. We determined the presence of bcl-2 protein immunohistochemically and apoptosis by in situ end labeling of DNA which detects cells in early stages of PCD not recognized morphologically. Nine NHL patients demonstrated PCD ranging from 1%-40%, while it was undetectable in 13/22 patients. Of these 13 cases, 6 showed the presence of bcl-2 expression. To understand the relationship of the microenvironment to the lymphoma cells, the presence of transforming growth factor beta (TGF-beta) was determined immunohistochemically. TGF-beta was present in all the cases where bcl-2 was present, except one. This study highlights some of the key biological features of NHL cells and their microenvironment.

Apoptosis↗
PubMed1994

Age-related changes in brainstem auditory neurotransmitters: measures of GABA and acetylcholine function.

This study was designed to determine if there are age-related alterations in the bio-synthetic enzyme glutamic acid decarboxylase (GAD), the degradative enzyme GABA-transaminase (GABA-T), and the uptake system for GABA in the central nucleus of the inferior colliculus (CIC), the cochlear nucleus (CN), and/or nuclei of the lateral lemniscus (NLL) of Fischer-344 rats. For purposes of comparison, the cholinergic neuronal system was studied in parallel in young adult (3-7 months), mature (15-17 months) and aged (24-26 months) rats. In young adults GAD activity was highest in the CIC (219 nmol/mg protein/h; N = 5), intermediate in NLL (82 nmol/mg protein/h), and lowest in CN (34 nmol/mg protein/h). Choline acetyltransferase (ChAT) activity was highest in NLL and CN, and approximately 35-40% lower in CIC. A more uniform pattern was observed with GABA-T activity. Reductions in GAD activity were seen in the CIC of mature (-31%) and aged (-30%) rats that were not graded with age when compared to young adult, P < 0.05 (N = 5). This effect was regionally selective, since the CN did not show any loss of GAD or ChAT activity. The neurotransmitter selectivity of this deficit in CIC is supported by the non-parallel changes in ChAT activity (-22%, aged vs. mature, P < 0.05) that occurred after the changes in GAD activity. In contrast to the loss of GABAergic biosynthetic capacity in aged CIC, high affinity uptake processes (Kd and Vmax) for 14C-GABA and 3H-D-aspartate were not significantly altered (P > 0.05). Similar to the CIC, the NLL showed remarkable age-related deficits, but these deficits were more substantial for the cholinergic system (ChAT activity: -56% aged vs. young adult, P < 0.05; GAD activity: -35% aged vs. mature). None of the areas examined showed a significant loss of GABA-T activity with aging. These data suggest: 1) Age-related loss of GABA-mediated inhibition in the CIC of Fischer-344 rats is not attributable to changes in uptake or degradation of GABA, but may be related loss of biosynthetic capacity (i.e. activity or quantity) of the GAD present; 2) processing centers of the central auditory pathway (i.e. CIC and NLL), but not necessarily primary (i.e. CN) integrative nuclei, demonstrate selective, age-related neurochemical deficits; and 3) age-related neurochemical changes in central auditory structures may contribute substantially to the abnormal perception of signals in noise and loss of speech discrimination observed in neural presbycusis.

4-Aminobutyrate Transaminase↗
PubMed1994

Parallel studies of clonogenic leukaemia cells and the leukaemia cell population as a whole in acute myelogenous leukaemia.

The clonogenic cells in patients with acute myelogenous leukaemia (AML) were evaluated with respect to the relationship between primary and secondary cloning capacity and the proliferative and molecular biological characteristics of the leukaemia cell population as a whole. Secondary cloning capacity was correlated with primary cloning efficiency, and with the ability of the clonogenic cells to produce large sized clones. The cloning capacity of AML cells was unrelated to the cell cycle characteristics of the leukaemia cell population in vivo or to the level of myc, myb, fms, or interleukin (IL)1 beta expression. The sensitivities of the clonogenic cells to cytosine arabinoside and daunorubicin were inversely correlated with the ability of the leukaemia cells to produce large sized clones in vitro. This latter observation may explain the reported relationships between the clonogenic capacity of AML cells and response to chemotherapy.

Cell Cycle↗
PubMed1994

Transmission-blocking antibodies against multiple, non-variant target epitopes of the Plasmodium falciparum gamete surface antigen Pfs230 are all complement-fixing.

We have studied the properties of 16 newly derived monoclonal antibodies (MoAbs) against Pfs230, a gamete surface protein of Plasmodium falciparum and a target of transmission-blocking antibodies. All 16 MoAbs recognized Pfs230 by immunoprecipitation from non-ionic detergent extracts of the protein radio-labelled with 125Iodine. The MoAbs also recognized this protein on Western blots under non-reducing conditions but none of them recognized the protein under reducing conditions. Using an immunoradiometric assay the MoAbs appear to define nine different epitope regions. The MoAbs were tested for their ability to lyse extra-cellular female gametes of P. falciparum isolate 3D7. Eight of the MoAbs, all of isotype IgG2a, mediated complement-dependent lysis of the gametes; seven of the MoAbs, all isotype IgG1, failed to lyse the gametes in the presence of active complement. The eight complement-fixing MoAbs mediated almost total suppression of infectivity of gametocytes of P. falciparum 3D7 to mosquitoes; where tested this suppression was mainly complement-dependent. The seven non-complement-fixing MoAbs had no significant effect on the infectivity of gametocytes of P. falciparum 3D7 to mosquitoes. When tested by immunofluorescence the target epitopes of all the MoAbs were conserved in each of the five different isolates of P. falciparum which were tested.

Animals↗
PubMed1994

Novel in situ double labeling for simultaneous detection of proliferation and apoptosis.

We describe a novel double-labeling method to simultaneously investigate proliferation and apoptosis from plastic-embedded biopsy specimens (PEBs). Infusions of bromo- and/or iododeoxyuridine (BrdU/IudR) were given to 10 patients, five with acute myeloid leukemia (AML) and five with myelodysplastic syndromes (MDS), and S-phase cells were measured in PEBs using a monoclonal anti-IudR/BrdU antibody. Apoptosis was measured by in situ end-labeling (ISEL) of DNA. The results demonstrate that both AML and MDS are highly proliferative disorders but that there is almost no apoptosis in the former, whereas extensive apoptosis was observed in the latter. Double labeling revealed that large numbers of S-phase cells in MDS were simultaneously undergoing apoptosis. We conclude that the high cell death in MDS cancels the high cell birth, resulting in a functionally aplastic marrow and thus accounting for the observed ineffective hematopoiesis. On the other hand, AML is rapidly fatal, probably owing to high cell birth with no or minimal cell death. Therapeutic strategies to prevent intramedullary programmed cell death of hematopoietic precursors should be evaluated in MDS, and efficacy of chemotherapy in AML can be assessed by measuring the induction of apoptosis in post-treatment biopsy specimens.

Acute Disease↗
PubMed1994

The relationship of the in vivo cell cycle characteristics and treatment outcome in acute myelogenous leukemia to the expression of the FMS and MYC proto-oncogenes.

Fms expression was measured by Northern blot analysis in the pretherapy leukemia cells of 30 patients with acute myelogenous leukemia (AML) who either entered CR or who were shown to have drug resistant disease. High levels of fms expression correlated with a CR outcome and were strongly associated with the fall in the number of leukemia cells during the first six days of remission induction therapy. In contrast, similar studies of myc expression failed to demonstrate any relationship to treatment outcome. Considering those genes whose expression have been measured to date, the level of expression of fms is the one which is most highly correlated with treatment outcome in AML. No significant relationship between the expression levels of these two genes and the cell cycle characteristics of leukemia cells in vivo were detected.

Blotting, Northern↗
PubMed1994

The study of acute leukemia cells by means of acridine orange staining and flow cytometry.

The studies described here explored the staining of acute leukemia cells with acridine orange (AO). The red fluorescence curve of AML specimens was usually bimodal, suggesting the presence of subpopulations of cells which have different RNA contents. In almost every AML specimen, small leukemic blast cells comprised at least part of the "low RNA content" subpopulation. Residual granulocytes and lymphocytes also contributed to this population. Frequently, the green fluorescence, indicative of the binding of AO to DNA, was slightly less in these cells than in the majority of cells present. There was no evidence however, that the leukemia cells with these characteristics represented a G0 or kinetically quiescent population of cells. In ALL specimens, the presence of multiple cytogenetically distinct clones was easily detectable in AO stained specimens. The red fluorescence curve of G0/G1 ALL cells was unimodal.

Acridine Orange↗
PubMed1993

Clinical and prognostic significance of in vivo differentiation in acute myeloid leukemia.

Bromodeoxyuridine (BrdU) was administered to 86 newly diagnosed patients with standard risk acute myeloid leukemia (AML) prior to starting induction therapy and the labeling index (LI), durations of S-phase (Ts), and the cell cycle (Tc) of myeloblasts were determined. Induction therapy with cytosine arabinoside and daunomycin was subsequently started. Bone marrow biopsies were obtained on days 6 and 17 and weekly thereafter, and were treated with a monoclonal anti-BrdU antibody to determine the fate of cells labeled on day 0 by BrdU. BrdU labeled granulocytes indicating the presence of in vivo differentiation (Diff+) were identified in 48 patients ranging from 1+ (1-10 labeled cells) to 4+ (greater than 31 labeled granulocytes). When compared to 38 differentiation negative (Diff-) patients, Diff+ group had longer Ts (14.5 hr vs. 10.95 hr, P = 0.015) and Tc (59.7 hr vs. 41.7 hr, P = 0.017). Remission duration was significantly longer (no median) for 3-4+ Diff+ as compared to Diff- (median = 220 days) patients (Wilcoxon P = 0.04). We conclude that the detection of in vivo differentiation in AML patients indicates a favorable long-term prognosis either due to the presence of a substantial amount of normal residual hematopoiesis prior to starting induction therapy or due to the ability of leukemic cells to undergo differentiation.

Acute Disease↗
PubMed1993

Biological characteristics of newly diagnosed poor prognosis acute myelogenous leukemia.

A pilot study was conducted of the biological characteristics of the leukemia cells of newly diagnosed patients with poor prognosis acute myelogenous leukemia (AML). This study included measurements of the pretherapy proliferative rate of the leukemia cells in vivo, assessment of differentiation in vivo during remission induction therapy, and the level of expression of the fms, myc, and IL1 beta genes in pretherapy leukemia cells. Short cell cycle times were characteristic of the best prognostic category and were associated with a rapid reduction in marrow leukemia cells in cytosine arabinoside (araC)-sensitive patients. Expression of c-fms was associated with rapid reduction in marrow leukemia cells during araC therapy and with a successful treatment outcome. Expression of the IL1 beta gene was associated with short remissions. These studies suggest that when compared to newly diagnosed standard prognosis AML, the leukemia of poor prognosis patients is more likely to exhibit long cell cycle times, low levels of fms expression, and is less likely to be associated with myeloid differentiation during remission induction therapy.

Aged↗