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Biomedical subjects

T C Fisher

Publications and source records attributed to T C Fisher.

At least 37 records · Page 2Linked to original sources
PubMed1994

Apoptosis and cancer chemotherapy.

The major disseminated cancers remain stubbornly resistant to systemic therapy. Drug-resistant tumours include both slow and fast growing types, with the carcinomas constituting the major problem. Strategies for drug discovery have, in the past, been focused on attempts to design antiproliferative agents, largely targeted to interfere with DNA integrity and replication. The malignant phenotype might be characterized by the emergence of cell populations with a greater survival potential: a lower proclivity to undergo apoptosis. This idea provides a possible explanation of the genesis and progression of cancer and of the inherent resistance of tumour cells to engage apoptosis. Work is described which identifies the molecular basis for differences in the survival potential of stem cells in the crypts of the colon and small intestine. The advantageous survival of colonic stem cells, provided by expression of bcl-2 and a muted p53 response to DNA damage, allows damaged cells to survive. Continued expression of bcl-2 renders tumour cells resistant to drug-induced DNA damage by a mechanism different from classical mechanisms of drug resistance. The attenuation of cell survival is described as a key component in strategies for the drug treatment of disseminated cancers.

Animals↗
PubMed1994

Polymorphonuclear leukocytes in ischemic vascular disease.

Over the last decade, an extensive amount of evidence has accumulated which implicates PMN in the etiology and pathophysiology of ischemic/thrombotic diseases. It has become apparent that PMN infiltration is not, as once thought, an innocent secondary phenomenon following ischemia. Rather, PMN are active participants in the pathophysiology of infarction, exacerbating the tissue damage. Since the development of means to achieve reperfusion after thrombosis, this phenomenon has become of critical importance. Many different approaches, targeted at prevention of PMN trapping in the capillaries of the ischemic, area, have been shown to effectively reduce the final infarct size, and will likely prove valuable adjuncts to reperfusion. However, perhaps the most significant aspect of the realization that PMN play a significant role in thrombotic disease may prove to be the potential for early intervention: Elevated PMN counts are predictive of ischemic events, and there is preliminary evidence that the elevated PMN count may be also associated with increased PMN activation, suggesting that research directed at the prophylactic use of anti-PMN agents might someday prove effective in reducing the incidence of MI and stroke.

Acute Disease↗
PubMed1994

Decreased polymorphonuclear leukocyte deformability in NIDDM.

OBJECTIVE: To determine the rheological properties of polymorphonuclear leukocytes (PMN) from non-insulin-dependent diabetes mellitus (NIDDM) patients. RESEARCH DESIGN AND METHODS: The deformability of PMN from 33 NIDDM subjects, 13 with impaired glucose tolerance (IGT), and 22 with normal glucose tolerance (NGT) was studied. A Cell Transit Analyzer that measures the transit time of PMN through 8-microns pores was used. Studies were performed under three different conditions: 1) basal state; 2) after incubation with cytochalasin B (20 microM) to dissociate f-actin from the cytoskeleton; and 3) following activation with N-formyl-methionyl-leucyl-phenylalanine (fMLP, 1 nM). RESULTS: PMN from diabetic patients were more rigid (i.e., had longer transit time) than those from subjects with NGT or IGT under basal conditions and after cytochalasin B, but not after stimulation with fMLP. The deformability of PMN from subjects with IGT was similar to those of the NGT group. In the pooled data, basal transit time correlated with age; systolic and diastolic blood pressure; HbA1c; and serum creatinine, cholesterol, and triglyceride concentrations (r = 0.29, 0.34, 0.37, 0.48, 0.25, 0.36, 0.29, respectively, P < 0.05 for each). Hypertensive diabetic patients had less deformable PMN than normotensive ones. No relation was found between PMN deformability and the duration of diabetes, type of treatment, or the presence of retinopathy. CONCLUSIONS: These data indicate increased rigidity of PMN in NIDDM that may contribute to development of microcirculatory disturbances and microangiopathy.

Adult↗
PubMed1994

Beta-S gene cluster haplotypes modulate hematologic and hemorheologic expression in sickle cell anemia. Use in predicting clinical severity.

PURPOSE: The rate of progression of major organ failure in sickle cell anemia is genetically controlled. It is the direct consequence of the sickle cell-evoked vasculopathy. PATIENTS AND METHODS: Presence of the beta S gene cluster haplotypes and alpha gene deletions as genetic markers indicate the expected frequency of illness and the risk of end-stage major organ failure. The risk of irreversible soft tissue organ failure is greatest in patients with a Central African Republic (CAR) chromosome, whereas morbidity is consistently lowest in patients with a Senegalese chromosome. Presence of alpha-thalassemia-2 decreases the risk of soft tissue organ failure in all haplotype combinations. RESULTS: Other laboratory abnormalities, when combined with haplotype and alpha gene status, also predict the risk of clinical morbidity. The mean hemoglobin level (or red blood cell count) is lowest in patients with the most severe clinical manifestations. On the other hand, the platelet count and leukocyte count as well as the plasma fibrinogen level are elevated in the sickest patients. A threshold level of hemoglobin F at 1.2 g/dl (approximately 20% hemoglobin F) decreases the risk of major organ failure and is attained most frequently in those with a Senegalese chromosome. Hemorheologic findings observed during the most stable state of patients with sickle cell anemia indicate two trends: (a) the mean percentage of dense red cells is nearly twice as high in the maximal severity patients as compared with the minimal severity patients; and (b) mean red cell rigidity is greatest in the maximal severity group and least in the minimal severity group. These findings suggest that a greater percentage of dense, poorly deformable red cells are present in sickle cell patients in the genotypic category of maximal severity. CONCLUSIONS: The combination of the beta S gene cluster haplotype and alpha-gene status correlates with both phenotypic laboratory findings (hematologic profile) and morbidity. These associations increase our ability to predict clinical severity and the future risk of major organ failure.

Anemia, Sickle Cell↗
PubMed1993

bcl-2 modulation of apoptosis induced by anticancer drugs: resistance to thymidylate stress is independent of classical resistance pathways.

The hypothesis was tested that expression of bcl-2 could provide protection against apoptosis induced by cytotoxic drugs via a mechanism which was different from the classical determinants of drug resistance. Sensitivity and resistance to inhibitors of thymidylate synthase (EC 2.1.1. 45) were chosen for study since these drugs have a well-defined and quantifiable locus of action with similarly well defined biochemical sequelae resulting from enzyme inhibition. Human lymphoma cells transfected with the vector alone readily underwent apoptosis after a 36-h exposure to various drugs. For example, 5-fluorodeoxyuridine (0.1 microM) induced 67% apoptosis in vector control cells 24 h after removal of the drug. In contrast, cells treated under identical conditions, but which expressed the bcl-2 protein, showed only basal levels of apoptosis (8%), with no significant fall in viability. Similar results were obtained using two quinazoline-based inhibitors of thymidylate synthase, N10-propargyl-5,8-dideazafolic acid (CB3717) and ICI M247496. Determinants of resistance to these three drugs were investigated. Analysis of the cell cycle, thymidylate synthase levels, and activity showed these to be unchanged by expression of bcl-2. Addition of the drugs brought about equivalent inhibition of proliferation in the presence or absence of bcl-2 expression. 5-Fluorodeoxyuridine treatment reduced TTP synthesis, induced strand breaks in nascent DNA, measured by alkaline elution, and increased the synthesis of thymidylate synthase; these changes preceded the onset of apoptosis and were identical in the vector controls and bcl-2 transfectants. Resistance to thymidylate stress in bcl-2-expressing cells therefore occurred by a mechanism different from those which classically define resistance to this type of cytotoxic drug.

Apoptosis↗
PubMed1992

Dipyridamole increases human red blood cell deformability.

An automated filtration technique has been used to investigate the effect of dipyridamole (DP) on red blood cell deformability in patients identified as having rigid red cells. They were patients on haemodialysis (HD) for chronic renal failure (n = 18), patients with peripheral vascular disease (PVD, n = 23) and controls (hospital outpatients, n = 33). Leucocytes and platelets were removed from heparinised blood by filtration through Imugard wool. Washed red cell suspensions in buffer at 5% haematocrit, without or with 5 microM DP, were filtered through Nuclepore Hemafil Membranes with 4.7 microns pores. The initial steady state relative filtration pressure (iRFP) was used to assess cell deformability. A low iRFP value reflects increased deformability and vice versa. The mean iRFP values were 0.33, 0.393 and 0.403 for controls, PVD and HD patients respectively, indicating that the red cells in the two groups of patients were significantly more rigid. DP reduced the iRFP to 0.266, 0.278 and 0.263 for controls, PVD and HD patients respectively. The results suggest that DP may be beneficial when red cell rigidity contributes to impaired microvascular perfusion.

Adult↗
PubMed1992

Circadian rhythm of white blood cell aggregation and free radical status in healthy volunteers.

Previous studies have demonstrated circadian rhythms in the onset of thrombotic events, which occur most commonly in the morning, and also in the fibrinolytic activity of the blood which has a peak in the evening and a trough in the morning. There has recently been increasing interest in the role of white blood cells (WBCs) and free radicals (FRs) in thrombosis. No one has yet investigated the potential circadian variation of WBC aggregation and FRs in humans. We studied the circadian rhythm of WBC aggregation and FR status in 10 healthy male volunteers. Six blood samples were collected at 4 hourly intervals from 12:00 (mid-day) until 08:00 the following morning. The volunteers carried out normal daily activities until 00:00 at which time they went to bed and they remained in bed until 08:00. The following were measured on each sample: WBC aggregation; thiobarbituric acid reactive substances (TBARs), plasma thiols (PSH), red cell lysate thiols (LSH), glutathione (GSH) and superoxide dismutase (SOD) which are all altered in the presence of FR activity. The following parameters demonstrated significant circadian rhythms, WBC aggregation p less than 0.001, TBARs p less than 0.015, PSH p less than 0.001, LSH p less than 0.002. WBC aggregation was lowest at 09:00 and highest at 00:00-04:00. TBARs and PSH both had a peak at 16:00 and a trough at 04:00. LSH had a peak at 12:00 and a trough at 08:00. As the behaviour of WBCs and FR status influence the flow properties of blood, a circadian rhythm in WBC function and FR status may contribute to the time of onset of thrombotic diseases. Moreover, with many studies being currently undertaken in this area, our work indicates the need to standardize sample times.

Adult↗
PubMed1992

Kinetics of granulocyte deformability following exposure to chemotactic stimuli.

In order to quantitate the kinetics of granulocyte deformability changes consequent to chemotactic stimulation, transit times (TT) of neutrophils through 8-microns diameter pores were studied via a modified cell transit analyzer (CTA). Cells isolated from normal human blood were tested at 20-second intervals for 5 minutes following stimulation with N-formyl-methionyl-leucyl-phenyl-alanine (fMLP) or zymosan-activated plasma (ZAP). The effects of cytochalasin B (CB), N-ethylmaleimide (NEM), and ibuprofen were also evaluated. Salient results included: (1) greater TT with increasing fMLP concentration (0.01-100 nM) with a peak response at 40-60 seconds followed by a return to or toward control at 5 minutes; (2) longer TT at 60 seconds for at least 75% of the cells at all fMLP concentrations, yet for 1 nM at 5 minutes nearly one half had TT less than unstimulated cells; and (3) similar temporal responses during a 5-minute period to ZAP simulation, with a nonlinear relation between cell rigidity and F-actin content. CB (20 microM) and NEM (1 mM) caused an immediate 30% to 35% decrease of TT for unstimulated cells; ibuprofen (10-1000 micrograms/ml) did not affect unstimulated TT, yet significantly reduced the response to fMLP and ZAP. Cell volume, as judged by CTA pulse height, decreased following fMLP stimulation, thus indicating that cell swelling does not contribute to the longer pore transit times of activated granulocytes. Our results, combined with literature reports describing microvascular occlusion by neutrophils, strongly suggest the importance of kinetic rather than static studies of granulocyte deformability; further, they indicate the usefulness of the modified CTA for such measurements.

Actins↗
PubMed1991

Effect of dipyridamole alone and in combination with aspirin on whole blood platelet aggregation, PGI2 generation, and red cell deformability ex vivo in man.

STUDY OBJECTIVE: The aim was to investigate the effects of dipyridamole, aspirin, and a combination of dipyridamole plus aspirin on platelet aggregation in whole blood, PGI2 generation, and red cell deformability ex vivo. SUBJECTS: were 16 male volunteers, aged 22-39 years, mean age, 26.6 years. DESIGN: This was a randomised, double blind, placebo controlled trial. The volunteer received each of the following treatments 10 days apart: dipyridamole 200 mg; aspirin 300 mg; dipyridamole 200 mg plus aspirin 300 mg; matched placebos. MEASUREMENTS AND MAIN RESULTS: Blood was taken for platelet function tests, PGI2 metabolite assay, and red cell deformability before and 2 h after the trial dose was taken. Platelet aggregation was quantified by measuring the fall in single platelet count after stimulation with 2 micrograms.ml-1 collagen or 50 nM platelet activating factor (PAF), or by rollermixing aliquots of blood to initiate spontaneous aggregation. The platelet function tests were completed at 37 degrees C within 10 min of venepuncture. The stable metabolite of PGI2, 6-keto PGF1 alpha, was measured in serum. There was inhibition of spontaneous platelet aggregation by dipyridamole (p less than 0.004), aspirin (p less than 0.005), and the combination of dipyridamole plus aspirin (p less than 0.0001) as compared with placebo. PAF induced platelet aggregation was inhibited by dipyridamole (p less than 0.002) and the combination of dipyridamole plus aspirin (p less than 0.0001) but aspirin alone had no inhibitory effect. Collagen induced platelet aggregation was inhibited by all three treatments: dipyridamole (p less than 0.06), aspirin (p less than 0.0001), and the combination of dipyridamole plus aspirin (p less than 0.0001). PGI2 generation was markedly inhibited by aspirin (p less than 0.0001) and the combination doses (p less than 0.0001) but was unaffected by dipyridamole alone. Of the three active treatments, only dipyridamole alone significantly (p less than 0.001) increased red cell deformability; there was a modest decrease in red cell deformability with aspirin. CONCLUSIONS: The results with PAF support the view that dipyridamole inhibits platelet activation by more than one mechanism; the effect on collagen induced and spontaneous platelet aggregation suggests that the effect of the combination doses is additive and that on red cell deformability the synergy is negative.

6-Ketoprostaglandin F1 alpha↗
PubMed1991

Circadian variation of endothelial cell function, red blood cell deformability and dehydro-thromboxane B2 in healthy volunteers.

There is a circadian variation in the time of onset of thrombotic disorders. The thrombotic tendency of blood is influenced by many factors including complex interactions between endothelial cells, platelets and red blood cells. We studied the circadian variation of various indices of these cells' functions in 10 young healthy male volunteers. Six blood samples were collected at 4 h intervals from 12.00 until 08.00 the following morning. The volunteers carried out normal daily activities until 00.00 at which time they went to bed. They remained in bed until 08.00 the following morning. The following were measured on each sample: plasma factor VIII von Willebrand factor antigen (FVIII vWF Ag), tissue plasminogen activator activity (tPA), plasminogen activator inhibitor (PAI), prostacyclin stimulating factor (PGI2 SF), fibrinopeptide A (FPA), 11 dehydro-thromboxane B2 (11-dehydro-TXB2) and red cell deformability (RCD). The following parameters demonstrated significant circadian variation: tPA P less than 0.001, PAI P less than 0.04 and 11-dehydro-TXB2 P less than 0.005 (two-way analysis of variance). tPA was highest at 16.00-20.00 and lowest at 08.00; PAI was highest at 08.00 and lowest at 20.00; TXB2 had a peak at 16.00 and trough at 04.00. As the behaviour of endothelial cells and platelets influence the rheological properties of blood, circadian variations in their behaviour may contribute to the time of onset of thrombotic disorders.

Adult↗
PubMed1990

P47 phosphoprotein of blood platelets (pleckstrin) is a major target for phorbol ester-induced protein phosphorylation in intact platelets, granulocytes, lymphocytes, monocytes and cultured leukaemic cells: absence of P47 in non-haematopoietic cells.

Aggregating agents including phorbol esters which activate protein kinase C induce the rapid phosphorylation of a Mr = 47,000 cytosolic protein in blood platelets (P47 or pleckstrin). This protein is well resolved by analytical 16-BAC----SDS two-dimensional PAGE and was purified from platelets by preparative 16-BAC----SDS PAGE. Polyclonal antibodies were raised to the protein in mice and rabbits. These antisera detected a single protein with the migration of P47 on Western blots of platelet extracts, and the rabbit antisera immunoprecipitated 32P-labelled P47 from platelet cytosol. The presence of P47 in other haematopoietic cells was determined by prelabelling them with 32P and observing increased 32P incorporation into the location of P47 on autoradiographs of 16-BAC----SDS analytical PAGE of cells exposed to phorbol ester. The identity of the phosphoprotein found in this location was further established by probing Western blots of SDS PAGE gels of cultured cell lines with the P47 antisera. P47 was detected in peripheral blood lymphocytes, monocytes and granulocytes (including the granulocytes of two unrelated patients with X-linked chronic granulomatous disease). P47 was also found in HL-60 promyelocytes (especially after differentiation with retinoic acid), U937 histiocytes, HEL leukaemia cells, and Raji 'B' lymphoblasts. It was not detected in normal erythrocytes, K562 leukaemic cells, MOLT-3 'T' lymphoblasts, or in wide range of non-haematopoietic cell lines. We conclude that P47 is a major target for the action of phorbol ester induced phosphorylation in platelets, normal leucocytes and some haematopoietic cell lines. These cells have as their common feature the ability when stimulated to develop adhesive functions on their plasma membranes.

Blood Platelets↗
PubMed1989

Human whole-blood granulocyte aggregation in vitro.

1. Aggregation assays are a commonly used technique for the study of granulocyte activation. These studies are usually performed using a pure cell suspension in buffer. This necessitates a separation procedure which is time-consuming and may modify the function of the cells. Interaction between different cell types is precluded. 2. To avoid these disadvantages a method was developed which quantifies granulocyte aggregation in whole blood. Samples drawn from an incubated vessel before and after the addition of a chemotactic stimulus were fixed with formaldehyde to prevent disaggregation. Erythrocytes were then removed by chemical lysis and using an electronic cell-sizing device the number of single cells and aggregates could then be easily measured. 3. Results from a group of volunteers showed a rapid and reversible response to a chemotactic tripeptide, with a fall in single granulocyte count and the appearance of doublets and triplets. Lymphocytes were unaffected. Intra-assay reproducibility was better than +/- 5%. 4. Using this assay, a significant elevation in aggregability was observed in blood from patients after acute myocardial infarction. 5. This novel technique, by avoiding the separation step, is faster, simpler and more physiological than previous methods, and as such is useful for both assays of drug action in vitro and the study of cell activation in disease states.

Adult↗
PubMed1986

Cytogenetic analysis of adult acute lymphoblastic leukemia including a Ph+ case surviving more than 5 years.

We have performed cytogenetic analysis on 25 consecutive adult patients with previously untreated acute lymphoblastic leukemia (ALL) who were subsequently treated with the same protocol at this institution. Ten of the 25 patients studied (40%) demonstrated karyotypic abnormalities. The most frequent abnormalities were hyperdiploidy (six patients) and presence of the Philadelphia (Ph) chromosome (three patients). Univariate analysis of 12 features identified only immunophenotype as differing between patients with abnormal and normal karyotype. The cells of patients with an abnormal karyotype were more often non-B, non-T and less often T cell in phenotype. One patient initially with Ph remains cytogenetically normal in complete remission 272 weeks post diagnosis. We confirm that cytogenetic abnormalities are frequent in adult ALL. The attainment of disease free survival in Ph-positive ALL of more than 5 years with persistently normal cytogenetics demonstrates that aggressive multimodal therapy can induce long-term remissions and possible cure of this usually unfavorable situation.

Adolescent↗
PubMedDate not supplied

The influence of suspending phase viscosity on the passage of red blood cells through capillary-size micropores.

Much attention has been paid to the study of blood flow in long, narrow tubes. While the influence of tube diameter and driving pressure have been examined in detail, the influence of suspending phase viscosity has generally been assumed only to affect the blood viscosity in a linearly proportional manner, hence the practice of normalizing apparent blood viscosity values by the suspending phase viscosity to give a relative viscosity (e.g., Pries et al., 1992). While this assumption is probably valid for long tubes, it apparently does not hold for blood flow in short tubes (and by extension also for flow in short or branching capillary segments in vivo) in which RBC deformation plays a more significant role. In this paper we present a series of experiments using the Cell Transit Analyzer (CTA) in which the influence of driving pressure and suspending phase viscosity on RBC passage through short, narrow tubes has been systematically evaluated. Over the range studied (1 to 10 cm water), the influence of driving pressure was found to be unremarkable, in that RBC velocity scaled directly and linearly with pressure. This finding is consistent with previous studies. However, a distinct intercept was observed in the linear relationship between RBC pore transit time and suspending phase viscosity, which presumably arises as a consequence of RBC deformation either at the pore entrance or within the pore. Two simple mathematical models for the suspending phase-viscosity/transit-time relationship were considered. The results show that making CTA measurements over a range of suspending medium viscosities is a simple and practical way to obtain additional information about RBC mechanical properties.

Blood Pressure↗
PubMedDate not supplied

Determination of particle sedimentation rate by ultrasonic interferometry: role of particle size, density and volume fraction.

The sedimentation rate (SR) of non-aggregated spherical particles in suspension was determined using an ultrasonic interferometry technique (Echo-Cell); this method is based on A-mode echography and measures the rate of formation of a sediment on a solid plate during settling. The particle accumulation rate, which is related to SR, is obtained from the interference of two waves reflected by two interfaces: one between the plate and the sediment and the other between the sediment and the suspension. Studies were carried out at 25 degrees C using latex spheres of different diameters (7 to 20 micron) and densities (1.062 to 1.190 g/cm3) suspended in distilled water at various volume fractions (1% to 5%). As anticipated by the Stokes model, linear relations were found between SR and both particle density and the square of particle radius. Experimental SR values decreased with increasing suspension particle concentration; these concentration effects were in good agreement with those predicted by the Steinour model. Our results thus serve to validate the theoretical aspects of the Echo-Cell method and suggest its usefulness as a tool for studies of RBC interaction and RBC aggregation.

Blood Sedimentation↗
PubMedDate not supplied

Pulse shape analysis of RBC micropore flow via new software for the cell transit analyser (CTA).

The Cell Transit Analyser (CTA) provides a means to rapidly measure the deformability of large numbers of individual cells. It combines many of the advantages of micropipette studies with the simplicity and speed of filtrometry methods by measuring the duration of each resistive pulse generated as a cell passes through one of 30 identical micropores in a membrane. However, in our opinion, the potential of the system is limited by the microcomputer and software supplied for data analysis. We have therefore written new software for a more-powerful microcomputer to examine the shape of each resistive pulse rather than just the duration. Seven new parameters are derived, which provide additional information regarding the passage of cells through the pores. In particular, the contribution of the entry and exit phases of the cell transit are evident in the rise time and fall time of the pulses. The software is user-friendly and allows the analysis of each pulse to be reviewed, which aids understanding of the system and helps to avoid errors in interpreting the data.

Adult↗