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Biomedical subjects

D J Fitzgerald

Publications and source records attributed to D J Fitzgerald.

At least 19 recordsLinked to original sources
PubMed2000

The platelet integrin alpha IIbbeta 3 has an endogenous thiol isomerase activity.

Integrins are cysteine-rich heterodimeric cell-surface adhesion molecules that alter their affinity for ligands in response to cellular activation. The molecular mechanisms involved in this activation of integrins are not understood. Treatment with the thiol-reducing agent, dithiothreitol, can induce an activation-like state in many integrins suggesting that cysteine-cysteine dithiol bonds are important for the receptor's tertiary structure and may be involved in activation-induced conformational changes. Here we demonstrate that the platelet-specific integrin, alpha(IIb)beta(3), contains an endogenous thiol isomerase activity, predicted from the presence of the tetrapeptide motif, CXXC, in each of the cysteine-rich repeats of the beta(3) polypeptide. This motif comprises the active site in enzymes involved in disulfide exchange reactions, including protein-disulfide isomerase (EC ) and thioredoxin. Intrinsic thiol isomerase activity is also observed in the related integrin, alpha(v)beta(3), which shares a common beta-subunit. Thiol isomerase activity within alpha(IIb)beta(3) is time-dependent and saturable, and is inhibited by the protein-disulfide isomerase inhibitor, bacitracin. Furthermore, this activity is calcium-sensitive and is regulated in the EDTA-stabilized conformation of the integrin. This novel demonstration of an enzymatic activity associated with an integrin subunit suggests that altered thiol bonding within the integrin or its substrates may be locally modified during alpha(IIb)beta(3) activation.

Amino Acid Sequence↗
PubMed2000

Cyclooxygenase-1 and -2-dependent prostacyclin formation in patients with atherosclerosis.

BACKGROUND: The formation of prostacyclin (PGI(2)), thromboxane (TX) A(2), and isoprostanes is markedly enhanced in atherosclerosis. We examined the relative contribution of cyclooxygenase (COX)-1 and -2 to the generation of these eicosanoids in patients with atherosclerosis. METHODS AND RESULTS: The study population consisted of 42 patients with atherosclerosis who were undergoing surgical revascularization. COX-2 mRNA was detected in areas of atherosclerosis but not in normal blood vessel walls, and there was evidence of COX-1 induction. The use of immunohistochemical studies localized the COX-2 to proliferating vascular smooth muscle cells and macrophages. Twenty-four patients who did not previously receive aspirin were randomized to receive either no treatment or nimesulide at 24 hours before surgery and then for 3 days. Eighteen patients who were receiving aspirin were continued on a protocol of either aspirin alone or a combination of aspirin and nimesulide. Urinary levels of 11-dehydro-TXB(2) and 2,3-dinor-6-keto-PGF(1alpha), metabolites of TXA(2) and PGI(2), respectively, were elevated in patients with atherosclerosis compared with normal subjects (3211+/-533 versus 679+/-63 pg/mg creatinine, P<0.001; 594+/-156 versus 130+/-22 pg/mg creatinine, P<0.05, respectively), as was the level of the isoprostane 8-iso-PGF(2alpha). Nimesulide reduced 2, 3-dinor-6-keto-PGF(1alpha) excretion by 46+/-5% (378.3+/-103 to 167+/-37 pg/mg creatinine, P<0.01) preoperatively and blunted the increase after surgery. Nimesulide had no significant effect on 11-dehydro-TXB(2) before (2678+/-694 to 2110+/-282 pg/mg creatinine) or after surgery. The levels of both products were lower in patients who were taking aspirin, and no further reduction was seen with the addition of nimesulide. None of the treatments influenced urinary 8-iso-PGF(2alpha) excretion. CONCLUSIONS: Both COX-1 and -2 are expressed and contribute to the increase in PGI(2) in patients with atherosclerosis, whereas TXA(2) is generated by COX-1.

6-Ketoprostaglandin F1 alpha↗
PubMed2000

Surface expression of glycoprotein ib alpha is dependent on glycoprotein ib beta: evidence from a novel mutation causing Bernard-Soulier syndrome.

Bernard-Soulier syndrome is a rare bleeding disorder caused by a quantitative or qualitative defect in the platelet glycoprotein (GP) Ib-IX-V complex. The complex, which serves as a platelet receptor for von Willebrand factor, is composed of 4 subunits: GPIb alpha, GPIb beta, GPIX, and GPV. We here describe the molecular basis of a novel form of Bernard-Soulier syndrome in a patient in whom the components of the GPIb-IX-V complex were undetectable on the platelet surface. Although confocal imaging confirmed that GPIb alpha was not present on the platelet surface, GPIb alpha was readily detectable in the patient's platelets. Moreover, immunoprecipitation of plasma with specific monoclonal antibodies identified circulating, soluble GPIb alpha. DNA-sequence analysis revealed normal sequences for GPIb alpha and GPIX. There was a G to A substitution at position 159 of the gene encoding GPIb beta, resulting in a premature termination of translation at amino acid 21. Studies of transient coexpression of this mutant, W21stop-GPIb beta, together with wild-type GPIbalpha and GPIX, demonstrated a failure of GPIX expression on the surface of HEK 293T cells. Similar results were obtained with Chinese hamster ovary alpha IX cells, a stable cell line expressing GPIbalpha that retains the capacity to re-express GPIX. Thus, we found that GPIbbeta affects the surface expression of the GPIb-IX complex by failing to support the insertion of GPIb alpha and GPIX into the platelet membrane. (Blood. 2000;96:532-539)

Animals↗
PubMed2000

Glycoprotein IIb/IIIa antagonists induce apoptosis in rat cardiomyocytes by caspase-3 activation.

The platelet integrin glycoprotein (GP) IIb/IIIa, which mediates platelet aggregation, has been the target for novel antiplatelet agents, the GPIIb/IIIa antagonists. Several GPIIb/IIIa antagonists have been developed based on the peptide RGDS present in adhesion proteins, including the principle ligand fibrinogen. The apoptosis enzyme, procaspase-3, contains an RGD-recognition sequence and is activated by RGDS. We examined the effects of RGDS and several GPIIb/IIIa antagonists on cell death and procaspase-3 activation in rat neonatal cardiomyocytes. These antagonists do not recognize rat integrins, yet RGDS, orbofiban, and xemilofiban induced dose-dependent apoptosis and procaspase-3 activation in cardiomyocytes over 72 h, particularly under hypoxic conditions. Scrambled peptide, the monoclonal antibody 7E3 or integrelin (a peptide containing a KGD sequence), had little or no effect. Immunoprecipitation of procaspase-3 followed by treatment with the compounds showed that procaspase-3 was activated directly by RGDS, orbofiban, xemilofiban, and by monoclonal 7E3 antibody, the latter demonstrating that compounds must enter cells to induce apoptosis through caspase activation. Integrelin had no effect. Binding studies with (3)H-SC52012B, a GPIIb/IIIa antagonist analogue of orbofiban, showed no specific binding to cardiomyocytes, but the radioligand accumulated intracellularly over 72 h. (3)H-SC52012B also bound directly to human recombinant caspase-3 (K(d), 59 +/- 2 nm), and this was prevented by orbofiban, xemilofiban, and the monoclonal 7E3 antibody but not by integrelin. Finally confocal microscopy showed that RGDS co-localized with caspase-3 inside the cell. These data show that RGDS and its mimetics induce cardiomyocyte apoptosis by direct activation of procaspase-3.

Alanine↗
PubMed2000

Streptokinase-induced platelet activation involves antistreptokinase antibodies and cleavage of protease-activated receptor-1.

Streptokinase activates platelets, limiting its effectiveness as a thrombolytic agent. The role of antistreptokinase antibodies and proteases in streptokinase-induced platelet activation was investigated. Streptokinase induced localization of human IgG to the platelet surface, platelet aggregation, and thromboxane A(2) production. These effects were inhibited by a monoclonal antibody to the platelet Fc receptor, IV.3. The platelet response to streptokinase was also blocked by an antibody directed against the cleavage site of the platelet thrombin receptor, protease-activated receptor-1 (PAR-1), but not by hirudin or an active site thrombin inhibitor, Ro46-6240. In plasma depleted of plasminogen, exogenous wild-type plasminogen, but not an inactive mutant protein, S(741)A plasminogen, supported platelet aggregation, suggesting that the protease cleaving PAR-1 was streptokinase-plasminogen. Streptokinase-plasminogen cleaved a synthetic peptide corresponding to PAR-1, resulting in generation of PAR-1 tethered ligand sequence and selectively reduced binding of a cleavage-sensitive PAR-1 antibody in intact cells. A combination of streptokinase, plasminogen, and antistreptokinase antibodies activated human erythroleukemic cells and was inhibited by pretreatment with IV.3 or pretreating the cells with the PAR-1 agonist SFLLRN, suggesting Fc receptor and PAR-1 interactions are necessary for cell activation in this system also. Streptokinase-induced platelet activation is dependent on both antistreptokinase-Fc receptor interactions and cleavage of PAR-1. (Blood. 2000;95:1301-1308)

Adenosine Diphosphate↗
PubMed2000

Prospective evaluation of the risk conferred by factor V Leiden and thermolabile methylenetetrahydrofolate reductase polymorphisms in pregnancy.

Factor V (FV) Leiden and thermolabile methylenetetrahydrofolate reductase (MTHFR) are 2 common polymorphisms that have been implicated in vascular thrombosis. We determined whether these mutations predicted an adverse outcome in pregnancy. Second, we looked for an interaction between these 2 mutations in patients with recurrent fetal loss or thrombosis in pregnancy. Primigravid subjects at their booking visit to the National Maternity Hospital (Holles Street, Dublin, Ireland) were screened for the polymorphisms. Thermolabile MTHFR and FV Leiden genotypes were detected by either restriction fragment length polymorphism or heteroduplex capillary chromatography. The carrier frequency of FV Leiden in the screened primigravid population was 2.7% (allele frequency 1.36%), all being heterozygous for the mutation. This value was lower than expected from previous studies in European populations. Forty-nine percent of the screened population (289 of 584) were heterozygous for thermolabile MTHFR, and 10.6% were homozygous (62 of 584). The frequency of the 2 polymorphisms was no higher in those who subsequently developed preeclampsia (n=12) or intrauterine growth retardation (n=9), and none of the screened population developed thrombosis. However, the frequency of FV Leiden was higher in patients who subsequently miscarried after the first trimester of pregnancy (allele frequency of 5.5%, P=0.0356). Among those positive for FV Leiden, 3 of 27 miscarried, compared with 24 of 572 of FV Leiden-negative patients (11% versus 4.2%). No interaction was found between the 2 mutations in the control or patient populations. In patients with a prior history of venous thrombosis, the carrier rate of FV Leiden was increased (4 of 33, allele frequency of 7.6%, P=0. 0115). In contrast, the carrier frequency for thermolabile MTHFR was no higher, and there was no interaction between the 2 mutations. Neither mutation occurred at a significantly higher frequency in patients with a prior history of recurrent fetal loss. In conclusion, FV Leiden is a risk factor for thrombosis in pregnancy and possibly for second-trimester miscarriage independent of thermolabile MTHFR. However, prospective analysis suggests that the risk conferred by FV Leiden is low in a primigravid population. The thermolabile MTHFR genotype was not implicated in any adverse outcome.

Abortion, Habitual↗
PubMed2000

An automated heteroduplex assay for the Pi(A) polymorphism of glycoprotein IIb/IIIa, multiplexed with two prothrombotic genetic markers.

Screening for the PIA polymorphism has been made faster and simpler with the advent of heteroduplex technology. Simultaneous screening for three common prothrombotic polymorphisms pl(A), factor V Leiden, and MTHFR(C677T) has been achieved with multiplex heteroduplex analysis. We describe a quick and simple method for PlA heteroduplex probe production. The probe was multiplexed with heteroduplex probes for MTHFR(C677T) and factor V Leiden polymorphisms in a one tube assay, allowing rapid automated genotyping of all three. This automated multiplex assay was applied to a cohort of 165 patients and showed excellent correlation with gel-based assays, both PAGE and RFLP. This approach will facilitate the analysis of multiple polymorphisms in complex disease in large populations.

Cohort Studies↗
PubMed1999

DNA distortion as a factor in nucleosome positioning.

In a previous report we constructed a synthetic DNA sequence that directed the deposition of histone octamers to a single site, and it was proposed that DNA distortion was involved in the positioning effect. In the present study we utilized the chemical probe potassium permanganate to identify sites of DNA distortion in the synthetic positioning sequence. A permanganate hypersite was identified 15 bp from the nucleosome pseudo-dyad at a site known to display DNA distortion in the mature nucleosome. The sequence of the site contained a TA step flanked by an oligo-pyrimidine tract. A series of substitutions were made in the region of the permanganate hypersite and the resulting constructs tested for affinity for histone octamers and translational positioning in in vitro studies. The results revealed that either a single base substitution at the TA step or in the adjacent homopolymeric tract dramatically affected affinity and positioning activity. The rotational orientation of the permanganate-sensitive sequence was shown to be important for functions, since altering the orientation of the site in a positioning fragment reduced positioning activity and octamer affinity, while altering the rotational orientation of the sequence in a non-positioning fragment had the opposite effects. A reconstituted 5 S rDNA positioning sequence from Lytechinus variegatus was also shown to display a permanganate hypersite 16 bp from its pseudo-dyad.

Animals↗
PubMed1999

Ticlopidine and clopidogrel.

The thienopyridines ticlopidine and clopidogrel are inhibitors of platelet function in vivo. Their mode of action has not been defined, but it appears that they require conversion to as yet unidentified metabolites that are noncompetitive antagonists of the platelet ADP receptor. Inhibition of platelet aggregation with these compounds is delayed until 24 to 48 hours after administration. Maximum inhibition occurs after 3 to 5 days, and recovery is slow after drug withdrawal. Ticlopidine is effective in preventing cardiovascular events in cerebrovascular, cardiovascular, and peripheral vascular disease, with an efficacy that is similar to aspirin. However, its use is associated with significant and sometimes fatal adverse reactions, specifically neutropenia and bone marrow aplasia. Gastrointestinal side effects and skin rashes are common and result in discontinuation of therapy in up to 10% of patients. Clopidogrel is at least as effective as aspirin in preventing cardiovascular events in patients with a history of vascular disease. It appears to be safer than ticlopidine, although its efficacy in acute coronary syndromes or post-coronary-stent insertion has not been reported. Important outstanding issues are whether clopidogrel adds to the benefit of aspirin and whether the combination of these agents is safe. If so, this combination may become the standard for antithrombotic therapy in cardiovascular disease.

Adenosine Diphosphate↗
PubMed1999

The relationship between cyclooxygenase-2 expression and colorectal cancer.

CONTEXT: Epidemiological studies have implicated the inducible form of cyclooxygenase (COX-2) in the pathogenesis of colorectal cancer; however, its role is not fully understood. OBJECTIVE: To examine the relationship between the expression of COX-2 in human colorectal cancer and patient survival. DESIGN: Patients diagnosed as having colorectal cancer were evaluated and followed up for up to 9.4 years (median follow-up, 2.7 years). Tumor sections were stained for COX-2 using a rabbit polyclonal antibody raised against human COX-2. The extent of COX-2 staining was graded by 2 observers blinded to outcome. Preabsorption of the anti-COX-2 antibody with a COX-2 peptide abolished the staining, demonstrating the specificity of the assay. SETTING: Gastrointestinal unit of a large general teaching hospital in Dublin, Ireland. PARTICIPANTS: Seventy-six patients (median age, 66.5 years) with colorectal cancer (Dukes tumor stage A, n = 9; Dukes B, n = 30; Dukes C, n = 25; Dukes D, n = 12) whose diagnosis was made between 1988 and 1991. Fourteen normal colon biopsies were stained for COX-2 as controls. MAIN OUTCOME MEASURES: Survival in years following diagnosis compared by extent of COX-2 epithelial staining (grade 1, <1%; grade 2, 1%-19%; grade 3, 20%-49%; grade 4, > or = 50%), Dukes stage, tumor size, and lymph mode metastasis. RESULTS: COX-2 was found in tumor epithelial cells, inflammatory cells, vascular endothelium, and/or fibroblasts. The extent of epithelial staining was heterogeneous, varying markedly among different tumors. Normal tissue adjacent to the tumors also stained weakly for COX-2. No COX-2 was detected in control tissue samples. The Kaplan-Meier survival estimate was 68% in patients who had grade 1 tumor epithelial staining compared with 35% in those with higher grades combined (log-rank chi2 = 5.7; P = .02). Greater expression of COX-2 correlated with more advanced Dukes stage (Kendall tau-b, 0.22; P = .03) and larger tumor size (Kendall tau-b, 0.21; P = .02) and was particularly evident in tumors with lymph node involvement (Kendall tau-b, 0.26; P = .02). CONCLUSIONS: Our data indicate that COX-2 expression in colorectal cancer may be related to survival. These data add to the growing epidemiological and experimental evidence that COX-2 may play a role in colorectal tumorigenesis.

Aged↗
PubMed1999

Selective nucleosome disruption by drugs that bind in the minor groove of DNA.

Previous studies have shown that drugs which bind in the DNA minor groove reduce the curvature of bent DNA. In this article, we examined the effects of these drugs on the nucleosome assembly of DNA molecules that display different degrees of intrinsic curvature. DAPI (4,6-diamidino-2-phenylindole) inhibited the assembly of a histone octamer onto a 192-base pair circular DNA fragment from Caenorhabditis elegans and destabilized a nucleosome that was previously assembled on this segment. The inhibitory effect was highly selective since it was not seen with nonbent molecules, bent molecules with noncircular shapes, or total genomic DNA. This marked template specificity was attributed to the binding of the ligand to multiple oligo A-tracts distributed over the length of the fragment. A likely mechanism for the effect is that the bound ligand prevents the further compression of the DNA into the minor groove which is required for assembly of DNA into nucleosomes. To further characterize the effects of the drug on chromatin formation, a nucleosome was assembled onto a 322-base pair DNA fragment that contained the circular element and a flanking nonbent segment of DNA. The position of the nucleosome along the fragment was then determined using a variety of nuclease probes including exonuclease III, micrococcal nuclease, DNase I, and restriction enzymes. The results of these studies revealed that the nucleosome was preferentially positioned along the circular element in the absence of DAPI but assembled onto the nonbent flanking sequence in the presence of the drug. DAPI also induced the directional movement of the nucleosome from the circular element onto the nonbent flanking sequence when a nucleosome preassembled onto this template was exposed to the drug under physiologically relevant conditions.

Animals↗
PubMed1999

Heterogeneity in the actions of drugs that bind in the DNA minor groove.

Distamycin and Hoechst 33258 have long served as the model compounds for biochemical, biophysical, and clinical studies of the drugs that bind in the DNA minor groove. However, the results presented in this investigation clearly show that 4,6-diamidino-2 phenylindole (DAPI) is superior to both of these drugs at negating the effects of intrinsic DNA curvature and anisotropic bendability as measured by electrophoretic and ligation analysis. In addition, DAPI was more effective than distamycin and Hoechst 33258 at inhibiting the assembly of nucleosomes onto synthetic and natural sequences that have multiple closely spaced oligo-AT sequences that serve as drug binding sites. Since these effects may be related to the biological action of the drugs, it was of interest to determine the mechanism that was responsible for the enhanced action of DAPI. The possibility that the differential drug potencies resulted from differential overall affinities of the ligands for A-tract molecules was considered, but drug binding studies suggested that this was not the case. It is also unlikely that the differential drug effects resulted from the binding of the drugs to different DNA sites since the oligo A/T binding sites for DAPI and Hoechst were centered on the same nucleotide positions as revealed by footprinting studies using exonuclease III, DNase I, and hydroxyl radical. However, the footprinting studies with DNase I did uncover a potentially important difference between the drugs. DAPI protected only the AT bp in the binding sites, while distamycin and Hoechst protected these bp as well as flanking Gs and Cs. These results permitted us to advance a preliminary model for the enhanced action DAPI. According to the model, the short length of DAPI and its absolute specificity for A/T bps with narrow minor grooves ensures that only particularly minor grooves that give rise to curvature and anisotropic bendability are occupied by the drug. Consequently, each helical deflection induced by an A-tract in the absence of the drug is countered by an opposite deflection induced by DAPI binding, thus effectively neutralizing intrinsic curvature and bending into the minor groove.

Animals↗
PubMed1999

Oxidative damage of cardiomyocytes is limited by extracellular regulated kinases 1/2-mediated induction of cyclooxygenase-2.

Oxidative stress causes cardiac damage following ischemia/reperfusion and in response to anthracyclines. Extracellular signal-regulated kinases (ERK) 1/2 are activated by oxidative stress in cardiac myocytes and protect cardiac myocytes from apoptosis. Prostaglandins (PG) also protect cells from injury in a number of tissues, including the cardiomyocyte. Cyclooxygenase (COX) the rate-limiting enzyme in PG biosynthesis has two isoforms, the constitutive COX-1 and an inducible COX-2. Here, we examined the effects of two oxidative stresses, hydrogen peroxide (H2O2) and the anthracycline doxorubicin on the activity of ERK1/2 and the expression of COX isoforms and PG formation in neonatal rat primary cardiomyocytes. These cells expressed COX-1 at rest and both COX isoforms on treatment with phorbol 12-myristate 13-acetate. Exposure to 50 microM H2O2 for 10 min or doxorubicin at 10 and 100 micrograms/ml caused expression of COX-2 that was prevented by free radical scavengers. COX-2 induction was associated with activation of ERK1/2 and the specific ERK-inhibitor PD098059 abolished COX-2 expression. Treatment of cells with decoy oligonucleotides corresponding to COX-2 promoter elements implicated the AP-1 and NF-kappaB-2 but not the NF-kappaB-1 in the transcription of COX-2. Induction of COX-2 mRNA and protein was accompanied by increased prostacyclin formation, which was abolished by the selective COX-2 inhibitor, NS-398, and PD098059. H2O2 and doxorubicin enhanced the release of lactate dehydrogenase and free radical scavengers prevented this. NS-398 enhanced the release of lactate dehydrogenase in response to H2O2 and doxorubicin, whereas the injury was prevented by iloprost, a stable prostacyclin analogue. In cardiomyocytes cell injury by H2O2 and doxorubicin is limited by an increase in prostacyclin formation that reflects induction of COX-2 mediated by ERK1/2 activation.

Animals↗
PubMed1999

The in vivo assessment of nimesulide cyclooxygenase-2 selectivity.

In man, nimesulide selectively inhibits cyclooxygenase-2 (COX-2) with little effect on haemostatic function or gastric prostaglandin formation. It causes significantly less gastrointestinal injury than naproxen, but has anti-inflammatory efficacy similar to that of naproxen and other currently available non-steroidal anti-inflammatory drugs. Naproxen suppressed arachidonic-acid-mediated platelet aggregation, reduced serum thromboxane B2 levels by 98% throughout the treatment period and reduced gastric mucosal prostaglandins (PGE2 and 6-keto-PGF1alpha) production by an average of 80%. This contrasts with nimesulide: platelet aggregation was not significantly affected, thromboxane B2 levels were only 29% lower and the gastric mucosal prostaglandins were inhibited in the order of approximately 20%. During the treatment period, both nimesulide and naproxen significantly inhibited COX-2-dependent PGE2 synthesis in the whole blood; however, naproxen had a lesser effect than nimesulide.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed1999

Stability of the blood lactate-heart rate relationship in competitive athletes.

INTRODUCTION: The identification of the HR (or RPE) associated with blood lactate concentrations of 2.5 mmol x L(-1)(aerobic threshold) (AerT) and 4.0 mmol x L(-1)(anaerobic threshold) (AnT) is a common method for defining training intensities. It is often assumed that the HR at AerT and AnT changes with changes in fitness, much as the power output (Watts: W) associated with AerT and AnT is known to change. METHODS: We studied speed skaters (N = 13, 7 male, 6 female) during spring (deconditioned) and fall (conditioned) evaluations, using cycle ergometry (stage duration = 5 min) to determine W, HR, and RPE at AerT, AnT, and at maximal exercise (3000 (female) and 5000 (male) m cycle time trials). RESULTS: In the spring vs. fall evaluations, the power output at AerT was 127+/-12 vs 162+/-9 W (P<0.05), at AnT was 216+/-14 vs. 230+/-13 W (P<0.05), and at maximal exercise was 341+/-15 vs. 364+/-19 W (P<0.05); HR at AerT was 129+/-6 vs. 130+/-7 bpm (P>0.05), at AnT was 162+/-7 vs. 164+/-7 bpm (P>0.05), and at maximal exercise was 196+/-6 vs. 198+/-5 bpm (P>0.05); RPE at AerT was 2.7+/-0.9 vs. 2.6+/-0.8 (P>0.05), at AnT was 5.3+/-1.0 vs. 5.3+/-0.9 (P>0.05). CONCLUSIONS: These data suggest that although power output at AerT, AnT, and maximal exercise changes significantly with conditioning, there is no systematic change in the associated values for HR and/or RPE used as practical markers of training intensity. Accordingly, a single well-conducted evaluation may allow evaluation of appropriate training markers that may be longitudinally stable.

Anaerobic Threshold↗
PubMed1999

Left ventricular function during interval and steady state exercise.

PURPOSE: Interval training (INT) is a commonly used method of exercise training in both athletic and clinical populations. Although we generally understand left ventricular (LV) function during steady state (SS) exercise, there are no data regarding LV function during INT. METHODS: We studied eight healthy, physically active volunteers during upright cycle ergometry during 15 min of both SS and INT, at the same average power output (90% individual anaerobic threshold), using first pass radionuclide ventriculography. During INT (60s/60s), measures of LV function were made during work (220 W) after 4 and 12 min and during recovery (120 W) after 7 and 15 min. These were compared with the average of four temporally matched measures made during SS (170 W). RESULTS: During INT, LV ejection fraction increased from rest (67 +/- 6%) to 77 +/- 5, 80 +/- 5, 77 +/- 5 and 79 +/- 4% after 4, 7, 12, and 15 min, respectively. During SS, LV ejection fraction was not significantly different at rest (70 +/- 4%) or during exercise (76 +/- 4, 79 +/- 4, 80 +/- 3, and 81 +/- 3%) after 4, 7, 12, and 15 min, respectively. Other measures of LV function (HR, BP, LV volumes, cardiac output, systemic vascular resistance, peak emptying, and filling rates) were likewise similar during temporally matched measurements during INT and SS. CONCLUSIONS: Although there were the expected transitions of ejection fraction with work and recovery, the overall hemodynamic picture during INT was very similar to SS. These data suggest that LV function during INT is not substantially different to that during SS.

Aged↗
PubMed1998

Unique translational positioning of nucleosomes on synthetic DNAs.

A computational study was previously carried out to analyze DNA sequences that are known to position histone octamers at single translational sites. A conserved pattern of intrinsic DNA curvature was uncovered that was proposed to direct the formation of nucleosomes to unique positions. The pattern consists of two regions of curved DNA separated by preferred lengths of non-curved DNA. In the present study, 11 synthetic DNAs were constructed which contain two regions of curved DNA of the form [(A5.T5)(G/C)5]4 separated by non-curved regions of variable length. Translational mapping experiments of in vitro reconstituted mononucleosomes using exonuclease III, micrococcal nuclease and restriction enzymes demonstrated that two of the fragments positioned nucleosomes at a single site while the remaining fragments positioned octamers at multiple sites spaced at 10 base intervals. The synthetic molecules that positioned nucleosomes at a single site contain non-curved central regions of the same lengths that were seen in natural nucleosome positioning sequences. Hydroxyl radical and DNase I digests of the synthetic DNAs in reconstituted nucleosomes showed that the synthetic curved element on one side of the nucleosomal dyad assumed a rotational orientation where narrow minor grooves of the A-tracts faced the histone surface with all molecules. In contrast, the curved element on the other side of the nucleosome displayed variable rotational orientations between molecules which appeared to be related to the positioning effect. These results suggest that asymmetry between the two halves of nucleosomal DNA may facilitate translational positioning.

Base Sequence↗