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Biomedical subjects

P Kierulf

Publications and source records attributed to P Kierulf.

At least 19 recordsLinked to original sources
PubMed2001

Flow cytometric evaluation of apoptosis, necrosis and recovery when culturing monocytes.

After developing and applying a method for cryopreserving monocytes, we found a substantial cell loss when culturing these cells. Monocytes were isolated from blood donors by density gradient centrifugation, purified by elutriation and cryopreserved. Thawed cells were cultured in ultra low attachment wells and studied with Annexin V, Propidium iodide, Dihexyloxacarbocyanine (DiOC(6)(3)), bromolated deoxyuridine triphosphate nucleotides (Br-dUTP), DNA ploidy and DNA ladder methodologies. The main cell loss was within the first 24 h and recovery on day 7 was 35-40%. The first 2-6 h of culture were found to be crucial for determining which cells survive. Initially (2-4 h), apoptosis was the main feature but after 6 h, necrosis dominated. Two populations of cells developed after 24 h: "A" consisting of larger cells with low levels of apoptosis and necrosis signals and population "B" comprising smaller cells with a high expression of necrotic but low levels of apoptotic signals. Signs of DNA fragmentation were slight. These early, dynamic changes may be important for the interpretation of experimental results when investigating monocytes in culture.

Apoptosis↗
PubMed2001

Non-mannose-capped lipoarabinomannan stimulates human peripheral monocytes to expression of the "early immediate genes" tissue factor and tumor necrosis factor-alpha.

In the present study, we have shown that stimulation of cryopreserved, human peripheral blood monocytes with the cell wall components from Gram-negative bacteria, lipopolysaccharide (LPS), and from rapid-growing Mycobacterium sp., non-mannose-capped lipoarabinomannan (AraLAM), both induce expression of the "early immediate genes" tissue factor (TF) and tumor necrosis factor-alpha (TNF-alpha). This was demonstrated both at the protein and the mRNA levels. Antibodies against the CD14 receptor could block the stimulating effects. AraLAM was a significantly weaker inducer than LPS, and we speculate that this may reside in the number of the fatty acids in the part of the molecule that interacts with the CD14/Toll-like receptors (TLR). Finally, both LPS and AraLAM activated the "early immediate genes" through translocation of the transcription factor proteins NF-kappaB/Rel and increasing the binding activity of AP-1.

Antigens, Bacterial↗
PubMed2001

Active recovery and post-exercise white blood cell count, free fatty acids, and hormones in endurance athletes.

Strenuous endurance exercise in fasted subjects is accompanied by increased plasma levels of catecholamines, leucocytosis, low insulin, and elevated plasma free fatty acids (FFA). Immediately after such exercise, plasma FFA may rise to high and potentially harmful levels, whereas the white blood cell count (WBCC) rapidly decreases towards or below baseline values. The present work investigated how active recovery (AR) for 15 min at 50% of maximal oxygen consumption (VO2max), after 60 min of uphill running at 83% of VO2max, influenced plasma FFA, lymphocyte, neutrophil, granulocyte, and monocyte count, as well as adrenaline, noradrenaline, insulin and cortisol concentrations until 120 min post-exercise. Thirteen endurance athletes participated in the study [24.2 (3.7) years, 1.82 (0.06) m, 76.7 (7.9) kg and VO2max 69.2 (6.8) ml min-1 kg-1]. In a randomized order, the subjects completed two sets of strenuous workouts, followed by either AR or complete rest in the supine position (RR). Compared with RR, AR strongly counteracted the rapid increase in plasma FFA 5 min post-exercise. The decreases in neutrophil and monocyte counts post-exercise were nullified by AR, and the cell count stayed above resting values throughout the observation period. AR also counteracted the rapid return of hormone concentration towards baseline levels. It would appear that active recovery at low intensity after strenuous exercise can maintain sufficient adrenergic activation to counteract the post-exercise drop in WBCC. However, in spite of keeping the catecholamine concentration high and insulin levels low, AR can also maintain a low plasma FFA concentration, probably because of the continued use of FFA in muscle. It remains to be elucidated whether the observed high FFA and low WBCC values after RR have a negative effect on health. If so, AR could be a preventive measure.

Adult↗
PubMed2001

Plasma fibrinogen level and long-term prognosis in Norwegian middle-aged patients with previous myocardial infarction. A 10 year follow-up study.

OBJECTIVES: To investigate the prognostic value of plasma fibrinogen level amongst middle-aged survivors of myocardial infarction (MI). DESIGN: Prospective cohort study. SETTINGS: Determination of fibrinogen and other prognostic variables in MI patients recruited in a presumably stable phase of coronary heart disease (CHD). SUBJECTS: A total of 247 middle-aged CHD patients (54 women and 193 men) who had their first MI at age < or = 60 (women) or < or = 55 (men) were recruited at least 3 months after (mean 2.1 years) the most recent MI. MAIN OUTCOME MEASURES: The primary endpoint was total mortality, and the secondary endpoint was cardiac deaths. The tertiary endpoint was major cardiac events (cardiac death, MI and cardiac arrest). RESULTS: During a follow-up period of 10 years a total of 44 patients had died, 36 from cardiac causes. Major cardiac event occurred in 70 patients. After adjusting for age, ejection fraction (EF), total serum cholesterol (TC), smoking and hypertension, patients in the top quartile of fibrinogen (> or = 4.0 g L-1) had a relative risk (RR) of 1.8 (95% CI 1.0-3.6) (P = 0.07) for death of all causes. The top quartile of fibrinogen was a stronger predictor of cardiac death; RR = 2.2 (95% CI 1.1-4.4) (P = 0.03), whilst the effect on the endpoint major cardiac event was not significant; RR=1.1 (95% CI 0.6-1.9) (P = 0.69). CONCLUSIONS: A plasma fibrinogen level in the top quartile predicted cardiac death in middle-aged patients who had suffered MI.

Female↗
PubMed2001

Neisseria meningitidis lipopolysaccharides in human pathology.

Neisseria meningitidis causes meningitis, fulminant septicemia or mild meningococcemia attacking mainly children and young adults. Lipopolysaccharides (LPS) consist of a symmetrical hexa-acyl lipid A and a short oligosaccharide chain and are classified in 11 immunotypes. Lipid A is the primary toxic component of N. meningitidis. LPS levels in plasma and cerebrospinal fluid as determined by Limulus amebocyte lysate (LAL) assay are quantitatively closely associated with inflammatory mediators, clinical symptoms, and outcome. Patients with persistent septic shock, multiple organ failure, and severe coagulopathy reveal extraordinarily high levels of LPS in plasma. The cytokine production is compartmentalized to either the circulation or to the subarachnoid space. Mortality related to shock increases from 0% to > 80% with a 10-fold increase of plasma LPS from 10 to 100 endotoxin units/ml. Hemorrhagic skin lesions and thrombosis are caused by up-regulation of tissue factor which induces coagulation, and by inhibition of fibrinolysis by plasminogen activator inhibitor 1 (PAI-1). Effective antibiotic treatment results in a rapid decline of plasma LPS (half-life 1-3 h) and cytokines, and reduced generation of thrombin, and PAI-1. Early antibiotic treatment is mandatory. Three intervention trials to block lipid A have not significantly reduced the mortality of meningococcal septicemia.

Cytokines↗
PubMed2000

Fulminant meningococcal septicemia: dissociation between plasma thrombopoietin levels and platelet counts.

Thrombopoietin (TPO), interleukin (IL)-6, and platelets were measured serially in 9 patients with fulminant meningococcal septicemia and consumption coagulopathy. The results were compared with those of patients with meningococcal meningitis and mild meningococcemia (n=10) and with those of healthy control subjects (n=19). TPO levels in control subjects were below the detection limit (<63 pg/mL). In patients with fulminant meningococcal septicemia, the median TPO level on admission was 193 pg/mL (range, 133-401 pg/mL), and the level peaked within 3-7 days (median, 488 pg/mL; range, 239-1334 pg/mL). Platelet counts remained low, despite the elevated TPO levels. In patients with meningitis or meningococcemia, the median TPO level on admission was 112 pg/mL (range, <63-695 pg/mL), and the TPO level was not detectable within 48 h. Platelet counts for these patients remained within normal limits. Maximum IL-6 levels in patients with septicemia were observed on admission (median, 5317 pg/mL; range, 188-651,000 pg/mL) and increased earlier than TPO levels. In patients with fulminant septicemia, TPO level increases significantly whereas the level of circulating platelets does not.

Adult↗
PubMed2000

Active recovery reduces the decrease in circulating white blood cells after exercise.

This study was conducted to examine the effects of different recovery regimens on white blood cell count (WBCC) and muscle enzyme activities following strenuous, submaximal, steady state workouts on a treadmill. Fourteen endurance trained, healthy, non-smoking college-aged males participated in the study. The workouts were followed by either 15-min of rest recovery (RR), or active recovery (AR). The AR consisted of running at 50% of VO2max whereas RR implied complete rest. Seven subjects completed two sets of 60-min running at 70% of VO2max (moderate intensity group, MI) followed by either RR or AR. The other seven completed two sets of 30-min running at 80% of VO2max (high intensity group, HI) followed by either RR or AR. Blood samples were drawn at rest, immediately after exercise, and at 15- and 120-min post-exercise (PE). Blood lactate concentrations increased throughout the running trials. Creatine kinase (CK), lactate dehydrogenase (LD), white blood cell count (WBCC) and thrombocyte count increased between rest and 0-min PE (p<0.05). Between 0-15-min PE, there were several significant differences between RR and AR in the HI-trial. RR was associated with a 35% reduction in WBCC, compared to only 6% decrease in AR (p<0.02). Neither during 15-120-min PE this period, nor in the 120-min sample alone, were there any significant differences in WBCC between the RR and AR experiments. In conclusion, the results show that AR as opposed to rest recovery prevents the initial 0-15-min PE fall in WBCC after strenuous endurance exercise.

Adult↗
PubMed2000

Isolation of monocytes from whole blood by density gradient centrifugation and counter-current elutriation followed by cryopreservation: six years' experience.

BACKGROUND: Monocyte purification by means of counter-current elutriation and subsequent cryopreservation for future use was initiated in 1986 and has been established as a routine since 1993. AIM: To sum up and evaluate our method for the isolation and preservation of monocytes. MATERIALS AND METHODS: Peripheral blood mononuclear cells (PBMC) were isolated from healthy donor blood by density gradient centrifugation, and monocytes were isolated from the PBMC by counter-current elutriation centrifugation using the Beckman J-6M/E centrifuge. The monocytes were then cryopreserved at 135 degrees C and thawed when required for experimental use. RESULTS: Results are given for the last 6 years, including 59 elutriations and the fractions containing monocytes. The mean purity of monocytes was 93% (range 64-98%); mean recovery was 51% (range 22-55%). Studies of CD14 expression and Annexin V indicate that there are no differences between elutriated fractions immediately upon purification or after freezing and thawing. The studies also indicate that interdonor variations are much larger than intradonor variations. DISCUSSION: Although it differs from other reports in certain respects, our procedure has nevertheless produced results in line with other findings. After extensive testing and use in different contexts we feel confident that we have established a method for producing a large number of purified and well-preserved monocytes. CONCLUSION: The goal of being able to perform a large number of experiments with monocytes of high purity and good functionality has been reached.

Annexin A5↗
PubMed2000

Aspirin potentiates LPS-induced fibrin formation (FPA) and TNF-alpha-synthesis in whole blood.

The effect of aspirin on LPS-incubation of whole blood was investigated. Aspirin induced a concentration dependent increase (2.5-5-fold at 5 mM aspirin) in LPS-induced appearance of TNF-alpha and fibrinopeptide A (FPA) in plasma, despite the concomitant increase in the inhibitory cytokine IL-100. Aspirin substantially raised the levels of LPS-induced TF-mRNA and TNFalpha-mRNA in monocytes isolated from whole blood. The median ratio for TF-/beta-actin mRNA increased from 1.5 +/- 0.44 in the presence of LPS-alone, to 2.5 +/- 0.51 when 5 mM aspirin was added. The TNFalpha/beta-actin mRNA ratios were 1.8 +/- 0.4 and 5.5 +/- 2.7 respectively. Addition of exogenous PGE2 before incubation nearly abrogated the effect of aspirin on TNF-alpha, substantiating the role of PGE2 as a regulator of TNF-alpha synthesis, whereas the effect on FPA was small. Thus, in the presence of LPS in this whole blood model, aspirin apparently had a pro-inflammatory rather than an anti-inflammatory effect.

Aspirin↗
PubMed2000

Cellular activating properties and morphology of membrane-bound and purified meningococcal lipopolysaccharide.

Neisseria meningitidis, the cause of epidemic meningitis and acute lethal sepsis, synthesizes surplus lipopolysaccharides (LPSs) during growth, which are released as outer membrane vesicles (OMV) or "blebs". Meningococcal disease severity is related to plasma LPS levels. We have compared the biological activities of native outer membrane vesicles (nOMV) to those of purified Nm-LPS (Nm-LPS) and LPS-depleted OMV (dOMV) prepared from N. meningitidis. The LPS content of nOMV was determined spectrophotometrically by quantifying KDO and by silver-stained SDS-PAGE gels. The morphology of the preparations was studied by transmission electron microscopy. The Limulus amoebocyte lysate (LAL) assay was used to quantify LPS in the plasma solutions. The preparations were diluted in endotoxin-free heparin plasma to equal amounts of LPS (w/w) in the range 50-5000 pg/ml. The biological reactivity was tested by: (i) a monocyte target-assay (monocyte purity > or =96%); and (ii) a whole blood model, measuring the secretion of TNF-alpha and IL-6 induction of procoagulant activity in monocytes (PCA). In both models, nOMV induced dose-dependent cell responses (TNF-alpha, IL-6, PCA) similar to purified Nm-LPS, whereas dOMV induced minimal responses. However, LAL activity was significantly higher for nOMV than for purified Nm-LPS and dOMV. The cellular responses of purified Nm-LPS and nOMV were reduced (>95%) by a specific anti-CD14-antibody.

Adult↗
PubMed1999

Effects of doxazosin and atenolol on circulating endothelin-1 and von Willebrand factor in hypertensive middle-aged men.

Elevated levels of endothelin-1 (ET-1) and von Willebrand factor (vWF), both markers indicative of endothelial function, are associated with hypertension. In a randomized open study we investigated the effect of antihypertensive treatment with the alpha-blocker doxazosin (n = 23) or the beta-blocker atenolol (n = 22) for 22 weeks on circulating levels of ET-1 and vWF in middle-aged men with essential hypertension. Blood pressure reduction was satisfactorily achieved with both drugs, although the decrease in the atenolol group was larger than that in the doxazosin group. A reduction in the levels of vWF occurred in both groups, being more pronounced in the alpha-blocker group compared with the decrease on beta blockers, p = 0.004 and p = 0.056, respectively. In the alpha-blocker group, there was a significant correlation (r = 0.50, p = 0.022) between the reduction in diastolic blood pressure and the decline in vWF. A highly significant decrease in plasma ET-1 was obtained during beta blockade (p = 0.007), whereas no significant change occurred within the alpha-blocker group. There was, however, no correlation between the decrease in blood pressure and the reduction in ET-1. The different favorable effects of alpha and beta blockers on endothelial function expressed as vWF and ET-1, could indicate that the effects are probably related not only to the blood pressure per se, but also to the different pharmacologic mechanisms of the drugs.

Adrenergic alpha-Antagonists↗
PubMed1999

Procoagulant activity and cytokine expression in whole blood cultures from patients with atherosclerosis supplemented with omega-3 fatty acids.

Omega-3 fatty acids (n-3 FA) may reduce atherogenesis and thrombosis. We investigated the effects of n-3 FAs on procoagulant activity and cytokine expression in whole blood cultures from patients with atherosclerosis. Eleven of the 23 included patients had received 5.1 g n-3 FA daily for 6 months (group I) whereas 12 patients had been on placebo (group II). All patients were then given 5. g n-3 FA daily for another 4 weeks. At baseline significantly lower levels of LPS-induced prothrombin fragment1+2 were found in group I (p = 0.010), this difference being eliminated after 4 weeks. Il-6 and TNFalpha were significantly higher at baseline in group I and the differences in changes from baseline between the groups were statistically highly significant with increasing values in group II(Il-6 p = 0.001, TNF alpha p = 0.002). The present results indicate a reduction in pro-thrombotic potential in patients receiving highly concentrated n-3 FA, whereas some proinflammatory responses might be adverse.

Adult↗
PubMed1998

Beta-thromboglobulin in urine and plasma: influence of coronary risk factors.

Blood platelet activation in vivo was evaluated by measuring beta-thromboglobulin in plasma and high molecular weight beta-thromboglobulin in urine in hypertensive smoking and nonsmoking middle-aged men (n=36) and in normotensive age-matched controls (n=40). We found no significant linear relationships between nocturnal or resting urinary high molecular weight beta-thromboglobulin and plasma beta-thromboglobulin in the combined hypertensive and normotensive groups. The excretion of high molecular weight beta-thromboglobulin correlated significantly with diastolic blood pressure when all subjects were pooled. After 60 minutes supine rest, nonsmokers had higher excretion of high molecular weight beta-thromboglobulin than smokers. Plasma beta-thromboglobulin levels tended to be higher in hypertensives. In multivariate analyses, resting high molecular weight beta-thromboglobulin excretion was positively related to diastolic blood pressure and negatively related to smoking, whereas plasma beta-thromboglobulin was positively related to diastolic blood pressure and inversely related to apolipoprotein A1 and B. We conclude that urinary high molecular weight beta-thromboglobulin and plasma beta-thromboglobulin are not closely related, but are complementary analyses, as there are methodological confounders for both variables.

Adult↗
PubMed1998

Replacement of partially hydrogenated soybean oil by palm oil in margarine without unfavorable effects on serum lipoproteins.

We have compared the effects of three different margarines, one based on palm oil (PALM-margarine), one based on partially hydrogenated soybean oil (TRANS-margarine) and one with a high content of polyunsaturated fatty acids (PUFA-margarine), on serum lipids in 27 young women. The main purpose of the study was to test if replacement of trans fatty acids in margarine by palmitic acid results in unfavorable effects on serum lipids. The sum of saturated fatty acids (12:0, 14:0, 16:0) was 36.3% of total fatty acids in the PALM-diet, the same as the sum of saturated (12:0, 14:0, 16:0) (12.5%) and trans (23.1%) fatty acids in the TRANS-diet. This sum was 20.7% in the PUFA-diet. The content of oleic acid was 37.9, 35.2, and 38.6%, respectively, in the three diets, whereas linoleic acid amounted to 16, 13.5, and 27.3%, respectively. Total fat provided 30-31% and the test margarines 26% of total energy in all three diets. The subjects consumed each of the diets for 17 d in a Latin-square crossover design. There were no significant differences in total cholesterol, low density lipoprotein (LDL)-cholesterol and apolipoprotein B (apoB) between the TRANS- and the PALM-diets. High density lipoprotein (HDL)-cholesterol and apoA-1 were significantly higher on the PALM-diet compared to the TRANS-diet whereas the ratio of LDL-cholesterol to HDL-cholesterol was lower, although not significantly (P = 0.077) on the PALM-diet. Total cholesterol, LDL-cholesterol, and apoB were significantly lower on the PUFA-diet compared to the two other diets. HDL-cholesterol was not different on the PALM- and the PUFA-diets but it was significantly lower on the TRANS-diet compared to the PUFA diet. Compared to the PUFA-diet the ratio of LDL- to HDL-cholesterol was higher on both the PALM- and the TRANS-diets whereas apoA-1 was not different. Triglycerides and lipoprotein (a) were not significantly different among the three diets. We concluded that nutritionally, palmitic acid from palm oil may be a reasonable alternative to trans fatty acids from partially hydrogenated soybean oil in margarine if the aim is to avoid trans fatty acids. A palm oil-based margarine is, however, less favorable than one based on a more polyunsaturated vegetable oil.

Adult↗
PubMed1998

Effect on plasma lipids and lipoproteins of replacing partially hydrogenated fish oil with vegetable fat in margarine.

We have compared the effects on lipoproteins and haemostatic variables of two hard margarines with similar functional properties, one traditional margarine containing partially hydrogenated fish oil (PHFO), and one experimental margarine based on vegetable oil (VO). Both were all-purpose cooking margarines with nearly identical functional properties. Trans fatty acids from PHFO in the traditional margarine were replaced mostly by saturated, monounsaturated and trans fatty acids of vegetable origin in the new formulation. Both test margarines contained approximately the same amount of cis polyunsaturated fatty acids. Sixteen female normolipidaemic students consumed each diet with the two test margarines for 14 d in random order (crossover design). The amount of fat was 31% energy in the PHFO diet and 32% energy in the VO diet. The test margarines provided approximately 26% energy in both diets. In the PHFO diet 7.8% of the energy was derived from trans fatty acids and 9.2% from saturated fatty acids (12:0, 14:0 and 16:0) while in the VO diet, 1.1% energy was derived from trans fatty acids and 13.3% from saturated fatty acids (12:0, 14:0 and 16:0). The natural content of cholesterol in PHFO was deliberately not balanced by addition of cholesterol to the VO diet, thus the PHFO diet contained 215 mg and the VO diet 86 mg cholesterol per 8.5 MJ. LDL-cholesterol concentration was 19% higher in subjects on the PHFO diet compared with the VO diet (P < 0.01). The ratio LDL-cholesterol:HDL-cholesterol was 12.6% higher in subjects on the PHFO diet compared with the VO diet (P < 0.01). The level of apolipoprotein (apo)A-I was 6% lower in subjects on the PHFO diet compared with the VO diet (P < 0.01). The ratio apoB:apoA-I was 10.4% higher in subjects on the PHFO diet than on the VO diet (P < 0.01). There were no significant differences in total cholesterol, HDL-cholesterol, triacylglycerols, apoB, lipoprotein(a) and haemostatic variables between the diets. Our results demonstrate that PHFO, with its unfavourable effects on plasma lipids, can be replaced by vegetable oils in margarine without appreciable loss of functional properties but with significant improvement in the effects on plasma lipoproteins.

Adult↗
PubMed1997

Macrophage tropism: fact or fiction?

Primary HIV-1 isolates can be distinguished by phenotypic qualities such as the ability to productively infect cells of established CD4-positive lines and to induce syncytia in MT-2 cells. Such viral phenotypes have also been reported to confer host cell specificity. It is perceived that primary isolates with the syncytium-inducing phenotype (SI or rapid/high) are T cell tropic and are therefore unable to infect primary cells of the monocyte/macrophage lineage. However, we have consistently found that these isolates are as capable of establishing infection in monocyte-derived macrophages (MDM) as the monocytotropic, non-syncytium-inducing variants (NSI or slow/low). It is known that differentiation, activation, and proliferation of human monocytes are affected by both isolation methods and culture conditions. Therefore, to test whether our inability to discriminate macrophage tropic HIV-1 isolates could be explained by differences in culturing techniques, we isolated monocytes by elutriation or short-term adherence and allowed the cells to mature and differentiate in either the presence or absence of autologous lymphocytes. After removal of nonadherent cells, MDM were infected with a panel of SI and NSI primary HIV-1 isolates. MDM were susceptible to infection by the SI as well as the NSI isolates, regardless of whether or not the cells were allowed to mature in the presence of autologous lymphocytes. However, MDM matured in the presence of autologous lymphocytes replicated HIV-1 isolates (both NSI and SI) to a higher titre than MDM matured in the absence of lymphocytes. In light of these findings and recently published reports on HIV-1 phenotype and chemokine receptor usage we believe that the term macrophage-tropic strains of HIV-1 is no longer appropriate.

Cell Differentiation↗