Effects of isoflavones equol and genistein on bone quality in a rat osteopenia model.

Phytoestrogens might be an alternative medication in prophylaxis and treatment of osteoporosis. In this study, the osteoprotective effects of genistein (GEN) and equol (EQO) were evaluated. After ovariectomy, 44 rats received soy-free food (Control, C) and developed substantial osteoporosis over the course of two months. After that period, the rats were divided into different groups and fed estradiol (E), GEN or EQO for 35 days. To analyze the osteoprotective effects of the tested substances, bone biomechanical properties and histomorphometric changes of the lumbar vertebrae were evaluated. In analyzing the vertebral body compression strength, we found that the EQO (103.8%) and GEN (96.8%) groups reached similar levels relative to the E group, while the C group reached 77.7% of the biomechanical properties of the E group. EQO was significantly superior to C. The histomorphometric evaluation demonstrated an increased number of nodes in EQO- and E-treated rats compared to GEN- and C-treated rats. E led to an improvement of cortical as well as trabecular bone, an advantage that was only partly seen in the other groups. Treatment with phytoestrogens induced improved bone quality. EQO and GEN might be alternatives for hormone replacement therapy, although further studies are needed to elucidate possible side effects.

The impact of anti-endotoxin core antibodies on endotoxin and cytokine release and ventilation time after cardiac surgery.

OBJECTIVES: We hypothesized that a temporary cardiopulmonary bypass (CPB)-induced reduction of endotoxin antibody levels contributes to elevated endotoxin levels and the associated inflammatory consequences, with a significant influence on the postoperative ventilation time period. BACKGROUND: Cardiac surgery using CPB induces a systemic inflammatory response syndrome with an associated risk of increased postoperative morbidity and mortality. METHODS: A total of 100 consecutive patients undergoing elective coronary artery bypass graft surgery using CPB were prospectively investigated. Endotoxin core antibodies (immunoglobulin [Ig] M/IgG against lipid A and lipopolysaccharide), endotoxin, interleukin (IL)-1-beta, IL-6, IL-8 and tumor necrosis factor-alpha were measured serially from 24 h preoperatively until 72 h postoperatively. RESULTS: Eighty-five patients had no complications (group 1), whereas 15 patients required prolonged ventilation (group 2). In both groups, there was a decrease of all antibodies 5 min after CPB onset, compared with baseline values (p < 0.001), an increase of endotoxin and IL-8 peaking at 30 min postoperatively (p < 0.001) and an increase of IL-6 peaking 3 h postoperatively (p < 0.001). In group 2, preoperative antibody levels were lower (p < 0.01)--specifically, the decrease in IgM was significantly stronger and of longer duration (p < 0.002)--and levels of endotoxin (p < 0.001) and IL-8 (p < 0.001) were higher at 30 min postoperatively. CONCLUSIONS: We conclude that an CPB-associated temporary reduction of anti-endotoxin core antibody levels contributes to elevated endotoxin and IL-8 release. Furthermore, lower levels of IgM anti-endotoxin core antibodies were associated with a greater rise in endotoxin and IL-8, as well as prolonged respirator dependence.

Hemostasis in normotensive and hypertensive men: results of the PROCAM study. The prospective cardiovascular Münster study.

BACKGROUND: The greater than normal cardiovascular risk of hypertensive patients could be partly due to an impairment of hemostatic balance found in such individuals. OBJECTIVE: To examine the relationship between hemostatic variables and blood pressures in 1950 apparently healthy male participants in the prospective cardiovascular Münster study aged 40-65 years. METHODS: Blood pressure and other variables were determined, including fibrinogen level, coagulation factor VII clotting activity, protein C level, antithrombin III level, plasminogen activator inhibitor-1 level, euglobulin fibrinolytic activity, and von Willebrand factor level. RESULTS: Age-adjusted mean values of coagulation factor VII clotting activity, plasminogen activator inhibitor-1 level, antithrombin III level, and protein C level in hypertensives and borderline hypertensives were significantly higher than those in normotensive men (e.g. for hypertensive versus normotensive men, coagulation factor VII clotting factor activity 111.5 versus 106.1%, plasminogen activator inhibitor-1 level 5.05 versus 3.22 arbitrary units/ml, and protein C level 111.1 versus 107.0%, P < 0.05-0.01). For most of the hemostatic variables we found positive bivariate correlations to blood pressure (P < or = 0.05). Exceptions were von Willebrand factor level (no correlation to blood pressure), and euglobulin fibrinolytic activity (a negative correlation to systolic blood pressure and no correlation to diastolic blood pressure). Significance persisted in the multiple logistic regression analysis with the exception of the relationships between systolic and diastolic blood pressures and fibrinogen level as well as euglobin fibrinolytic activity after adjustment for age. After adjustment for age and body mass index significance for relationships between systolic blood pressure and coagulation factor VII clotting activity as well as protein C level was also lost. CONCLUSIONS: We conclude that the greater than normal cardiovascular risk of hypertensive patients is partly due to an imbalance in hemostasis.

The relation of interleukin-6, tumor necrosis factor-alpha, IL-2, and IL-2 receptor levels to cellular rejection, allograft dysfunction, and clinical events early after cardiac transplantation.

Immunoregulatory cytokines have been implicated in the pathophysiology of graft dysfunction after heart transplantation (HTx). In 15 consecutive patients undergoing HTx we prospectively determined levels of interleukin-6 (IL-6), tumor-necrosis-factor-alpha (TNF-alpha), interleukin-2 (IL-2), and soluble-interleukin-2-receptor (sIL-2-R) at eight points in time during biopsy and right heart catheterization and within 12 hr of echocardiography during the first three months after HTx. Blood was taken from the pulmonary arterial line. IL-6-levels correlated positively with hemodynamic and echocardiographic parameters of pump dysfunction--namely, pulmonary capillary wedge pressure, pulmonary arterial pressure, right atrial pressure, heart rate--and negatively with isovolumic relaxation time and stroke volume independent of the degree of cellular rejection as classified by the ISHLT criteria. A similar pattern was found for TNF-alpha- and sIL-2-R, while IL-2 correlated negatively with left and right heart filling pressures and positively with fractional shortening. In the three patients who died of sepsis or multiorgan failure within the study period IL-6-, TNF-alpha, and sIL-2-R-levels were elevated and IL-2-levels were suppressed compared with the 12 patients with a stable clinical course. IL-6 and sIL-2-R correlated positively while IL-6 and IL-2 correlated negatively. In this pilot study, a cytokine pattern with elevated levels of IL-6, TNF-alpha, and sIL-2-R as well as suppressed levels of IL-2 in the early period after HTx corresponds to impaired hemodynamics independent of cellular rejection and may indicate an unfavorable prognosis. These cytokines may therefore be useful for monitoring and warrant further study.

T-cell and monocyte subsets, inflammatory molecules, rejection, and hemodynamics early after cardiac transplantation.

BACKGROUND: In the early period after cardiac transplantation, differential diagnosis of graft failure due to rejection, infection, and other causes is important but difficult. METHODS: In 22 consecutive patients undergoing heart transplantation, we prospectively determined levels of interleukin-6 as well as T-cell and monocyte subsets at eight points in time during biopsy and right heart catheterization and within 12 hr of echocardiography during the first 3 months after transplantation. RESULTS: Worse hemodynamic parameters, as characterized by dichotomization according to median values (pulmonary capillary wedge pressure >10 mmHg, mean pulmonary arterial pressure > 18 mmHg, pulmonary vascular resistance > 115 dyn x sec x cm(-5), right atrial pressure > 5 mmHg, cardiac index <3 L/min/m2, early mitral deceleration time < 135 msec, and isovolumic relaxation time <80 msec), were associated with higher levels of interleukin-6, C-reactive protein, polymorphonuclear cells, CD71+/CD14+ monocytes, and IgM levels and, in contrast, with lower levels of immunocompetence markers such as CD3+ T cells, CD4+ T cells, CD8+ T cells, CD3+/CD25+ T cells, CD4+/ CD45RO+ T cells, NK cells, and lower biopsy scores. CONCLUSION: Early after cardiac transplantation, elevated levels of inflammatory cells and soluble inflammatory molecules and lower levels of immunocompetence markers are associated with impaired allograft function in the absence of cellular rejection.

Effects of diets containing olive oil, sunflower oil, or rapeseed oil on the hemostatic system.

Various studies have already shown that the fatty acid composition of dietary fat has different effects on hemostasis and platelet function. However, knowledge on this topic is incomplete. In the present study, fifty-eight healthy students received either a 4-week rapeseed oil [high content of monounsaturated fatty acids (MUFA) and high n-3/n-6 PUFA ratio], an olive oil (high content of MUFA, low n-3/n-6 PUFA ratio) or a sunflower oil (low content of MUFA, low n-3/n-6 PUFA ratio) diet. In each group, effects on hemostatic parameters were compared with a wash-in diet rich in saturated fatty acids with respect to intermediate-time effects on the hemostatic system and platelet function. With the olive oil diet, a reduction of coagulation factors VIIc, XIIc, XIIa, and Xc was found, whereas sunflower oil led to lower values of coagulation factors XIIc, XIIa, and IXc. In all study groups levels of plasmin-alpha2-antiplasmin were lower in week 4 than at baseline. Lower fibrinogen binding on platelets was found after the sunflower oil diet, whereas expression of CD62 and spontaneous platelet aggregation were slightly higher after the olive oil diet. However, given the major differences in the fatty acid compositions of the diets, the differences between the groups with respect to hemostasis tended to be small. Therefore, the clinical significance of the present findings remains to be evaluated.

Prediction of clinical outcome after cardiac surgery: the role of cytokines, endotoxin, and anti-endotoxin core antibodies.

Coronary artery bypass grafting (CABG) using cardiopulmonary bypass (CPB) can lead to a systemic inflammatory response syndrome with organ failure and increased morbidity and mortality. The mechanisms of these findings are still under discussion. We investigated whether anti-endotoxin core antibodies, endotoxin, and proinflammatory cytokines influence the clinical course after cardiac surgery. Seventy-eight patients undergoing CABG using CPB were investigated. Anti-endotoxin core antibodies, endotoxin, interleukin (IL)-6, IL-8, IL-1beta, and TNF-alpha were measured 24 h preoperatively and up to 72 h postoperatively. Patients with a postoperative mechanical ventilation time below 24 h (n = 65; Group A) were compared to patients with prolonged respirator therapy (>24 h; n = 13; Group B). Preoperative antibody levels were significantly lower in Group B (P < 0.001). In this group, antibody levels remained decreased during the observation period (P < 0.001). Endotoxin significantly increased 30' postoperatively in both groups (P < 0.002). The increase in Group B was 3-fold higher (P< 0.001). IL-8 increased postoperatively in both groups, peaking 3 h after surgery (P < 0.001). In Group B, the IL-8 release was significantly higher than in Group A (P < 0.001). IL-6 significantly increased in both groups, reaching its maximum 24 h postoperatively (P < 0.001). No differences between groups were observed. No significant changes of IL-1beta and TNF-alpha were observed. We conclude that anti-endotoxin core antibodies may be predictive of adverse outcome after cardiac surgery. The imbalance between antibodies and endotoxin results in an exaggerated increase in endotoxin and IL-8 with an impact on clinical outcome.

Activation of the cardiac interleukin-6 system in advanced heart failure.

OBJECTIVES: The study objective was to assess the cardiac expression of interleukin-6 (IL6) and its receptor (IL6R) in advanced heart failure. BACKGROUND: While IL6 plasma levels are elevated and associated with an impaired prognosis in advanced heart failure, little is known about the intracardiac expression of the IL6 system. METHODS: Heart tissue was obtained from 20 patients (n=10, idiopathic dilated cardiomyopathy, age 44+/-15 years; n=10, ischemic cardiomyopathy, age 55+/-8 years) at the time of transplantation. Left and right ventricular tissue was subjected to in situ hybridization, Northern blot analysis, and RT-PCR. Signals were quantified by densitometric scanning and corrected for G3PDH-mRNA levels. Right ventricular biopsy specimens (n=11) of patients with arrhythmias and normal cardiac function served as controls. In addition, data were correlated with cardiac catheterization and echocardiography data obtained at transplant evaluation. RESULTS: Ventricular IL6 and IL6R transcripts were detected in all explant specimens examined. Expression of both mRNA species was higher than in controls (P=0.001). Left ventricular IL6 mRNA levels correlated positively with heart rate (r=0.77; P=0.009), pulmonary capillary wedge pressure (r=0.53; P=0.03), right atrial pressure (r=0.77; P=0.003), and inversely with left ventricular ejection fraction (r=-0.61; P=0.03). Right ventricular IL6 mRNA levels correlated inversely with cardiac index (r=-0.48; P=0.05). IL6R expression did not correlate with hemodynamic data. CONCLUSIONS: In advanced heart failure, cardiac IL6/IL6R mRNA expression is increased and may play a role in the pathophysiology of advanced heart failure.

Metallothionein: localization in human transplant endomyocardium, relation to cytokines and allograft function.

BACKGROUND: The aim of this study was to investigate the role of metallothionein in cardiac transplants in relation to cytokines and allograft function. Recent studies have revealed an association of allograft dysfunction with elevated proinflammatory cytokines independent of cellular rejection. In animal experiments, cytokines induced overexpression of metallothionein, a low-molecular-weight protein implicated in cellular stress response. METHODS: In 105 consecutive biopsies from 15 patients during the first 3 months after heart transplantation, metallothionein expression was investigated immunohistochemically. Its relation to serum interleukin-6, tumor necrosis factor-alpha, interleukin-2 (IL-2), soluble interleukin-2 receptor rejection, and echocardiographic parameters was determined. Forty-three biopsies of 12 patients with idiopathic ventricular tachycardia served as controls. RESULTS: Metallothionein expression was demonstrated in small vessels, cardiomyocytes, fibrocytes, and interstitial round cells. A positive relation between interleukin-6 levels and the number of metallothionein-positive small vessels (p < 0.028) was observed. Patients with lower serum IL-2 levels showed significantly higher numbers of metallothionein-positive small vessels (p < 0.043). Grafts with prolonged ischemic time (>150 minutes) showed a significantly higher myocardial metallothionein score (p < 0.021). Metallothionein expression was associated with lower fractional shortening, larger left ventricular end-systolic diameter, and lower mean arterial pressure but not with acute cellular rejection. CONCLUSIONS: Metallothionein expression is associated with elevated interleukin-6 and decreased interleukin-2 serum levels and left ventricular allograft dysfunction in the absence of rejection.

Can fat explain the human brain's big bang evolution?-Horrobin's leads for comparative and functional genomics.

When David Horrobin suggested that phospholipid and fatty acid metabolism played a major role in human evolution, his 'fat utilization hypothesis' unified intriguing work from paleoanthropology, evolutionary biology, genetic and nervous system research in a novel and coherent lipid-related context. Interestingly, unlike most other evolutionary concepts, the hypothesis allows specific predictions which can be empirically tested in the near future. This paper summarizes some of Horrobin's intriguing propositions and suggests as to how approaches of comparative genomics published in Cell, Nature, Science and elsewhere since 1997 may be used to examine his evolutionary hypothesis. Indeed, systematic investigations of the genomic clock in the species' mitochondrial DNA, the Y and autosomal chromosomes as evidence of evolutionary relationships and distinctions can help to scrutinize associated predictions for their validity, namely that key mutations which differentiate us from Neanderthals and from great apes are in the genes coding for proteins which regulate fat metabolism, and particularly the phospholipid metabolism of the synapses of the brain. It is concluded that beyond clues to humans' relationships with living primates and to the Neanderthals' cognitive performance and their disappearance, the suggested molecular clock analyses may provide crucial insights into the biochemical evolution-and means of possible manipulation-of our brain.

Impact of left ventricular dysfunction on cytokines, hemodynamics, and outcome in bypass grafting.

BACKGROUND: Although patients with reduced left ventricular ejection fraction undergoing cardiac operation experience a higher rate of perioperative complications, the contribution of proinflammatory cytokines released during extracorporeal circulation is not well defined. METHODS: We compared arterial and mixed venous levels of interleukin-6, tumor necrosis factor-alpha, soluble interleukin-2 receptor, and interleukin-2 at 10 points in time (24 hours before until 48 hours after extracorporeal circulation) in 21 patients with an ejection fraction of less than 0.45 (study group) to 15 patients with an ejection fraction of more than 0.55 (control group) undergoing elective coronary artery bypass grafting. The study and control group differed with regard to left ventricular ejection fraction (0.37 +/- 0.05 versus 0.66 +/- 0.11, p < 0.05) and reperfusion time (35 +/- 42 minutes versus 18 +/- 4 minutes, p = 0.07), but not age, sex, vessel involvement, number of grafts performed, cross-clamp time, extracorporeal circulation time, core temperature, and duration of ventilation. RESULTS: Six patients in the study group required mechanical support and 1 died. There were no complications in the control group. In the study group, there were higher preoperative interleukin-2 and tumor necrosis factor-alpha levels and a higher maximum cytokine response to extracorporeal circulation for interleukin-2, soluble interleukin-2 receptor, interleukin-6, and tumor necrosis factor-alpha (all p < 0.05). Interleukin-6 correlated with duration of extracorporeal circulation, dose of norepinephrine and epinephrine support, pulmonary capillary wedge pressure, mean pulmonary arterial pressure, right atrial pressure, heart rate, cardiac index, and inversely with systemic vascular resistance. Interleukin-6 was highest in patients with complications. Arterial and venous cytokine levels correlated closely. CONCLUSIONS: Preoperative left ventricular dysfunction is associated with a higher degree of proinflammatory cytokine release during elective coronary artery bypass grafting. This response is associated with impaired hemodynamics and a higher incidence of perioperative complications.

The biologic lower detection limit of six ultrasensitive PSA assays.

BACKGROUND: The benefit of monitoring patients with prostate cancer (PCA) by ultrasensitive measurement of prostate specific antigen (PSA) is frequently discussed. Usually, the analytic lower detection limit of an ultrasensitive assay is determined by the manufacturer. As the analytic lower detection limit does not take into account interfering factors of human serum, the biologic lower detection limit, which is defined as PSA concentration detected in PSA-free human serum, plus 3 standard deviations, is of greater interest. MATERIALS AND METHODS: We investigated the biologic lower detection limit of six ultrasensitive PSA assays. Sera from 15 men with bladder cancer after radical cystoprostatectomy and from 30 healthy women were applied. Hence, we expected no PSA of prostatic origin. RESULTS: The biologic lower detection limit obtained using these sera was up to 30 fold higher (men, 0.29-0.63 ng/ml; women, 0.03-0.69 ng/ml) than the analytic lower detection limit (0.01-0.09 ng/ml). CONCLUSIONS: PSA measurement in sera obtained from men without prostate and women results in PSA values above the ultrasensitive range. Therefore, advantages provided by ultrasensitive PSA measurement in monitoring PCA patients after radical prostatectomy are limited.

Ischemic versus idiopathic cardiomyopathy: differing neurohumoral profiles despite comparable peak oxygen uptake.

OBJECTIVE: We tested the hypothesis that neurohormonal and immunological activation differs in ischemic and idiopathic dilated cardiomyopathy since recent intervention trials indicate that ischemic cardiomyopathy seems to carry a worse prognosis than idiopathic cardiomyopathy of comparable clinical severity. METHODS: In ten patients with ischemic cardiomyopathy undergoing spiroergometric evaluation venous levels of norepinephrine, epinephrine, renin, angiotensin, atrial natriuretic peptide as well as soluble interleukin-2-receptor were determined before, during and 10 min after exercise. Results were compared to sixteen patients with idiopathic cardiomyopathy with similar peak oxygen uptake (13.3+/-3 vs. 13.6+/-3 ml/kg/min; P=ns). RESULTS: In ischemic patients, norepinephrine, angiotensin, and interleukin-2 receptor levels were significantly higher before, during and after exercise. Interleukin-2-receptor levels correlated with angiotensin. CONCLUSIONS: We conclude that in ischemic as compared to idiopathic cardiomyopathy, a more pronounced activation of the sympathetic, renin-angiotensin and T-cell immune system is present at rest, during and after exercise. These data may contribute to explain differences in response to intervention and in prognosis. They warrant further investigation.

Interleukin-6 correlates with hemodynamic impairment during dobutamine administration in chronic heart failure.

Proinflammatory cytokines have been implicated in the pathophysiology of chronic heart failure. We determined mixed venous levels of interleukin-6 (IL6) in 18 heart transplant candidates before, 1, 4, and 24 h after initiation of dobutamine infusion (3 micrograms/kg/min) during hemodynamic evaluation. During the first 4 h of dobutamine, systemic vascular resistance decreased (1358 to 1024 dyn x s x cm-5, P = 0.01) while cardiac index (2.3 to 2.9 l/min/m2, P = 0.008) increased. Both returned to baseline after 24 h. IL6 was elevated at baseline compared to age-matched controls (1.5 (0/4.3) vs. 0 (0/0.5) P = 0.003). There was an increase in IL6 from 1.5 (0/4.3) to 3.6 (0.3/5.3) pg/ml after 24 h (P = 0.04). We found higher IL6 levels in the sicker half of patients as defined by pulmonary capillary wedge pressure > 24 mmHg (P = 0.005), mean pulmonary arterial pressure > or = 35 mmHg (P = 0.01), right atrial pressure > 13 mmHg (P = 0.02), and heart rate > or = 87/min (P = 0.02) as well as mean arterial pressure < 82 mmHg (P = 0.005). In conclusion, in this pilot study IL6 correlates with the severity of chronic heart failure during low dose dobutamine infusion.

Arterial and venous cytokine response to cardiopulmonary bypass for low risk CABG and relation to hemodynamics.

During and after cardiopulmonary bypass (CPB), cytokines may affect cardiac performance and the immune response and are therefore of diagnostic and therapeutic interest. We have used EIA/EASIA kits to measure arterial and venous levels of interleukin-1-beta (IL-1-beta), IL-2, IL-2 receptor (IL-2-R), IL-6, tumor necrosis factor (TNF)-alpha and interferon (IFN)-gamma in 12 men and 3 women (mean age 59.4 +/- 8.5 years, mean left ventricular ejection fraction 66 +/- 11%, average of 2.5 +/- 0.64 vessels affected by disease) undergoing elective coronary artery bypass grafting (CABG). On average each patient received 3 +/- 0.85 bypass grafts and required a postoperative maximum dopamine-dose of 3.8 micrograms/kg per min. Mean CPB and operation times were 60 +/- 21 min, and 132 +/- 16 min, respectively. During CPB, the venous levels of IL-2 temporarily decreased from 234 to 0 (p < 0.05) pg/ml and arterial and venous levels of IL-2-R temporarily decreased from 28 to 16, and 36 to 18 pM (p < 0.05), respectively. After termination of CPB, there was an increase in the arterial and venous levels of IL-6 from below 3 to 253 and 277 pg/ml (p < 0.05) and TNF-alpha from 1.1 to 5.7 and 0.7 to 4.0 pg/ml, respectively (p < 0.05). Tumor necrosis factor-alpha-increases peaked 30 min, and IL-6 increases peaked 4 h after termination of CPB. Twenty-four hours after the end of CPB, IL-6 showed a tendency to return to baseline, but still remained significantly elevated.(ABSTRACT TRUNCATED AT 250 WORDS)

Receptor assay based on surface plasmon resonance for the assessment of the complex formation activity of cyclosporin A and its metabolites.

OBJECTIVE: A new automated receptor assay has been used to determine the complex formation activity of cyclosporin A (CsA) and its metabolites in whole blood. METHODS: CsA in vivo forms a complex with cyclophilin A and calcineurin leading to an inhibition of the calmodulin-dependent phosphatase activity of calcineurin. The equilibrium complex formation gives information about the potential immunosuppressive activity of CsA and its metabolites. To measure the amount of this complex the authors developed an automated receptor assay based on an optical biosensor (Biacore) with surface plasmon resonance (SPR) technology. RESULTS: In the range of 50-300 nM CsA, the intra-day coefficient of variation (CV) was 7.2%, and the inter-day CV was 10.1%. Measuring range of the assay was 10-500 nM with a detection limit of 5 nM and a processing time of 10 min. Recovery rate for sample pretreatment was 74 +/- 5%. 193 blood specimens from heart transplant recipients were analyzed with 3 different methods. The results determined with the receptor assay were correlated with those obtained by fluorescence polarization immunoassay (FPIA; r = 0.599) and high-performance liquid chromatography (HPLC; r = 0.615). CONCLUSION: The receptor assay determines the complex formation activity of CsA and its metabolites with high sensitivity and precision.