Cutaneous and muscular microcirculation in patients with terminal heart failure awaiting transplantation.

Heart failure patients are clinically characterized by extreme cardiomegaly, breathlessness, fluid retention and an early onset of fatigue. Studies have shown generalized restricted blood flow in those patients. Furthermore animal experiments proved an impaired blood flow and a diminished oxygen supply of the skeletal muscle in animals with chronic heart failure. Patients with chronic heart failure are limited to the extent of their ability to regulate their arterial pressure, especially in physical activity. It is however unclear in what way restriction of blood flow in the main arteries correlates with those in capillaries and to what extent. In this study it was examined the depth of capillary circulatory restriction as well as the disregulation of oxygen partial pressure in skeletal muscle in rest and stress conditions, in patients with terminal heart failure.

A quantitative real-time PCR assay for Ehrlichia ruminantium using pCS20.

Heartwater is a tick borne disease that affects ruminants and wild animals in Africa south of the Sahara. It is caused by Ehrlichia ruminantium and transmitted by the tick Amblyomma hebraeum. The protocols currently used to detect heartwater take several days to complete. Here, we describe the development of a pCS20 quantitative real-time PCR TaqMan probe assay to detect E. ruminantium in livestock blood and ticks from the field. The assay is based on the conserved pCS20 gene region of E. ruminantium that contains two overlapping genes, rnc and ctaG [Collins, N.E., Liebenberg, J., De Villiers, E.P., Brayton, K.A., Louw, E., Pretorius, A., Faber, F.E., Van Heerden, H., Josemans, A., Van Kleef, M., Steyn, H.C., Van Strijp, M.F., Zweygarth, E., Jongejan, F., Maillard, J.C., Berthier, D., Botha, M., Joubert, F., Corton, C.H., Thomson, N.R., Allsopp, M.T., Allsopp, B.A., 2005. The genome of the heartwater agent Ehrlichia ruminantium contains multiple tandem repeats of actively variable copy number. PNAS 102, 838-843]. The pCS20 quantitative real-time PCR TaqMan probe was compared to the currently used pCS20 PCR and PCR/32P-probe test with regards to sensitivity, specificity and the ability to detect DNA in field samples and in blood from experimentally infected sheep. This investigation showed that the pCS20 quantitative real-time PCR TaqMan probe was the most sensitive assay detecting seven copies of DNA/mul of cell culture. All three assays, however, cross react with Ehrlichia canis and Ehrlichia chaffeensis. The pCS20 real-time PCR detected significantly more positive field samples. Both the PCR and pCS20 real-time PCR could only detect E. ruminantium parasites in the blood of experimentally infected sheep during the febrile reaction. The PCR/32P-probe assay, however, detected the parasite DNA 1 day before and during the febrile reaction. Thus, because this new quantitative pCS20 real-time PCR TaqMan probe assay was the most sensitive and can be performed within 2h it is an effective assay for epidemiological surveillance and monitoring of infected animals.

Effect of preeclampsia on umbilical cord blood hematopoietic progenitor-stem cells.

OBJECTIVE: The aim of the present study was to determine the influence of preeclampsia on cord blood hematopoietic progenitor-stem cells obtained at delivery because cord blood is increasingly used clinically for stem cell retrieval as an alternative to bone marrow. STUDY DESIGN: Umbilical cord blood was collected from patients fulfilling the criteria for preeclampsia and from gestational age- and birth weight-matched control subjects at delivery (patient/control subjects ratio, 1:2). Cord blood volume and nucleated cell content were measured, and the number of hematopoietic progenitor-stem cells was determined by means of fluorescence-activated cell sorting with the CD34(+) epitope and by means of colony assays with different hematopoietic growth factors. In addition, the expression of adhesion molecules by CD34(+) progenitor-stem cells was examined. RESULTS: In pregnancies affected by preeclampsia, volume and nucleated cell and total CD34(+) cell contents in the collected cord blood were significantly smaller compared with those of control subjects. Furthermore, there was a trend toward a smaller relative number of CD34(+) cells and colony-forming units per nucleated cell in cord blood samples from preeclamptic patients. No difference in the expression of the cell-adhesion molecules leukocyte function-associated antigen 1, very late activation antigen 4, and L-selectin by CD34(+) cells could be found. CONCLUSION: This study shows that preeclampsia affects umbilical cord blood volume and nucleated cell and progenitor-stem cell numbers obtained at birth.

Developmental changes in adhesion molecule expressions in umbilical cord blood CD34 hematopoietic progenitor and stem cells.

OBJECTIVE: The purpose of this study was to determine whether expressions of the cell adhesion molecules LFA-1 (CD11a), VLA-4 (CD49d), and L -selectin (CD62L ) on CD34(+) stem and progenitor cells in umbilical cord blood change during gestation. STUDY DESIGN: In a prospective observational study 3-color fluorescence-activated cell sorting was used to assess the levels of expression of CD11a, CD49d, and CD62L on CD34(+) cells in fresh cord blood samples collected at delivery between 22 and 42 weeks' gestation. RESULTS: The relative number of CD34(+) cells decreased as gestational age increased (r = -0.71; P<.001). Conversely, we found significant increases in cell adhesion molecule expression by CD34(+) cells during gestation (LFA-1, r = 0.47; P =.001; VLA-4, r = 0.33, P =.031; L -selectin, r = 0.61; P<.001). Comparisons between grouped samples from early preterm (22-32 weeks' gestation), late preterm (33-37 weeks' gestation), and term (38-42 weeks' gestation) infants confirmed this correlation and revealed that the major increases occurred between early and late preterm gestation. CONCLUSION: These results suggest a role for cell adhesion molecule expression in the process of migration and homing of circulating stem cells to the fetal bone marrow toward the end of pregnancy. The findings may have implications for the use of preterm cord blood for hematopoietic stem cell transplantation and also for prenatal gene therapy.

Preterm birth and the availability of cord blood for HPC transplantation.

BACKGROUND: Cord blood from deliveries at term can be used for HPC transplantation. The objective of this study was to determine the amounts of cord blood nucleated cells (NCs) and HPCs that were collectable from preterm deliveries. STUDY DESIGN AND METHODS: Cord blood collected from preterm deliveries between 22 and 36 weeks of gestation was compared with regard to volume, NC count (/mL), CD34+ cell count (/mL), and the NC and CD34+ cell counts per cord blood sample and at different gestational ages. RESULTS: A correlation was found between gestational age and NC count (r = 0.52, p<0.001), and an inverse relation was found between gestational age and CD34+ cell count (r = - 0.68, p<0.001). The CD34+ cell count per cord blood sample was independent of gestational age (r = - 0.13, p = NS), and no significant difference between early (22-32 week) and late (33-36 week) preterm deliveries was found (p = 0.870). Comparison with published data from cord blood transplantations revealed that up to one-third of preterm samples contained at least as many NCs (or CD34+ cells) as the median cell dose transplanted (calculated for the median recipient weight) in the respective study. Furthermore, 77 percent of all preterm samples contained at least 1 x 10(7) NCs (and 42% at least 1 x 10(5) CD34+ cells) per kg for transplantation in a recipient of 20-kg body weight, which corresponds to the lower threshold of cells per kg in the graft recommended by Eurocord. CONCLUSION: Preterm delivery should not be a reason to exclude cord blood collection if allogeneic cord blood transplantation in a sibling is planned.

Umbilical cord blood collection before placental delivery during cesarean delivery increases cord blood volume and nucleated cell number available for transplantation.

OBJECTIVE: We sought to determine whether umbilical cord blood collection during cesarean delivery can be improved by collecting cord blood before delivery of the placenta. STUDY DESIGN: Patients undergoing cesarean delivery were randomly assigned to cord blood collection before or after placental delivery. Closed sterile collection systems were used for blood sampling. Cord blood characteristics and maternal outcome parameters were compared between the 2 groups. RESULTS: A total number of 40 patients were available for analysis. No differences in maternal and neonatal characteristics were found. A larger amount of cord blood volume (mean +/- SEM, 93 +/- 7.5 vs 66 +/- 6.6 mL; P =.013) and total nucleated cell number (11.1 +/- 1.2 vs 7.4 +/- 0.8 x 10(8) cells; P =.026) was obtained in the samples collected before compared with those collected after placental delivery. Similarly, there was a trend toward higher total CD34(+) cell number in samples collected in situ (30.0 +/- 6.0 vs 17.4 +/- 2.4 x 10(5) cells; P =.076). Estimated intraoperative blood loss, difference between prepartum and postpartum hemoglobin values, operating time, and puerperal infection rates were similar in both groups. CONCLUSION: If a cesarean delivery is performed, cord blood sampling is more efficacious if performed before delivery of the placenta. This collection method seems beneficial and safe and might therefore be preferably used for related, as well as unrelated, cord blood stem cell banking and transplantation.

In vivo heat shock protects rat myocardial mitochondria.

Heat shock (HS)/stress proteins (HSP) provide protection from a variety of stresses other than HS, including oxidative stress and mitochondria have been implicated as the target of HS-related protection in stressed cultured cells. Here we investigated whether mitochondria also are targets for the HS-mediated protection in vivo. Sprague Dawley rats were exposed, or not, to HS (41 degrees C, 15 min). After a 21 h recovery period, hearts were excised and perfused with or without H2O2 (0.15 mM). Myocardial mitochondria were then isolated, and their oxygen consumption was analyzed. HS prevented H2O2-induced alterations in state 3 respiration while increasing the expression of Hsp70 and heme oxygenase (HO). Thus, in vivo HS protects rat myocardial mitochondrial respiration against the deleterious effects of oxidative injury, a protection relating to Hsp70 and/or HO and targeting state 3 respiration.

Two roles for caveolae in the cardiac myocyte?

The presence of caveolae in many cell types including heart myocytes is well established. It is hypothesized that caveolae may play a role in the storing of excess Ca2+ and may be instrumental in Ca2+ transients during contraction and relaxation in pathological conditions. Furthermore, the presence of substances in caveolae and in their membranes may imply a role in the importing and exporting of key molecules under physiological and pathological conditions. Secretory activity is also suggested by an electron micrograph of rat heart muscle.

Numerical simulation of pulsating flow in the aortic arch.

In order to investigate the flow profiles in the aorta a numerical three-dimensional model of the aortic arch was created. The velocity fields were simulated by applying an inlet velocity corresponding to the physiological velocity of the pressure wave at the aortic valve. The velocity field distribution was found to be uniform throughout the model during the time of increasing inlet velocities. With decreasing inlet velocities a region of low flow developed in the descending portion of the model leading to recirculating flow at the inner wall. At this region of low flow the variation in velocity with time at the inner wall was approximately twice the variation at the outer wall. As a result of the recirculating flow, the wall shear stresses at the inner wall are low and oscillating, predisposing to the development of atherosclerosis. This model shows that transient fluid flow in the aortic arch can be simulated. Biological studies are needed to prove that this model can be used to predict sites of pathology.

[Tau protein. A potential biological indicator for early detection of Alzheimer disease]. / Tau-Protein. Ein potentieller biologischer Indikator zur Früherkennung der Alzheimer-Krankheit.

In 40 patients with Alzheimer's disease (AD), in 5 patients with non-AD dementia and in 36 cognitively normal controls the concentration of protein tau was determined in the cerebrospinal fluid (CSF). Of the AD patients, 19 were very mildly demented (MMSE score from 25 to 28). Even in these patients, CSF tau was significantly more elevated than in controls. In the non-AD patients protein tau was less increased. Among the AD patients there was no association between CSF tau and severity of cognitive impairment or deficit in cerebral blood flow, determined by SPECT. Our findings suggest that CSF tau may be elevated even at the predementia stage of AD and be useful as a biological marker for the early recognition of the disease.

Does urokinase play a role in renal stone formation?

Renal stone formation is a complex multifactorial disease, and it is believed that the initial step in the pathogenesis of urolithiasis must be the precipitation of an organic matrix of mucoproteins followed by precipitation of minerals onto this matrix. An important factor in this process may be the activity and/or concentration of the urinary enzyme, urokinase, which would affect the level of urinary mucoproteins such as uromucoid. In support of this hypothesis, ELISA studies were conducted to investigate the urokinase concentrations in urine obtained from males (22-60 years) with and without renal stones. These results showed a significant decrease in urinary urokinase concentration of renal stone patients which, once again, underlines the possible involvement of urokinase in renal stone formation. Therefore, it seems logical to conclude that urokinase may play an integral role in this multifactorial disease.

The conservation of mitochondrial function by ischaemia--the ischaemia paradox.

The aim of this study was to compare oxidative phosphorylation of rat heart mitochondria, isolated: (i) immediately after excision of the heart; (ii) after 60 min of low flow normoxic perfusion; (iii) after 60 min of high flow normoxic perfusion;: (iv) after 30 min of no-flow ischaemia and (v) after 30 min ischaemia followed by 20 min reperfusion. The results, using a retrogradely perfused rat hart as a model, show that mitochondrial oxidative phosphorylation after 30 min of no-flow ischaemia showed no difference from oxidative phosphorylation measured on mitochondria isolated directly after excision of the heart. However, after 30 min no-flow ischaemia followed by 20 min of reperfusion, oxidative phosphorylation deteriorated in comparison with oxidative phosphorylation after 30 min of ischaemia only. Furthermore, oxidative phosphorylation after 60 min of non-ischaemic high flow perfusion (thus high coronary flow), compared better with oxidative phosphorylation after ischaemia-reperfusion than with oxidative phosphorylation after ischaemia alone.

Acclimatization, preconditioning and heat shock proteins.

During prior acclimatization, mineworkers acquire tolerance against the adverse effects of working in a warm environment. In this article, the possible involvement of heat shock proteins as mediators of acclimatization is proposed. Acclimatization is compared with preconditioning. Preconditioning of isolated cells or organs by prior exposure to a temperature higher than normal or exposure to an ischaemic insult endow tolerance on them when later confronted with a severe ischaemic stress. This tolerance is possibly mediated by heat shock proteins induced by the heating or ischaemic preconditioning episode. Functioning of the induced heat shock proteins may therefore underlie the protective mechanisms in both acclimatization and preconditioning.

The possible role of sodium-potassium adenosine triphosphatase in preterm labour.

An unacceptably high incidence of preterm labour is seen in the black and coloured communities of South Africa. This hypothesis proposes that sodium-potassium adenosine triphosphatase activity plays an important role in preterm labour. The impaired activity of the sodium pump leads to increased cytosolic calcium levels, which may trigger contraction of myometrial smooth-muscle cells, resulting in preterm labour.

Normal binding of calcium to five fibrinogen variants with mutations in the carboxy terminal part of the gamma-chain.

Calcium ions are known to accelerate polymerization of fibrin monomers. Each of the two carboxy terminal domains of normal fibrinogen contains one high-affinity calcium binding site that seems to be situated close to the polymerization site in the gamma-chain. Most hitherto described functionally defective fibrinogen variants showed impaired clot formation. Since the tightly bound calcium ions may influence the conformation of the polymerization site, the question arises whether the abnormal clotting of a dysfibrinogen might be due to defective calcium binding. We investigated binding of calcium to fibrinogen and the effect of calcium on the clotting properties of five heterozygous fibrinogen variants showing normal thrombin-induced fibrinopeptide release but abnormal polymerization of fibrin monomers. Each of these dysfibrinogens has one single amino acid substitution in the carboxy-terminal part of the gamma-chain: fibrinogen Claro (gamma 275 Arg-->His), Milano V (gamma 275 Arg-->Cys), Milano I (gamma 330 Asp-->Val), Bern I (gamma 337 Asn-->Lys), and Milano VII (gamma 358 Ser-->Cys). The shortest thrombin clotting time and the earliest onset of turbidity increase were observed in fibrinogen gamma 358 Ser-->Cys; both parameters were little affected by calcium concentration. In the variant gamma 337 Asn-->Lys, the thrombin time was abnormally prolonged at 0.01 mM Ca2+, but it was normalized at 1 mM calcium. In contrast, the abnormal fibrin polymerization of fibrinogen gamma 330 Asp-->Val was barely improved at increasing calcium concentrations. Both variants with the substitution of gamma 275 Arg, the residue indispensable for normal D:D interactions, showed the slowest rate of fibrin polymerization and the lowest turbidity of fibrin clots at any Ca2+ concentration used. High affinity calcium binding was found to be normal in all five fibrinogen variants studied, suggesting that their abnormal clotting was not due to defective binding of calcium. The gamma-chain in the fragment D1 derived from the variant gamma 337 Asn-->Lys was further degraded by plasmin in the presence and in the absence of calcium, whereas fragments D1 from the other four gamma-chain variants as well as from normal fibrinogen were protected against plasmic degradation in the presence of 1 mM Ca2+.