Tissue factor pathway inhibitor (TFPI) levels in the plasma and urine of children with meningococcal disease.

Tissue factor pathway inhibitor (TFPI) is a potent inhibitor of the TF-dependent coagulation system. In meningococcal disease, up-regulation of tissue factor expression on blood monocytes and possibly on endothelial cells has the potential to trigger the activation of the TF-dependent pathway of coagulation. Intravascular coagulation is considered to be a major pathogenic factor in meningococcal disease. We postulated that imbalance between TF expression and TFPI concentration might lead to uncontrolled coagulation in meningococcal disease. The aim of this study was to assess the levels of total TFPI in the plasma of patients with meningococcal disease and assess whether increased leaking of the TFPI was occurring. TFPI antigen levels and activity were measured in the plasma of 54 patients with meningococcal disease, and 13 healthy control children. TFPI antigen level were also determined in the urines of 14 of the 54 and 9 healthy control children. Plasma TFPI activity was reduced in the meningococcal diseased patients (mean of 0.503 +/- 0.341 U/ml; control, 1.010 +/- 0.199 U/ml: p <0.0001), as was the TFPI antigen levels (mean of 54.85 +/- 35.05 ng/ml; Control, 94.51 +/- 11.44 ng/ml; p <0.0001). In contrast, TFPI antigen levels were increased in the urine of these patients when compared to the levels found in the urine of the healthy control children (mean of 12.96 +/- 5.392 ng/mmol creatinine; Control, 0.239 +/- 0.191 ng/mmol creatinine; p <0.035). A lack of correlation between TFPI-activity and TFPI-antigen plasma levels was observed (r = 0.002, p = 0.85). This data is consistent with the hypothesis that in meningococcal disease there is increased inactivation of plasma TFPI by the up regulation of tissue factor expression but in addition increased clearance of TFPI in urine is occurring.

Suppression of human monocyte tissue factor synthesis by antisense oligodeoxynucleotide.

Tissue factor (TF) is a cellular receptor and cofactor for factor V11/V11a which initiates the blood coagulation cascade. An understanding of TF cell biology is therefore of critical importance to the pathophysiology of many disorders. We have utilized an antisense oligodeoxynucleotide (aODN), directed against human blood monocyte TF mRNA, to investigate the feasibility of inhibiting the synthesis of TF. CD14 receptor-mediated endocytosis was used as a means of delivering TF aODN to monocytes. This DNA carrier system consists of the fab portion of anti-CD14 antibody covalently linked to poly (L-lysine). Co-treatment of monocytes with TF aODN and endotoxin resulted in 80.4 +/- 2.2% suppression of TF activity when compared with control endotoxin stimulated cells. Control experiments with TF sense ODN, mismatched aODN, and an irrelevant aODN were performed to exclude nonspecific inhibitory effects. The cytotoxicity of the DNA carrier complex was also evaluated. These results demonstrate that this TF mRNA antisense ODN specifically suppressed the synthesis of biologically active monocyte TF and that antisense ODNs might therefore represent a useful tool in the investigation of monocyte/macrophage TF function.

Decision making in the neonatal intensive care environment.

Consideration as to whether withdrawal of intensive care support might be a more appropriate line of action than to continue with full intensive care has become a part of the life and death decision making process undertaken in neonatal intensive care units. After outlining the moral objectives of delivery of health care, the arguments for taking quality of life and its various components into account during these deliberations are presented. The circumstances in which the appropriateness of continuing care should be considered are highlighted and the care options presented. The crucial importance of allowing time for parents to come to terms with the situation is emphasised as is the need for giving clear guidelines to junior staff over resuscitation issues. Finally, an environment for providing optimal family support during the process of withdrawal is suggested.

Co-localization of tissue factor protein and its mRNA in human blood monocytes.

Triggering of both the extrinsic and intrinsic coagulation pathways is mediated by the cell surface receptor Tissue Factor (TF). The ability to observe the cell surface TF protein and its mRNA at the single cell level would facilitate our understanding of the cellular biology of TF in health and diseased states. Employing the methods of immuno-gold silver staining and in situ hybridization using non-isotopically labelled oligoprobes, tissue factor antigen and mRNA were detected simultaneously in endotoxin stimulated human peripheral blood monocytes.

Production of tissue factor by monocyte progenitor cells.

Tissue factor (TF) is known to be produced by monocytes from human peripheral blood. However the production of this factor by haematopoietic progenitor cells is not yet known. We thus studied human monocyte progenitor cells isolated from bone marrow of normal and diseased individuals. These cells were non-adherent, monocytic and able to phagocytose particles ranging from 0.3-1 microns. Unactivated partial thromboplastin time clotting assay demonstrated procoagulant activity consistent with TF function, which was blocked by a neutralizing anti-TF monoclonal antibody, G12. The production of TF messenger RNA was demonstrated on dot blot and northern blot analysis utilizing an oligonucleotide probe.

Temporary inhibition of growth and adrenal suppression associated with the use of steroid nose drops.

Inhibition of growth and adrenal suppression are reported following the use of intranasal beta-methasone (0.1%) in a 9-year-old boy with cystic fibrosis and nasal polyps and in a 3-year-old girl with allergic rhinitis. On stopping treatment catch-up growth occurred and adrenal function returned to normal.

The variation with gestational age of the rheological properties of the blood of the new-born.

A study has been made of the variation of blood viscosity and related factors with the gestational age of neonates from 24 weeks to normal term. Viscosity increases significantly over this period by 36% at high shear rate (128.5 s-1) and 250% at low shear (0.277 s-1). The high shear rate changes can be explained largely by the effects of variations in haematocrit and plasma viscosity. At low shear rate the same factors are involved together with changes in the plasma protein composition, in particular the age-related increase in the concentration of the proteins known to induce rouleaux formation. The variation in the degree of sialination of fibrinogen with gestational age may also play a part.

The development of anti-HLA antibodies in multiply transfused preterm infants.

The development of antihuman leucocyte antigen antibodies (aHLAA) in response to multiple transfusions in preterm infants was studied prospectively. Fifty seven infants requiring a minimum of two blood transfusions were recruited after obtaining informed written parental consent. They were randomised to receive either whole blood or blood that had been passed through a leucocyte filter. Anti-HLAA were sought in maternal and cord blood so as to ensure that any aHLAA detected after transfusion had not been passively transferred antenatally, and in 1 ml samples drawn monthly from the baby, at least 10 days from a previous transfusion, until discharge from hospital. Anti-HLAA were detected by microlymphocytotoxicity assay. Results were obtained in 42 babies, 19 in the filter and 23 in the no filter group. Fifteen babies had to be excluded because of protocol violation or because they died. None of the babies receiving filtered blood developed aHLAA, but seven babies in the no filter group developed aHLAA. In conclusion, multiply transfused preterm infants have the ability to elaborate antibodies to HLA and leucocyte filters may prevent this.

The expression of surface tissue factor apoprotein by blood monocytes in the course of infections in early infancy.

The expression of surface tissue factor procoagulant activity and its shedding by blood monocytes can be induced by several stimuli. Few of these defined situations, other than the presence of bacteria and their toxins, are commonly present in the young human infant. In this study, measurements were made of the percentage of monocytes expressing surface tissue factor apoprotein (TFA) in blood taken from babies in the early weeks of life. Mononuclear cells were separated from blood in an environment free of detectable endotoxin. After exposure to a polyclonal rabbit antibody raised to purified brain TFA and subsequent exposure to a fluorescin-labeled murine anti-rabbit IgG, the cell fluorescent activity was analyzed by flow cytometry. The percentage of monocytes showing strong fluorescence was determined. In every instance when systemic bacterial infection was present, more than 60% of the monocytes examined showed fluorescence indicative of the presence of surface TFA. In a single case of fungal Candida septicemia, none of the monocytes was positive. More than 60% of cells were found to be positive in certain instances where infection was highly probable but not proven. Positive cells were found in three cases of isoimmune hemolytic disease of the newborn, as had been anticipated from previous studies, whereas less than 25% of monocytes derived from babies in the absence of discernible infection or isoimmune hemolytic disease expressed surface TFA (p less than 0.001). These findings provide insight into a possible mechanism of coagulation activation in sepsis and may prove to be a useful predictor of the presence of infection or endotoxemia in young infants.

Low dose intraventricular fibrinolytic treatment to prevent posthaemorrhagic hydrocephalus.

Posthaemorrhagic ventricular dilatation (PHVD) is thought to be due to clots from intraventricular haemorrhage obstructing cerebrospinal fluid pathways involved in reabsorption. Over 60% of infants with progressive PHVD have gone on to require surgical shunt placement. Previous treatments all have major problems. The object of this pilot study was to achieve enough fibrinolysis to restore pathways of cerebrospinal fluid reabsorption and so avoid shunt surgery. Nine preterm infants with progressive PHVD were treated with intraventricular infusion of streptokinase for 12-72 hours. All the infants survived and surgical shunting was required in only one case. A 200% increase in fibrinolytic activity was demonstrated in both ventricular and spinal fluid during streptokinase treatment. There were no cases of infection. Minor rebleeding occurred in one case and was not a serious problem. This represents the first direct therapeutic approach to the pathology of PHVD.

The cardiovascular and oncotic effects of albumin infusion in premature infants.

We determined the oncotic and cardiovascular effects of a standardised infusion of human albumin (1.2 g/kg over 2 h as a 20% solution) in 12 premature infants on 18 occasions when hypovolaemia was suspected on clinical grounds. Blood volume increased by a median value of 15.5%, and fell to preinfusion values by 3 h post infusion in all but four cases. Albumin concentration and colloid osmotic pressure rose during infusion and remained raised even when blood volume had fallen to preinfusion levels. Blood pressure rose in 3 cases only and heart rate fell by greater than 5 beats/min in 6 cases. Indices of long- and short-term heart rate variability were unchanged, but blood pressure variability fell in the second hour of infusion (P = 0.03), an effect which was independent of changes in lung inflation. No changes in blood gases or oxygenation occurred during infusion and no evidence of pulmonary oedema was found. There were wide variations in oncotic and cardiovascular responses to the standardised infusion both between and within subjects. When human albumin is infused in this manner some protection against respiration-induced variability in blood pressure can result, but the circulatory response may prove difficult to predict in the individual.

Postprandial motor response of the small intestine to enteral feeds in preterm infants.

The postprandial motor response to enteral feeds of the small intestine of preterm infants is dependent on the bolus volume of feed that the infants receive and the number of days over which enteral feeds have been given. The postconceptional age of the infant is not an important determinant. The early introduction of enteral nutrition to preterm infants is therefore likely to enhance the motor response of the small intestine to feeds.

A comparison of the rheologic properties of neonatal and adult blood.

A number of studies have indicated that the rheologic properties of neonatal blood are different from those of the adult. The frequent administration of blood components to the neonate during intensive care make it important that these differences be established and their causes understood. The purpose of this study was to make a detailed comparison of the rheologic properties of neonatal and adult blood, with particular emphasis on low shear rate viscosity and rouleaux-related phenomena. The viscometric data was obtained from seven preterm (PT) and 18 normal term (NT) babies and compared with those from 18 adults (A). In the present study, viscometry was performed over a wide range of shear rates, from about 0.3 to 130 s-1, and the low shear rate data were compared with direct measurement of rouleaux formation using the Myrenne Erythrocyte Aggregometer. A major factor leading to the viscometric differences observed was the high hematocrit common in the newborn (46.8 +/- 2.1% PT, 52.8 +/- 6.1% NT, 44.1 +/- 2.5% A males, 40.5 +/- 1.9% A females). However, this tended to be compensated for by the lower plasma viscosity (1.05 +/- 0.07 mPas PT, 1.23 +/- 0.14 mPas NT, 1.34 +/- 0.08 mPas A--no sex difference) and reduced rouleaux formation observed in the newborn and more marked in the preterm baby. The lowered levels of red cell aggregation were found not to be due to cellular differences between the adults and the babies but rather to differing plasma components. The presence of the fetal variant of fibrinogen and low levels of immunoglobulins, especially IgM and IgA, are likely to be of particular importance.

Ontogeny of fasting small intestinal motor activity in the human infant.

A clearly defined progression of fasting small intestinal motor development is seen in the human infant from disorganised low amplitude motor activity before 31 weeks gestation through an intermediate phase of increasing motor organisation and amplitude to the development of a normal cyclical pattern of motor activity with clearly defined phase I, II, and III activity between 37 weeks gestation and term. With increasing maturity smooth muscle contractility [gastric antral pressure (5-30 mmHg), average duodenal pressure (2-12 mmHg)], propagation and slow wave frequency (10.5-12.5 cpm) all increased in a significant fashion (p less than 0.01). The stage of development of fasting motor activity in the small intestine of the preterm infant can now be readily predicted from the gestational age of the infant.

Measurement of small-intestinal motor activity in the preterm infant.

The development of motor activity within the small intestine of preterm human infants is very poorly understood. Using a miniaturized multilumen continuous perfusion manometric technique the motor activity of the small intestine was followed longitudinally in a group of preterm infants. The system performed well at very low perfusion rates and clearly showed increases in both the magnitude and organization of motor activity with increasing gestational age.

Quantification of cardiovascular instability in premature infants using spectral analysis of waveforms.

Spectral analysis was applied to blood pressure and cerebral blood flow velocity recordings in premature infants with respiratory distress in order to quantify respiration-induced cardiovascular variability. Aortic blood pressure was transduced via an umbilical arterial catheter and cerebral blood flow velocity measured in the anterior cerebral artery using a 10 MHz continuous wave Doppler velocimeter in 16 infants less than or equal to 32 wk gestational age. Spectral analysis of the resulting waveforms revealed heart rate and respiratory rate components whose relative amplitudes (heart rate/respiratory rate amplitude ratio) represent an index of that component of variability induced by respiratory events. The mean (heart rate/respiratory rate amplitude) ratio was 47.2 in spontaneously breathing infants and rose to 165.9 in infants who were ventilated during muscle paralysis (p = 0.0003). Cerebral blood flow velocity recordings showed R components in only 22 of 38 simultaneous recordings. This method can be used to quantify respiration-induced cardiovascular variability and its response to therapy, and may provide a means of identifying infants at risk from brain injury due to an inability to regulate cerebral blood flow.