Otwin Linderkamp

Fas- or ceramide-induced apoptosis is mediated by a Rac1-regulated activation of Jun N-terminal kinase/p38 kinases and GADD153.

In the present study, we show that Fas receptor ligation or cellular treatment with synthetic C6-ceramide results in activation or phosphorylation, respectively, of the small G-protein Rac1, Jun N-terminal kinase (JNK)/p38 kinases (p38-K), and the transcription factor GADD153. A signaling cascade from the Fas receptor via ceramide, Ras, Rac1, and JNK/p38-K to GADD153 is demonstrated employing transfection of transdominant inhibitory N17Ras, N17Rac1, c-Jun, or treatment with a specific p38-K inhibitor. The critical function of this signaling cascade is indicated by prevention of Fas- or C6-ceramide-induced apoptosis after inhibition of Ras, Rac1, or JNK/p38-K.

The Effect of Epoetin-beta (recombinant-human-erythropoietin) On the Need for Transfusion in Very-low-birth-weight Infants

BACKGROUND Anemia of prematurity is characterized by low reticulocyte counts and inadequate erythropoietin response, for which many very-low-birth-weight infants receive multiple blood transfusions. We investigated whether early treatment of such infants with recombinant human erythropoietin would reduce their need for transfusions. METHODS We performed a controlled, blinded trial in 241 infants with very low birth weights at 12 centers in six European countries. When three days old, the infants were randomly assigned either to the epoetin group or to the control group. Those in the epoetin group received 250 IU of epoetin beta per kilogram of body weight subcutaneously three times a week from day 3 to day 42 (for a total of 17 doses); those in the control group did not receive this drug. Infants in both groups received oral iron (2 mg per day) from day 14 onward. RESULTS The control infants needed a mean of 1.25 transfusions each, as compared with 0.87 transfusion for epoetin-treated infants (P = 0.013). The median cumulative volume of blood transfused per kilogram per day was 0.41 ml in the control group (first quartile, 0 ml; third quartile, 0.8 ml) and 0.09 ml in the epoetin group (first quartile, 0 ml; third quartile, 0.8 ml) (P = 0.044). The rate of success, defined as an absence of need for transfusions and a hematocrit that never fell below 32 percent, was 4.1 percent in the control group and 27.5 percent in the epoetin group (P = 0.008). Epoetin was most beneficial in boys with birth weights of 1200 g or more and a base-line hematocrit of 48 percent or more. No toxic effects were observed in the epoetin group; as compared with the control group, the epoetin group had an increased incidence of septicemia (14 vs. 7 episodes, P not significant) and reduced weight gain (520 vs. 571 g, P = 0.02). CONCLUSIONS Infants with very low birth weights have less need of transfusions if given epoetin beta during the first six weeks of life (250 IU per kilogram three times a week). We recommend early epoetin treatment for all such infants, but further studies of nutrition and iron supplementation during treatment are needed.

Cleavage of L1 in Exosomes and Apoptotic Membrane Vesicles Released from Ovarian Carcinoma Cells

Purpose: The L1 adhesion molecule (CD171) is overexpressed in human ovarian and endometrial carcinomas and is associated with bad prognosis. Although expressed as a transmembrane molecule, L1 is released from carcinoma cells in a soluble form. Soluble L1 is present in serum and ascites of ovarian carcinoma patients. We investigated the mode of L1 cleavage and the function of soluble L1. Experimental Design: We used ovarian carcinoma cell lines and ascites from ovarian carcinoma patients to analyze soluble L1 and L1 cleavage by Western blot analysis and ELISA. Results: We find that in ovarian carcinoma cells the constitutive cleavage of L1 proceeds in secretory vesicles. We show that apoptotic stimuli like C2-ceramide, staurosporine, UV irradiation, and hypoxic conditions enhance L1-vesicle release resulting in elevated levels of soluble L1. Constitutive cleavage of L1 is mediated by a disintegrin and metalloproteinase 10, but under apoptotic conditions multiple metalloproteinases are involved. L1 cleavage occurs in two types of vesicles with distinct density features: constitutively released vesicles with similarity to exosomes and apoptotic vesicles. Both types of L1-containing vesicles are present in the ascites fluids of ovarian carcinoma patients. Soluble L1 from ascites is a potent inducer of cell migration and can trigger extracellular signal-regulated kinase phosphorylation. Conclusions: We suggest that tumor-derived vesicles may be an important source for soluble L1 that could regulate tumor cell function in an autocrine/paracrine fashion.

Risk factors in retinopathy of prematurity : A multivariate statistical analysis

Objective: To analyze risk factors in retinopathy of prematurity (ROP). Patients and Methods: Four hundred forty-seven surviving very-low-birth-weight infants (birth weight ≤1,500 g) were enrolled in a retrospective study. Clinical data underwent multivariate analysis using stepwise logistic forward regression. Results: For 402 of the babies a complete dataset of 38 possible risk factors was available. Multivariate analysis showed 10 factors to be independently significant variables. Low birth weight, low gestational age, artificial ventilation for more than 7 days, high volume of blood transfusion and surfactant therapy were risk factors associated with higher rates of ROP. Necrotizing enterocolitis, maternal pre-eclampsia, lung maturation by antepartum betamethasone application, vitamin E and phototherapy were accompanied by a lower incidence of ROP. Conclusions: This study confirms several risk factors recognized by previous statistical analysis. In addition, this study reveals maternal pre-eclampsia, which is associated with chronic intrauterine stress, as a statistically independent factor and as having an influence on the incidence of ROP.

The effect of early and late cord‐clamping on blood viscosity and other hemorheological parameters in full‐term neonates

This study was done to compare postnatal alterations in blood viscosity (capillary viscometer) and its determinants: hematocrit, plasma viscosity (capillary viscometer), red cell aggregation (Myrenne aggregometer) and red cell deformability (rheoscope) in the first five days of postnatal life in full‐term neonates with early (< 10 s) and late (3 min) cord‐clamping. The fetal blood volume of the placenta (“residual placental blood volume”) decreased from 52±8 ml/kg of neonatal body weight after early cord‐clamping to 15±4 ml/kg after later cord‐clamping. Neonatal blood volume, calculated as the difference between an assumed total feto‐placental blood volume of 115 ml/kg and the measured fetal blood volume of the placenta, was 50% higher in the late cord‐clamped infants than in the early cord‐clamped infants. Both groups showed similar viscosity, hematocrit and other rheological parameters in cord blood. In the infants with early cord‐clamping, the hematocrit decreased from 0.48±0.04 1/1 at birth to 0.43±0.61/1 after 24 h (p < 0.05). Whole blood viscosity did not change significantly with age. After late cord‐clamping, the hematocrit rose from 0.50±0.04% at birth to 0.63±0.051/1 at 2 h of age (p < 0.005) and dropped to 0.59±0.51/1 (p < 0.05) at 24 h. Blood viscosity increased by 40% (p < 0.001) within the first 2 h, but did not change significantly during the following five days. In both groups, plasma viscosity and red cell aggregation increased significantly (p < 0.05) on day 5 due to significant increases in total plasma protein and fibrinogen concentrations (p < 0.01). Red cell deformation was not affected by the mode of cord‐clamping and did not change with age. We conclude that late cord‐clamping results in marked rise in blood viscosity due to increasing hematocrit. The postnatal rise in plasma viscosity and red cell aggregation may keep the blood viscosity at a high level despite falling hematocrit.

Effect of caffeine on oxygen consumption and metabolic rate in very low birth weight infants with idiopathic apnea.

Objective. Methylxanthines are among the most commonly prescribed drugs in neonatal intensive care. This study evaluates the effect of caffeine on oxygen consumption and metabolic rate in premature infants with idiopathic apnea. Methods. Eighteen preterm infants at gestational ages from 28 to 33 weeks and birth weights of 890 to 1680 g were enrolled in the study. Nine preterm infants received caffeine therapy, and 9 served as a control group. Oxygen consumption and energy expenditure were examined before, during, and after caffeine treatment. Results. Oxygen consumption increased significantly from 7.0 ± 0.9 before caffeine to 8.8 ± 0.7 mL/kg/min after 48 hours of caffeine therapy, and energy expenditure increased from 2.1 ± 0.3 to 3.0 ± 0.2 kcal/kg/hour. During the observation period of 4 weeks of caffeine treatment, oxygen consumption increased significantly in the caffeine group compared with the control patients. In the caffeine group, a lower environmental temperature was sufficient to maintain a normal body temperature. With similar caloric intake in both groups during the study period, daily weight gain in the control group was significantly higher (21 ± 4 vs 42 ± 2 g/d). None of the other parameters recorded changed during caffeine therapy. Conclusion. Long-term administration of caffeine in preterm infants is associated with an increase in oxygen consumption and with a reduction of weight gain. This may have implications for clinical practice as nutritional regimens need to be adjusted during this therapy.

Metabolic rate and energy balance in very low birth weight infants during kangaroo holding by their mothers and fathers

The aim of the study was to compare effects of maternal and paternal kangaroo care on oxygen consumption, carbon dioxide production, energy expenditure, skin and rectal temperatures, heart and respiratory rates, arterial saturation, and behavioral states. Eleven preterm infants with gestational age of 28 to 31 weeks, birth weight of 560 to 1390 gm, and postnatal age of 8 to 48 days were studied before, during, and after maternal and paternal kangaroo care. Skin temperature (lower leg) increased significantly during both maternal (36.2 +/- 0.9 degrees vs 36.9 +/- 1.2 degrees C) and paternal (36.3 +/- 0.9 degrees vs 36.8 +/- 0.9 degrees C) kangaroo care. The other parameter changed neither during maternal nor during paternal kangaroo care. We conclude that both maternal and paternal kangaroo care have no adverse effects on energy expenditure.

Fas-induced programmed cell death is mediated by a Ras-regulated O2- synthesis.

Fas induces apoptosis in lymphocytes via a poorly defined intracellular signalling cascade. Previously, we have demonstrated the involvement and significance of a signalling cascade from the Fas receptor via sphingomyelinases and ceramide to Ras in Fas‐induced apoptosis. Here we demonstrate rapid and transient synthesis of reactive oxygen intermediates (ROI) via activation of Ras after Fas. Genetic inhibition of Ras by transfection of transdominant inhibitory N17Ras blocked Fas‐mediated ROI synthesis and programmed cell death. Likewise, the antioxidants N‐acetyl‐cysteine and N‐t‐butyl‐phenylnitrone abolished Fas‐induced cell death, pointing to an important role for Ras‐triggered ROI synthesis in Fas‐mediated programmed cell death.

Postnatal changes in neonatal acylcarnitine profile.

Electrospray-tandem mass spectrometry represents a powerful method for detection of inborn errors of fatty acid metabolism. In the present study, it was used to examine neonatal carnitine metabolism, which reflects fatty acid metabolism. In 70 healthy neonates, blood samples were taken from the umbilical cord and by heel-stick puncture in full-term neonates on postnatal d 5. Cord blood specimens were also obtained from 15 preterm and 10 small-for-gestational-age infants. Acylcarnitine concentrations were measured in dried blood spots by electrospray tandem mass spectrometry. Compared with cord blood, the levels of nearly all acylcarnitine species were significantly higher on the postnatal d 5, whereas free carnitine remained unchanged. Total acylcarnitine/free carnitine-ratio increased, whereas the free carnitine/total carnitine-ratio (0.54 ± 0.05;p < 0.01) further decreased. A reduced availability of free carnitine in the early neonatal period may affect fatty acid oxidation and thus be of potential pathophysiological relevance under conditions with higher energy demands, e.g. in sepsis. Cord blood concentrations of free carnitine, total carnitine, and total acylcarnitines were strongly related to birth weight (p < 0.01). Lower umbilical artery pH, i.e. mild hypoxia, caused accumulation of mainly long-chain acylcarnitines. This implicates that long-chain acylcarnitines could serve as a parameter of perinatal asphyxia.

L-selectin activates JNK via src-like tyrosine kinases and the small G-protein Rac.

Selectin and &agr;4&bgr;7‐integrins have been shown to mediate transient leucocyte interactions with endothelial cells which is a crucial step in the initial immune response to pathogens. We have previously shown that stimulation of T lymphocytes via L‐selectin results in activation of a signalling cascade from the L‐selectin molecule via the tyrosine kinase p56lck and tyrosine phosphorylation of L‐selectin to the stimulation of p21Ras and Rac proteins. In the present study we demonstrate that stimulation of Jurkat T lymphocytes via L‐selectin results in an activation of Jun N‐terminal kinase (JNK) but not of p38‐K. L‐selectin‐initiated activation of JNK is mediated by src‐like tyrosine kinases and the small G‐protein Rac 1/2, since genetic or pharmacological inhibition of p56lck or Rac proteins prevent the stimulation of JNK by L‐selectin. Thus, the data point to a novel signalling cascade from L‐selectin via src‐like tyrosine kinases and Rac proteins to JNK.