Biomedical subjects
Publications and source records attributed to Christoph Bührer.
We report on a newborn infant with complex congenital heart disease (CHD) featuring double outlet right ventricle and hypoplastic left ventricle who had postductal oxygen saturation well above 95% and thus eluded pulse oximetry screening for CHD.
PURPOSE: Thalidomide and its analogues have shown promise in the treatment of multiple myeloma but their therapeutic potential has not been evaluated in models of acute lymphoblastic leukemia (ALL). EXPERIMENTAL DESIGN: We assessed the effects of the thalidomide analogue, CC-4047, on the growth and apoptosis signaling of human B cell precursor (BCP) ALL cell lines and freshly obtained childhood BCP-ALL cells grown with or without stromal cells. In addition, we studied the effects of CC-4047 on the progression and dissemination of xenotransplanted human BCP-ALL cells in nonobese diabetic/severe combined immunodeficiency mice. RESULTS: CC-4047 reduced the proliferation of human BCP-ALL cell lines in vitro. In contrast with the antileukemic effect of cytarabin, this was more pronounced when cell lines or freshly obtained childhood BCP-ALL cells were cocultured with stromal cells. CC-4047 induced the cleavage of caspase-3, caspase-9, and poly(ADP-ribose) polymerase in stroma-cocultured BCP-ALL cells. The inhibition of tumor growth, caspase-3 cleavage, and reduced microvessel density was observed in nonobese diabetic/severe combined immunodeficiency mice inoculated s.c. with childhood BCP-ALL cells upon CC-4047 treatment. After i.v. BCP-ALL xenotransplantation, CC-4047 reduced splenic dissemination. CONCLUSIONS: The thalidomide analogue, CC-4047, displays profound cytostatic effects on stroma-supported human ALL cells both in vitro and in vivo.
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In the immature human brain, periventricular leukomalacia (PVL) is the predominant white matter injury underlying the development of cerebral palsy. PVL has its peak incidence during a well-defined period in human brain development (23-32 weeks postconceptional age) characterized by extensive oligodendrocyte migration and maturation. We hypothesized that the dramatic rise of oxygen tissue tension associated with mammalian birth and additional oxygen exposure of the preterm infant during intensive care may be harmful to immature oligodendrocytes (OLs). We therefore investigated the effects of hyperoxia on rat oligodendroglia cells in vitro and in vivo. Immature OLs (OLN-93), their progenitors [preoligodendrocytes (pre-OL)], and mature OLs were subjected to 80% hyperoxia (24-96 hr). Flow cytometry was used to assess cell death. Cell viability was measured by metabolism of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium (MTT). In addition, 6-day-old rat pups were subjected to 80% oxygen (24 hr) and then sacrificed, and their brains were processed for immunfluorescence staining. Apoptosis was detected at various stages (annexin-V, activated caspase-3) after 24-48 hr of incubation in 80% oxygen in pre- and immature OLs. Mature OLs were resistant to oxygen exposure. These results were confirmed by MTT assay. This cell death was blocked by administration of the pan-caspase inhibitor zVAD-fmk. Degeneration of OLs was confirmed in 7-day-old rat brains by positive staining for activated caspase-3. Hyperoxia triggers maturation-dependent apoptosis in immature and pre-OLs and involves caspase activation. This mechanism may be relevant to the white matter injury observed in infants born preterm.
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Coumarin embryopathy (CE) is a well-documented sequelae of prenatal exposure to vitamin K antagonists. We report on a female premature infant (25 weeks' gestation) born to a mother who had received phenprocoumon during pregnancy following mechanical heart valve replacement. The infant presented with impaired coagulation, intraventricular and minor parenchymal cerebral haemorrhages and midface hypoplasia typical of CE. In addition, there was hepatopathy with conjugated hyperbilirubinemia, elevated liver enzymes and repeated episodes of hypoglycemia upon attempts to discontinue glucose supplementation, all lasting for 4 months. There was corneal opacity with anterior segment dygenesis in the left eye, and persistent pupillary membrane, cataract and persistent hyperplastic primary vitreous in the right eye. While liver disease is an uncommon but serious side effect of vitamin K antagonists, this is the first report describing neonatal hepatopathy as part of CE. In anticoagulation of pregnant women with mechanical heart valves, vitamin K antagonists should be used with utmost restraint.
A case of a premature infant with tracheal atresia together with an exceptional combination of congenital abnormalities that partially corresponds to the TACRD and VACTERL associations is presented. Tracheal atresia was not detectable in the prenatal ultrasound due to lacking of the typical diaphragmatic and pulmonary findings because of the esophago-tracheal fistula, and therefore the resuscitation team was not prepared for this severe airway complication. After prolonged resuscitation efforts were terminated and the newborn expired after birth. Even without typical warning signs physicians have to be aware of tracheal atresia and airway obstruction if VACTERL or TACRD associations are diagnosed.
Serum concentrations of soluble L-selectin by far exceed those of other soluble adhesion molecules, and serum soluble L-selectin concentrations are remarkably stable upon prolonged storage. We present evidence for Ca(2+)-dependent binding interactions between human serum amyloid P (SAP), a proteolysis-resistant pentraxin glycoprotein, and L-selectin, as shown by surface plasmon resonance measurements, protein band shift assays in a native PAGE system, and after SDS-PAGE and membrane transfer. Monoclonal antibodies to L-selectin strongly reduced binding of biotinylated SAP to L-selectin-IgG chimeras immobilized on microtiter plates. As binding was reduced by prior glycopeptidase F treatment of L-selectin but not of SAP, it appears to be based on SAP lectin domain interactions with N-linked L-selectin carbohydrates. In freshly prepared human lymphocytes, SAP incubation induced expression of a beta2 integrin neoepitope associated with high-affinity binding. This was partially blocked by pre-incubation with Fab fragments of two anti-L-selectin antibodies. In flow chamber experiments, SAP inhibited the adherence of human neutrophils to activated endothelium under shear stress. Thus, SAP binds to human L-selectin and affects L-selectin-dependent leukocyte-endothelial interactions.
BACKGROUND: Elevated serum concentrations of S-100B, a 21-kDa protein expressed in astroglial cells, has been used to assess cerebral damage after head trauma, infection, ischemia, and perinatal asphyxia. OBJECTIVE: As S-100B is eliminated by the kidneys, we investigated the feasibility of measuring S-100B in urine of newborns with severe perinatal asphyxia, and in very low birth weight (VLBW) preterm infants at risk for neurodevelopmental impairment. METHODS: We first analyzed urine samples of 8 term or near-term newborns without major medical problems, followed by urine samples of 2 term newborns with severe birth asphyxia, and finally urine samples of 8 VLBW (gestational age 24-28 weeks) infants collected every 4 h for up to 10 days. RESULTS: Urinary S-100B concentrations in 8 term or near-term newborns without major medical problems were consistently <1 microg/l. In 2 term newborns with severe asphyxia (Apgar 0/0/0 and 0/2/4) who subsequently had widespread cerebral damage on magnetic resonance imaging, peak urinary S-100B concentrations on the first day of life were 28.1 and 28.4 microg/l, respectively. In 5/8 VLBW infants, urinary S-100B was> microg/l in samples obtained on the first day of life (range 1.2-44.9 microg/l, median 6.8 microg/l). Peak S-100B in urine samples collected during the first 12 h of life were negatively related to gestational age (R(s)=-0.882, p=0.009). Three of the 8 preterm infants had peak urinary concentrations>0 microg/l but neither ultrasound signs of brain damage nor neurodevelopmental delay at 1 year corrected age. CONCLUSIONS: The determination of urinary S-100B concentrations might be helpful in term infants with severe asphyxia, while high urinary S-100B concentrations in preterm infants are to be attributed to immaturity.
Glutamate antagonists limit the growth of human cancers in vitro. The mechanism of anticancer action of NMDA antagonists is not known, however. In this article, we report that the NMDA antagonist dizocilpine inhibits the extracellular signal-regulated kinase 1/2 pathway, an intracellular signaling cascade that is activated by growth factors and controls the proliferation of cancer cells. Dizocilpine reduces the phosphorylation of cAMP-responsive element binding protein, suppresses the expression of cyclin D1, up-regulates the cell cycle regulators and tumor suppressor proteins p21 and p53, and increases the number of lung adenocarcinoma cells in the G(2) and S phases of the cell cycle. Silencing of the tumor suppressor protein p21 abolishes antiproliferative action of dizocilpine. Consistent with inhibition of the extracellular signal-regulated kinase 1/2-signaling cascade, dizocilpine reverses the stimulation of proliferation induced by epidermal, insulin, and basic fibroblast growth factors in lung adenocarcinoma cells. Furthermore, dizocilpine prolongs the survival of mice with metastatic lung adenocarcinoma and slows the growth of neuroblastoma and rhabdomyosarcoma in mice. These findings reveal the mechanism of antiproliferative action of dizocilpine and indicate that it may be useful in the therapy of human cancers.
We report on three male newborn infants of a highly inbred Lebanese family presenting with a characteristic phenotype: arthrogryposis multiplex, deafness, large inguinal hernia, hiccup-like diaphragmatic contractions, and inability to suck, requiring nasogastric gavage feeding. All three boys died from respiratory failure during the first 3 months of life. Intra vitam or post mortem examinations revealed myopathic changes and elevated glycogen content of muscle tissue. This new syndrome is probably transmitted in an autosomal recessive mode, although X-linked inheritance cannot be excluded.
Hyperphenylalaninemia caused by phenylalanine hydroxylase (PAH) deficiency requires lifelong rigorous diet starting in early infancy to prevent severe neurodevelopmental handicap. In a considerable number of children with mild hyperphenylalaninemia, long-term tetrahydrobiopterin (BH4) treatment significantly improves phenylalanine (phe) tolerance, but it has never been investigated in classic phenylketonuria (PKU). We performed a BH4-loading test in 40 consecutive infants with phe serum concentrations exceeding 240 microM, who had been detected by newborn screening programs. Eighteen out of 40 infants were found to be BH4 responsive. Five of them, responding to the neonatal BH4-loading test, showed a phe tolerance of less than 20 mg/kg/day and a phe pretreatment level of >1000 microM. They were treated with BH4 (20 mg/kg/day) over a period of 24 months. All five children had a sustained response to BH4, allowing substantial easing of dietary restrictions. Before BH4 treatment daily phe tolerance was 18-19 mg/kg, increasing to 30-80 mg/kg on BH4 treatment and decreasing again to 12-17 mg/kg after termination of BH4 treatment. Mutation analysis revealed compound heterozygosity for a putative null and a variant PAH mutation in four patients and homozygosity for a variant PAH mutation in one patient. We conclude that BH4 sensitivity is not restricted to mild hyperphenylalaninemia and that long-term BH4 treatment may also improve phenylalanine tolerance in a considerable number of children with a more severe PKU phenotype.
Both mild hypoxia and exogenous erythropoietin may protect the brain against subsequent severe hypoxia, and the conditioning effect of transient hypoxia is partly mediated by hypoxia-induced endogenous erythropoietin. We now observed in several experimental models that combining transient hypoxia and exogenous erythropoietin may cause neuronal damage. High-dose erythropoietin (40 IU/ml) profoundly impeded synaptic transmission of rat hippocampal slice cultures when used in conjunction with moderate hypoxia (10% O2 for two 8-h periods). Addition of erythropoietin increased viability of cultured rat embryonic cortical neurons at 21% O2 but decreased viability under hypoxic conditions (2% O2) in a dose-dependent fashion. Death of human neuronal precursor cells challenged by oxygen and glucose deprivation was increased by erythropoietin when cells were cultured under hypoxic but not under normoxic conditions. In neonatal rats exposed to moderate hypoxia plus erythropoietin, numbers of degenerating cerebral neurons were increased, as compared to controls or rats subjected to either hypoxia or erythropoietin alone. Thus, erythropoietin may aggravate rather than ameliorate neuronal damage when administered during transient hypoxia.
Fatal congenital nonlysosomal cardiac glycogenosis has been attributed to a subtype of phosphorylase kinase deficiency, but the underlying genes and mutations have not been identified. Analyzing four sporadic, unrelated patients, we found no mutations either in the eight genes encoding phosphorylase kinase subunits or in the two genes encoding the muscle and brain isoforms of glycogen phosphorylase. However, in three of five patients, we identified identical heterozygous R531Q missense mutations of the PRKAG2 gene, which encodes the gamma 2-subunit of AMP-activated protein kinase, a key regulator of energy balance. Biochemical characterization of the recombinant R531Q mutant protein showed >100-fold reduction of binding affinities for the regulatory nucleotides AMP and ATP but an enhanced basal activity and increased phosphorylation of the alpha -subunit. Other PRKAG2 missense mutations were previously identified in patients with autosomal dominant hypertrophic cardiomyopathy with Wolff-Parkinson-White syndrome, characterized by juvenile-to-adult clinical onset, moderate cardiac glycogenosis, disturbed excitation conduction, risk of sudden cardiac death in midlife, and molecular perturbations that are similar to--but less severe than--those observed for the R531Q mutation. Thus, recurrent heterozygous R531Q missense mutations in PRKAG2 give rise to a massive nonlysosomal cardiac glycogenosis of fetal symptomatic onset and rapidly fatal course, constituting a genotypically and clinically distinct variant of hypertrophic cardiomyopathy with Wolff-Parkinson-White syndrome. R531Q and other PRKAG2 mutations enhance the basal activity and alpha -subunit phosphorylation of AMP-activated protein kinase, explaining the dominant nature of PRKAG2 disease mutations. Since not all cases displayed PRKAG2 mutations, fatal congenital nonlysosomal cardiac glycogenosis seems to be genetically heterogeneous. However, the existence of a heart-specific primary phosphorylase kinase deficiency is questionable, because no phosphorylase kinase mutations were found.
Glutaryl-CoA dehydrogenase deficiency is an inherited metabolic disease characterized by elevated concentrations of glutaric acid (GA) and its metabolites glutaconic acid (GC) and 3-hydroxy-glutaric acid (3-OH-GA). Its hallmarks are striatal and cortical degeneration, which have been linked to excitotoxic neuronal cell death. However, magnetic resonance imaging studies have also revealed widespread white matter disease. Correspondingly, we decided to investigate the effects of GA, GC, and 3-OH-GA on the rat immature oligodendroglia cell line, OLN-93. For comparison, we also exposed the neuroblastoma line SH-SY5Y and the microglia line BV-2 to GA, GC, and 3-OH-GA. Cell viability was measured by metabolism of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium. Flow cytometry was used to assess apoptosis via annexin-V, anti-active caspase-3 antibody, and propidium iodide staining. GA, GC, and 3-OH-GA reduced OLN-93 oligodendroglia cell viability in a dose-dependent manner. Toxicity of GA, GC, and 3-OH-GA was abrogated by preincubation with the pan-caspase inhibitor z-VAD-fmk. Apoptosis but not necrosis was detected at various stages (early: annexin-V; effector: caspase-3) after 24-48 h of incubation with GA, GC, or 3-OH-GA in OLN-93 but not in neuroblastoma or microglia cells. OLN-93 lacked expression of N-methyl-d-aspartate receptors, making classical glutamatergic excitotoxicity an unlikely explanation for the selective toxicity of GA, GC, and 3-OH-GA for OLN-93 cells. GA, GC, and 3-OH-GA directly initiate the apoptotic cascade in oligodendroglia cells. This mechanism may contribute to the white matter damage observed in glutaryl-CoA dehydrogenase deficiency.
Infants born prematurely may develop neurocognitive deficits without an obvious cause. Oxygen, which is widely used in neonatal medicine, constitutes one possible contributing neurotoxic factor, because it can trigger neuronal apoptosis in the developing brain of rodents. We hypothesized that two caspase-1-processed cytokines, interleukin (IL)-1beta and IL-18, are involved in oxygen-induced neuronal cell death. Six-day-old Wistar rats or C57/BL6 mice were exposed to 80% oxygen for various time periods (2, 6, 12, 24, and 48 hours). Neuronal cell death in the brain, as assessed by Fluoro-Jade B and silver staining, peaked at 12 to 24 hours and was preceded by a marked increase in mRNA and protein levels of caspase 1, IL-1beta, IL-18, and IL-18 receptor alpha (IL-18Ralpha). Intraperitoneal injection of recombinant human IL-18-binding protein, a specific inhibitor of IL-18, attenuated hyperoxic brain injury. Mice deficient in IL-1 receptor-associated kinase 4 (IRAK-4), which is pivotal for both IL-1beta and IL-18 signal transduction, were protected against oxygen-mediated neurotoxicity. These findings causally link IL-1beta and IL-18 to hyperoxia-induced cell death in the immature brain. These cytokines might serve as useful targets for therapeutic approaches aimed at preserving neuronal function in the immature brain, which is exquisitely sensitive to a variety of iatrogenic measures including oxygen.
Impaired psychomotor development, often anteceded by major intraventricular hemorrhage or periventricular leukomalacia, constitutes the most important long-term morbidity of very preterm infants. We reviewed randomized controlled trials aimed at reducing the incidence of brain damage, as detected by ultrasound, or neurodevelopmental impairment during follow-up of preterm infants. Preliminary reports of reduced rates of intraventricular hemorrhage obtained with administration of fresh frozen plasma, ethamsylate, phenobarbitone, or morphine have not been confirmed in subsequent larger trials. Early administration of indomethacin may reduce intraventricular hemorrhage without affecting long-term outcome. Pancuronium, inositol, and vitamin E decreased intraventricular hemorrhage rates but later psychomotor development was not examined. Thyroxin supplementation failed to improve neurodevelopmental outcome while protein enrichment of formula and individualized developmental care appear to be beneficial. The largest reductions in cerebral palsy and neurodevelopmental impairment were achieved by avoidance of postnatal steroids. This finding emphasizes the need to include these late endpoints in any randomized trial involving preterm infants.
Interleukin (IL)-18 is a potent inflammatory cytokine of the IL-1 family. It is synthesized as an inactive precursor (pro-IL-18), which is cleaved into its functionally active form by caspase-1. Resident cells of the CNS express IL-18 and caspase-1 constitutively, thus providing a local IL-18-dependent immune response. Recent studies have highlighted a crucial role for IL-18 in mediating neuroinflammation and neurodegeneration in the CNS under pathological conditions, such as bacterial and viral infection, autoimmune demyelinating disease, and hypoxic-ischemic, hyperoxic and traumatic brain injuries. This review provides a synopsis of the current knowledge of IL-18-dependent mechanisms of action during acute neurodegeneration in immature and adult brains.