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Terje Rootwelt

Publications and source records attributed to Terje Rootwelt.

15 recordsLinked to original sources
PubMed2006

Expression of complement regulators and receptors on human NT2-N neurons--effect of hypoxia and reoxygenation.

Complement activation can cause tissue damage in cerebral stroke by the release of biologically potent activation products and impaired function of regulatory proteins. We investigated the constitutive and hypoxia-reoxygenation-dependent expression of complement receptor 1 (CD35), membrane cofactor protein (CD46), decay-accelerating factor (CD55), protectin (CD59), and complement C3a and C5a receptors (C3aR and C5aR) on human NT2-N neurons. The effect of hypoxia-reoxygenation on C3d-deposition on neurons and endothelial cells was also investigated. NT2-N neurons were examined by cellular enzyme-linked immunosorbent assay and immunofluorescence microscopy. Endothelial cells were examined by flow cytometry. Three hours 1% or 0.1% hypoxia and 21h reoxygenation with 50% AB-serum were used to investigate the effect of hypoxia-reoxygenation on regulators and C3d-deposition. NT2-N neurons expressed significant amounts of CD59 (Clone H19/Clone BRIC229: p=0.000006/p=0.000003), CD46 (p=0.00006), CD55 (p=0.003) and C3aR (p=0.00003). CD35 and C5aR were not significantly expressed. There were no effects of hypoxia-reoxygenation on any of the regulators or receptors after 1% hypoxia and reoxygenation. However, CD55 (p=0.02) was down-regulated after 0.1% hypoxia and subsequent reoxygenation with AB-serum. There were no difference observed in the C3d-deposition during hypoxia-reoxygenation in either neurons or endothelial cells. In conclusion, human NT2-N neurons constitutively express C3aR, CD46, CD55 and, in particular, CD59. The cells may respond to locally produced C3a and, at the same time, be well protected against complement attack. Although severe hypoxia-reoxygenation may down-regulate CD55 expression, it does not seem to influence C3d-deposition.

Cell Hypoxia↗
PubMed2006

Effect of acidosis on IL-8 and MCP-1 during hypoxia and reoxygenation in human NT2-N neurons.

Inflammation probably plays a significant role in perinatal brain injury. To study the contribution of locally produced cytokines, the effect on cell death of addition of IL-8 and MCP-1 or antibodies to these, and the impact of acidosis, human postmitotic NT2-N neurons were exposed to 3 h of hypoxia and glucose deprivation and reoxygenated for 21 h. After 3 h of hypoxia with neutral medium, IL-8 was significantly increased compared to controls (150 (100-250)% vs. 100 (85-115)%, p=0.023). After 21 h of neutral reoxygenation, both IL-8 (380 (110-710)% vs. 150 (85-260)%, p=0.041) and monocyte chemoattractant protein-1 (MCP-1) (650 (440-2000)% vs. 310 (230-340)%, p=0.007) were significantly increased compared to controls. After 3 h of hypoxia, both IL-8 (p=0.002) and MCP-1 (p=0.008) were significantly lower in cells with acidotic compared with cells with neutral medium. Acidosis during reoxygenation, however, significantly increased IL-8 release, whereas MCP-1 release was diminished. Similar effects of acidosis were seen in normoxic controls. The cells also secreted RANTES and IP-10, but not 8 other cytokines tested. We found no effect on cell death, measured by MTT assay, of addition of IL-8, MCP-1 or antibodies to these. We conclude that human NT2-N neurons release IL-8 and MCP-1 during 21 h of reoxygenation after 3 h of hypoxia. Acidosis led to a differential effect on IL-8 and MCP-1, with increased IL-8 and decreased MCP-1, both during reoxygenation and in normoxic controls. IL-8 and MCP-1 had no effect on cell death.

Acidosis↗
PubMed2006

Recovery after short-term bilirubin exposure in human NT2-N neurons.

We used human NT2-N neurons to investigate delayed effects of short-term exposure to unconjugated bilirubin (UCB). Cell viability was evaluated with MTT reduction assays and nuclear morphology. A 6-h exposure to 1, 5, or 25 microM UCB and serum deprivation (SED) significantly diminished MTT reduction. 96 h after rescue of neurons with removal of UCB and re-incubation in the original serum-containing medium, delayed effects were evident as recovery (1 microM UCB), intermediate cell death (5 microM UCB), or near complete cell death (25 microM UCB). The impact of 6 h of SED alone appeared to be modest in rescued neurons. In this model, co-treatment with the specific caspase-3 inhibitor, zDEVD.FMK (100 microM), or the pancaspase inhibitor zVAD.FMK (100 microM) did not improve viability in rescued neurons exposed to 5 microM UCB, while treatment with the NMDA receptor antagonist MK-801 (1 microM) enhanced the number of undamaged nuclei (86 +/- 14% versus 50 +/- 12%, P = 0.001). MK-801 had, however, no impact on MTT reduction. In a different model with a 102-h continuous exposure to UCB and SED, we found a significant additional toxic impact of serum deprivation. Separate experiments suggested that this was a result of late caspase-mediated toxicity. We conclude that UCB-mediated effects may be reversible in this model. Blockade of excitotoxic mechanisms, but not caspase activity may prevent delayed cell death.

Apoptosis↗
PubMed2006

[Mitochondrial beta-oxidation defects].

BACKGROUND: Mitochondrial beta-oxidation of fatty acids is an important source of energy for the cells, especially during fasting. Since 1973 several inherited defects in beta-oxidation have been described. Defects in mitochondrial beta-oxidation are one of the largest groups of inborn errors of metabolism. MATERIAL AND METHODS: This review article is based on the experience of the authors and on literature studies. The authors' experience is from laboratory diagnostics and clinical experience in the departments of medical biochemistry and peadiatrics at our hospital. RESULTS AND INTERPRETATION: Beta-oxidation defects are potentially fatal disorders. Symptoms are usually seen during fasting, e.g. during childhood infections. Organs which preferably oxidize fatty acids or ketone bodies are especially vulnerable. Often, but not always, the patients have hypoketotic hypoglycaemia. In addition one can see affection of the liver, heart, muscular and nervous systems. The diseases can manifest both in childhood and adulthood and are often less severe in adulthood. The main principles of symptomatic treatment are avoidance of fasting and regular intake of a low-fat, high-carbohydrate diet. The diagnosis can be difficult to establish, especially in asymptomatic phases.

Adult↗
PubMed2006

Dexamethasone induces cell death which may be blocked by NMDA receptor antagonists but is insensitive to Mg2+ in cerebellar granule neurons.

Since dexamethasone may elevate the Ca2+ influx through NMDA receptors, we have investigated mechanisms of dexamethasone toxicity in rat cerebellar granule neurons. Dexamethasone concentrations over 0.1 microM induced cell death that reached about 20% of the death induced by glutamate. Dexamethasone-induced cell death was reduced by more than 80% by the mineralocorticoid antagonist RU 28318 or the NMDA receptor antagonists MK 801 and CGP 39551, whereas RU 28318 rescued only approximately 30% of cells treated with glutamate, indicating that dexamethasone requires NMDA receptors to induce acute neuronal toxicity and that a fraction of the neurons showed this toxicity. Mg2+ reduced the cell death induced by glutamate at potassium concentrations of 1 mM and 5 mM, but not at 25 mM. In contrast, cell death induced by dexamethasone was not significantly reduced by Mg2+ in any of the potassium concentrations. Both glutamate and dexamethasone induced toxicity with translocation of the apoptosis inducer NGFI-B to the mitochondria seen after 30 min-2 h concomitant with activation of apoptosis inducing factor (AIF) and caspase-3. In conclusion, dexamethasone induces a rapid toxicity which is blocked by NMDA receptor antagonists other than Mg2+, and involves mitochondrial apoptosis inducer NGFI-B.

Animals↗
PubMed2005

Synergistic protection of a general caspase inhibitor and MK-801 in bilirubin-induced cell death in human NT2-N neurons.

Unconjugated bilirubin (UCB) induces both apoptosis and necrosis in neurons. To investigate the role of caspases and excitotoxicity in UCB-induced cell death, we exposed NT2-N neurons to 5 microM UCB (a concentration known to induce apoptosis) or 2 microM staurosporine (positive apoptosis control) and investigated the effects of treatments with the specific caspase-3 inhibitor, zDEVD.FMK (20 and 100 microM), or the general caspase inhibitor, zVAD.FMK (20 and 100 microM), and/or the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 (10 microM) during a 24- or 48-h exposure. UCB increased caspase-3 activity 2.3-fold after 6 h. Despite this, treatment with zDEVD.FMK did not prevent cell death. zVAD.FMK enhanced neuronal survival by reducing apoptotic nuclear fragmentation, while MK-801 enhanced survival by reducing apoptotic nuclear condensation; both without affecting the MTT assays. Combined treatment reduced both apoptotic morphologies (without affecting necrosis), and this effect was also reflected in the MTT assays [corrected] We conclude that NMDA receptor-mediated pathways and caspase-mediated pathways are involved in UCB-induced cell death in human NT2-N neurons. Concomitant inhibition of both pathways results in synergistic protection.

Amino Acid Chloromethyl Ketones↗
PubMed2005

Acidosis during reoxygenation has an early detrimental effect on neuronal metabolic activity.

We recently showed that acidosis is protective during hypoxia and detrimental during reoxygenation. We hypothesized that the detrimental effect of acidosis during reoxygenation was due to a negative effect on mitochondrial function. Human postmitotic NT2-N neurons were exposed to 3 h of hypoxia and glucose deprivation and then reoxygenated for 0, 1, 4, 9, or 21 h. The detrimental effect of acidotic reoxygenation on metabolic activity was evident already after 1 h of reoxygenation, when MTT [3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide] reduction (percentage of normoxic controls) was significantly higher in cells reoxygenated with neutral compared with acidotic medium both after acidotic hypoxia (83+/-26% versus 67+/-27%, p=0.006) and after neutral hypoxia (51+/-12% versus 41+/-7%, p=0.005). Hypoxanthine, a marker of cellular energy failure, increased more with acidotic compared with neutral reoxygenation both after acidotic hypoxia (after 21 h: 7.7+/-2.7 versus 3.1+/-1.9 microM, p<0.001) and after neutral hypoxia (10.4+/-2.6 versus 7.9+/-2.8 microM, p=0.001). During hypoxia and reoxygenation, there was an earlier reduction in the activity of complex IV compared with complexes II+III, and the ratio between these complexes fell during the first hour of reoxygenation. The reduction in complex IV activity was alleviated with acidotic hypoxia. Acidosis during reoxygenation, however, had no effect on the activity of either complex IV or complexes II+III. We conclude that acidosis during hypoxia increases neuronal survival and preserves complex IV activity. Acidosis during reoxygenation has an early detrimental effect on metabolic activity, but this is not mediated through an effect on the mitochondrial complexes IV or II+III.

Acidosis↗
PubMed2005

Response to resuscitation of the newborn: early prognostic variables.

AIM: To characterize the development of clinically relevant variables the first minutes after birth and identify early prognostic markers in newborn infants requiring resuscitation. METHODS: A database of 591 infants resuscitated with either 21% or 100% oxygen was analysed. Time to first breath, development in heart rate, Apgar scores, arterial oxygen saturation (SaO(2)), and base deficit (BD) are described in relation to different degrees of birth depression and outcomes. RESULTS: Heart rate and Apgar scores increased quickly even in the most depressed infants but were significantly lower in those having a poor outcome. By contrast, BD normalized at the same rate, 6-7 mmol/l/h, in the first hour of life regardless of the degree of birth depression and outcome. SaO(2) values increased as quickly in room air as in 100%-oxygen-resuscitated infants. Time to first breath was prolonged threefold, from 1 to 3 min, in the most depressed (1-min Apgar score < 4) compared with the less depressed infants. Highest odds ratio (OR) for death in the first week of life or for development of hypoxic-ischaemic encephalopathy (HIE) stage 2 and 3 was a 5-min heart rate < or =60 bpm (OR 16.5 for both death and HIE) and Apgar < 4 (OR 14 and 18.8). Neonatal survival for HIE stage 1, 2, and 3 was 93%, 63%, and 11%, respectively. OR for early neonatal death, if SaO(2) < or =60% at 1 min, was 8.6 (sensitivity 0.82 and specificity 0.65). CONCLUSION: Apgar scores, heart rate, SaO(2), and time to first breath in newly born infants in need of resuscitation may be used for early identification of infants with a poor prognosis. These data may be helpful in describing the severity of depression in single infants and to select infants in need of interventional therapy.

Air↗
PubMed2004

Bilirubin induces apoptosis and necrosis in human NT2-N neurons.

Studies on primary cultures of newborn rodent neurons have suggested that neuronal death induced by unconjugated bilirubin (UCB) is mainly apoptotic in nature. We exposed a human teratocarcinoma-derived cell line, NT2-N neurons, to different concentrations of UCB and albumin at a 1.5 molar ratio and used multiple, independent measures of cell damage to evaluate neuronal injury after 6, 24, and 48 h. Low doses of UCB (0.66, 2, and 5 microM) induced a moderate loss of 3-4[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazoliumbromide (MTT) cleavage accompanied by delayed morphologic changes consistent with apoptosis (2 and 5 microM). Moderate concentrations of UCB (10 and 25 microM) resulted in early (6 h) necrosis in a subset of neurons, while remaining neurons underwent progressive impairment of MTT cleavage and increasing lactate dehydrogenase (LDH) release accompanied by predominantly delayed apoptosis. High concentrations of UCB (100 microM) induced severe impairment of MTT cleavage, extensive LDH release, and morphologic changes consistent with necrosis within 6 h. Used as a positive control for apoptosis, 2 microM STS induced progressive impairment of MTT cleavage and morphologic changes consistent with apoptosis over the entire observation period. DNA electrophoresis at 48 h was compatible with apoptosis both after treatment with STS and UCB concentrations <or=25, but not at 100 microM. Cleavage of poly (ADP-ribose) polymerase (PARP) was only seen in neurons treated with low UCB concentrations and STS. We conclude that UCB induces early necrosis at high and moderate concentrations and predominantly delayed apoptosis at low and moderate concentrations in cultured human NT2-N neurons.

Apoptosis↗
PubMed2004

Induction of Cbeta splice variants and formation of novel forms of protein kinase A type II holoenzymes during retinoic acid-induced differentiation of human NT2 cells.

Cyclic AMP (cAMP) and cAMP-dependent protein kinase (PKA) are critical regulators of neuronal differentiation. The expression, levels and activities of PKA subunits were studied prior to and during differentiation of the human neuronal precursor cell line NTera 2 (NT2). Undifferentiated NT2 cells expressed mainly cytoplasmic PKA type I, consisting of the regulatory subunit RIalpha and the catalytic subunit Calpha. Low levels of PKA type II consisting of RIIalpha or RIIbeta associated with Calpha were also detected, mainly in the cytoplasm and in the Golgi-centrosomal area. During retinoic acid-induced differentiation, the RIalpha and RIIalpha expressions remained in the cytoplasm, while we observed a strong upregulation of RIIbeta, located to the whole cytoplasm including neurite extensions. This upregulation coincided with increased PKA-specific activity accompanied by a strong induction of a number of neuronal-specific Cbeta splice variants that together with RIIbeta form novel PKAII holoenzymes. Formation of novel PKAII holoenzymes may imply specific PKA features which may have consequences for the process of neuronal differentiation and nerve cell function.

Alternative Splicing↗
PubMed2003

Severe phenotype of phosphorylase kinase-deficient liver glycogenosis with mutations in the PHKG2 gene.

Phosphorylase kinase-deficient liver glycogenosis manifests in infancy with hepatomegaly, growth retardation, and elevated plasma aminotransferases and lipids. It can be caused by mutations in three different genes of phosphorylase kinase subunits: PHKA2, PHKB, and PHKG2. It is usually a benign condition, often with complete resolution of symptoms during puberty. A minority of patients displays a more severe phenotype with symptomatic fasting hypoglycemia and abnormal liver histology that may progress to cirrhosis. Three patients with liver cirrhosis in childhood analyzed previously all had PHKG2 mutations. This suggested that this genotype may generally cause a more severe clinical manifestation, but to date PHKG2 mutations have been identified in only seven patients. Here, we report mutation analysis in three new patients with liver phosphorylase kinase deficiency and recurrent hypoglycemia, liver fibrosis, and lack of glucagon response but no overt cirrhosis. In all three patients, PHKG2 mutations were found (H89fs[insC], E157K, D215N, W300X). Three of these mutations are novel, bringing the total number of distinct human PHKG2 mutations to 11, found in 10 patients. We conclude that liver phosphorylase kinase deficiency with a severe phenotype, with or without cirrhosis, is indeed often caused by PHKG2 mutations. These patients require active measures to maintain normoglycemia (raw cornstarch, nocturnal tube feeding), which may also alleviate growth retardation and the development of abnormal liver histology.

Female↗
PubMed2003

Acidosis has opposite effects on neuronal survival during hypoxia and reoxygenation.

To study the effect of extracellular acidosis on apoptosis and necrosis during ischemia and reoxygenation, we exposed human post-mitotic NT2-N neurones to oxygen and glucose deprivation (OGD) followed by reoxygenation. In some experiments, pH of the cell medium was lowered to 5.9 during either OGD or reoxygenation or both. Staurosporine, used as a positive control for apoptosis, caused Poly(ADP-ribose)-polymerase (PARP) cleavage and nuclear fragmentation, but no PARP cleavage and little fragmentation were seen after OGD. Low molecular weight DNA fragments were found after staurosporine treatment, but not after OGD. No protective effect of caspase inhibitors was seen after 3 h of OGD and 21 h of reoxygenation, but after 45 h of reoxygenation caspase inhibition induced a modest improvement in 3-(4,5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium bromide (MTT) cleavage. While acidosis during OGD accompanied by neutral medium during reoxygenation protected the neurones (MTT: 228 +/- 117% of neutral medium, p < 0.001), acidosis during reoxygenation only was detrimental (MTT: 38 +/- 25%, p < 0.01). We conclude that apoptotic mechanisms play a minor role after OGD in NT2-N neurones. The effect of acidosis on neuronal survival depends on the timing of acidosis, as acidosis was protective during OGD and detrimental during reoxygenation.

Acidosis↗
PubMed2003

Resuscitation of newborn infants with 21% or 100% oxygen: follow-up at 18 to 24 months.

OBJECTIVE: To follow-up children who had been resuscitated at birth with either 21% or 100% oxygen (O2). METHODS: A multicenter study with 10 participating centers recruited 609 infants to the Resair 2 study where resuscitation was performed with either 21% or 100% O2. A follow-up between ages 18 and 24 months was performed. However, during follow-up registration, it was found that 18 infants had been enrolled twice in the original Resair 2 study with different registration numbers, leaving 591 enrolled in the Resair 2 study and 410 enrolled in the 7 centers participating in the follow-up. Of these 410 infants, 79 died (76 in the neonatal and 3 in the postneonatal period). Furthermore, for 8 infants informed consent was not obtained, leaving 323 eligible for follow-up. Of these, 213 infants (66%) were followed-up: 91 (62%) had been resuscitated with 21% O2, and 122 (69%) with 100% O2. At a median age of 22 and 20 months (not significant) in the 21% and 100% groups, respectively, a simple questionnaire was filled out and neurologic assessment was performed in addition to measuring anthropometric data. RESULTS: There were no significant differences in weight, height, or head circumference between the 2 groups. Cerebral palsy developed in 10% and 7%, respectively, in the 2 groups (not significant). In total, 11 cases (12%) in the 21% versus 11 cases (9%) in the 100% O(2) group (odds ratio: 1.39, 95% confidence interval: 0.57-3.36) developed cerebral palsy and/or mental or other delay. Furthermore, it was concluded that 14 (15%) in the 21% group and 12 (10%) in the 100% group were not normal (odds ratio: 1.67, 95% confidence interval: 0.73-3.80). CONCLUSIONS: There were no significant differences in somatic growth or neurologic handicap at an age of 18 to 24 months in infants resuscitated with either 21% or 100% O2 at birth. Based on these data, resuscitation with ambient air seems to be safe, at least in most cases. More studies are needed to settle this issue.

Birth Weight↗
PubMed2002

Neuronal formation of free radicals plays a minor role in hypoxic cell death in human NT2-N neurons.

Free radicals are suggested to play an important role in hypoxic-ischemic neuronal death. However, the importance in human disease is not known. Furthermore, whether posthypoxic free radical formation mainly occurs in endothelium and neutrophils, or whether neuronal production is important, is not finally determined. To study this we differentiated human Ntera2 teratocarcinoma cells into postmitotic NT2-N neurons and exposed them to free radicals, hypoxia, or oxygen and glucose deprivation. These cells are devoid of nitric oxide synthase, and we hypothesized that free radicals are important mediators downstream of N-methyl-D-aspartate stimulation. Production of free radicals, evaluated with the fluorescent dyes dihydrorhodamine and 2',7'-dichlorodihydrofluorescein, was significantly higher in neurons deprived of oxygen and glucose after 40 min of reoxygenation than in normoxic cells. The antioxidant trolox, the flavonoid quercetin, thiopental, and the N-methyl-D-aspartate-glutamate receptor antagonist MK-801 reduced the formation of free radicals. Treatment with the flavonoid rutin (86 +/- 16% of hypoxic cells without drug, p < 0.01), trolox (86 +/- 20%, p < 0.01), and MK-801 (57 +/- 12%, p < 0.01) reduced lactate dehydrogenase release after 6 h of hypoxia. Trolox, salicylate, and quercetin also significantly reduced lactate dehydrogenase release after 3 h of oxygen and glucose deprivation. The protection offered by these antioxidants was, however, limited compared with the effect of MK-801. We conclude that oxygen and glucose deprivation causes a moderate increase in the formation of free radicals in NT2-N neurons that can be inhibited by antioxidants and by blocking of the N-methyl-D-aspartate-glutamate receptor. Although MK-801 conveys profound protection, antioxidants provide only a limited improvement in neuronal survival. Thus in this model, mechanisms downstream of the N-methyl-D-aspartate-glutamate receptor other than free radicals and nitric oxide have to be invoked.

Antioxidants↗