[Long-term results after the total cavopulmonary connection].

Between July, 1988 and November, 2002, 108 patients underwent total cavopulmonary connection (TCPC) at Kobe Children's Hospital. The primary malformation was univentricular heart in 40 tricuspid atresia in 21, mitral atresia in 16, and other complex cardiac defects in the remaining 31. Fenestrated TCPC, staged TCPC, and off-pump TCPC were performed in 39, 26, and 15 high risk patients, respectively. Nitric oxide inhalation was administered in 46 patients. The mean follow-up period was 4.3 years (range, 1 month to 14 years). There were 10 early deaths due to low cardiac output syndrome in 4, thrombosis in 3, tracheal bleeding in 2, and disseminated intravascular coagulation in 1. There were 5 late deaths due to congestive heart failure in 2 patients, arrhythmia in 1, cerebral infarction in 1, and subarachnoid hemorrhage in 1. Late complications included arrhythmia in 17 patients, systemic desaturation caused by abnormal systemic venous channels in 10, pleural or pericardial effusion in 3, chylothorax in 1, and aortic valve incompetence in 1.

A neurofibromatosis type 1 patient with severe kyphoscoliosis and intrathoracic meningocele.

The patient presented with neurofibromatosis and a dystrophic kyphoscoliosis around the cervico-thoracic junction. When the patient was 59 years old, he started to suffer from dyspnea caused by an intrathoracic meningocele in the upper left thoracic cavity. A wide laminectomy from T2 to T5 was performed and the meningocele was resected. Although the dyspnoea disappeared postoperatively, the patient started to neurologically deteriorate. Laminectomy alone caused instability around the apex of the kyphosoliosis and spinal cord compression. Halo cast was applied and brought remarkable recovery of neurologic deficits. This result encouraged us to perform posterior fusion in situ from C3 to L2 with bone graft from the iliac crests and the Luque technique in conjunction with the Isola system. This resulted in the patient being able to walk again. The removal of the posterior element predisposes the patient to unstable postlaminectomy kyphosis and removes valuable bone stock required for posterior spinal fusion. For this reason, spinal fusion should have been conducted during surgery for the patient's meningocele.

Involvement of NF-kappaB in the protection of cell death by tumor necrosis factor in L929 derived TNF resistant C12 cells.

The Tumor necrosis factor (TNF)-resistant C12 cell line was established by continuous exposure of a toxic concentration of TNF to parental murine fibrosarcoma L929 cells. Introduction of the cytosolic phospholipase A2 (cPLA2) gene to C12 cells resulted in restoration of the TNF sensitive phenotype (CPL4 cells). DNA ladder formation and nuclear condensation by TNF exposure suggested that TNF induced apoptotic cell death in L929, C12 and CPL4 cells. TNF-induced activation of transcription factor nuclear factor-kappaB (NF-kappaB) was observed in all 3 cell lines. The activation reached the maximum level at 30 min after the exposure to TNF and thereafter declined slowly. The amount of activated NF-kappaB in C12 cells was about twice as high as that of L929 cells with either dose of TNF tested in this study. It was found that C12 cells expressed latent NF-kappaB twice that of L929 cells. This abundance of latent NF-kappaB would provide a higher response of NF-kappaB in C12 cells. Pretreatment with the known potent NF-kappaB inhibitor pyrolidine dithiocarbamate (PDTC) profoundly suppressed the activation of NF-kappaB induced by TNF and potentiated TNF cytotoxicity in all 3 cell lines. These results are consistent with the recently found anti-apoptotic action of NF-kappaB and suggest that NF-kappaB acts as an acquired TNF resistant factor in C12 cells.

Reduced sensitivity of HeLa cells to cis-platinum by simultaneous overexpression of copper, zinc-superoxide dismutase and catalase.

The overexpression of catalase or Cu,Zn-superoxide dismutase (Cu,Zn-SOD) did not affect the sensitivity of HeLa cells to cis-platinum. However, the cytotoxicity of cis-platinum was depressed significantly by the simultaneous overexpression of catalase and Cu,Zn-SOD. We concluded that cis-platinum accelerated the generation of superoxide anion in the cells, and the superoxide anion produced was converted into H2O by the cooperative roles of catalase and Cu,Zn-SOD.

Effect of intracellular glutathione on the production of prostaglandin D2 in RBL-2H3 cells oxidized by tert-butyl hydroperoxide.

The liberation of arachidonic acid and the production of prostaglandin D2 (PGD2) were significantly influenced by peroxide and the level of intracellular glutathione (GSH). The productions of free arachidonic acid and PGD2 in RBL-2H3 cells were enhanced considerably by exposure to tert-butyl hydroperoxide (t-BHP). The liberation of arachidonic acid induced by t-BHP was not inhibited by EGTA. The productions of PGD2 and arachidonic acid induced by t-BHP were significantly facilitated by the depletion of intracellular GSH using buthionine sulfoximine (BSO) or diethyl maleate (DEM), although the depletion of GSH had no effect on the production of PGD2 induced by A23187. t-BHP failed to activate the conversion of free arachidonic acid to PGD2, since the formation of PGD2 from exogenously added arachidonic acid was not enhanced by treatment with t-BHP. The level of lipid hydroperoxides in t-BHP-treated cells was significantly elevated by treatment with DEM. These results suggest that hydroperoxides increase the free arachidonic acid available for the synthesis of PGD2 by activating phospholipase A2 (PLA2) and that the depletion of GSH by DEM accelerates the activation of PLA2 by raising peroxide levels in cells. Thus, the observed alterations in GSH levels are large enough to cause increased PGD2 synthesis in RBL-2H3 cells exposed to oxidative stress.

Brain stem frequency-following response to dichotic vowels during attention.

Frequency-following responses (FFRs) were elicited by English long vowels (female /a/ and male /e/) in a dichotic listening task. Stimuli were simultaneous and of equal duration, but differing spectra permitted unique identification of vowel components in the compound FFR. Horizontal and vertical montage FFRs were recorded with putative origins in the acoustic nerve and central brain stem, respectively. FFRs obtained during attention to each vowel showed significant effects for the voice fundamental frequency, f0, which is perceptually salient and conveys paralinguistic information such as the sex of the speaker. Amplitudes of f0 were larger when vowels were attended than when ignored. These findings provide evidence of short-latency attention effects in humans and suggest that linguistic attention may initially filter inputs based on salient paralinguistic cues.

Overexpression of manganese-superoxide dismutase prevents methylmercury toxicity in HeLa cells.

HeLa cells were stably transformed with plasmid constructs that allowed constitutive expression of antioxidant enzymes such as catalase, glutathione peroxidase (GSH-Px), Cu,Zn-superoxide dismutase (Cu,Zn-SOD) or Mn-superoxide dismutase (Mn-SOD) to examine the involvement of reactive oxygen generation in methylmercury toxicity. Overexpression of catalase, GSH-Px or Cu,Zn-SOD did not affect the sensitivity of HeLa cells against methylmercury. However, the sensitivity of HeLa cells against methylmercury was decreased by overexpression of Mn-SOD, an enzyme localized in matrix of mitochondria and which decomposes superoxide anions. These results suggest that formation of superoxide anions in the mitochondria might be involved in the mechanism of the cytotoxicity of methylmercury.

Mutagenicity of cobalt and reactive oxygen producers.

Oxidative stress has been implicated in carcinogenesis yet there are chemicals that produce oxidative stress that are not carcinogenic. Mutations are the inherited results of DNA damage and are critical events in carcinogenesis. The mutagenicity of oxidative stress induced by peroxide, paraquat and cobalt compounds was examined in transgenic gpt+ Chinese hamster cell lines (G12 and G10). These two cell lines are known to be more sensitive to mutagens such as X-rays and UV than their parental V-79 cells. In these studies, the mutagenic activity of cobalt chloride, a metal that induces oxidative stress but is not carcinogenic, was measured to be 7.7 times higher than the spontaneous mutant frequency in G12, but was only 1.5 to 2.5 times higher than spontaneous mutant frequency in G10 cells. The mutant frequency of cobalt sulfide was somewhat lower. Hydrogen peroxide was found to be only weakly mutagenic in G12 cells, and treatment of cells with a combination of hydrogen peroxide and cobalt did not alter the mutation frequency induced by cobalt sulfide alone. Paraquat did not elicit mutagenesis in either cell line. These results indicate that agents producing oxidative stress are not necessarily mutagenic and these results are discussed in the context of the oxidative stress produced by other carcinogens such as nickel compounds.

Heterochromatic proteins specifically enhance nickel-induced 8-oxo-dG formation.

7,8-Dihydro-8-oxo-2'-deoxyguanosine (8-oxo-dG) was measured as an indicator of nickel-induced oxidative base damage in the presence of H2O2. Heterochromatic proteins isolated from Chinese hamster liver cells enhanced the formation of 8-oxo-dG induced by NiCl2 and H2O2 in vitro, whereas euchromatic proteins inhibited this reaction. The inhibitory effect of euchromatic proteins on dG oxidation may be due to the oxygen radical scavenging effects of low molecular weight protein-rich fractions. Gel electrophoresis confirmed that histone H1 was present at a higher concentration in heterochromatin than in euchromatin. It is believed that the presence of nickel-protein complexes in cells is crucial for the formation of reactive oxygen species (ROS). We found that Ni2+ binds to histone H1 and core histones as determined by 63Ni autoradiography of proteins on nitrocellulose membranes. In vitro studies showed that commercially purified histone H1, and to a considerably lesser extent core histones, enhanced the NiCl2 and H2O2 catalyzed formation of 8-oxo-dG in a reaction containing free dG base. Since histone H1 is a lysine- and alanine-rich protein, the levels of 8-oxo-dG induced by NiCl2 and H2O2 were studied in the presence of these amino acids and found to be enhanced by them. These results suggest that nickel may specifically produce oxidative DNA damage in heterochromatin because of the nature of its binding to histone H1 and core histones. This selective oxidation of genetically inactive heterochromatin may explain why nickel compounds which generate oxygen radicals and oxidize DNA bases are inactive in most gene mutation assays.

[Relationship between proliferative activity, and tissue differentiation and invasive mode in human oral squamous cell and colorectal carcinomas analysed by PCNA immunostaining].

The proliferative activity of carcinoma cells is generally considered to relate to the degree of the malignancy of carcinoma tissues. In this study, the proliferative activity at the tumor-stromal border was studied in 17 cases of oral squamous cell carcinoma (OSCC) and in 30 cases of colorectal adenocarcinoma (CAC) by means of proliferating cell nuclear antigen (PCNA) immunostaining, to evaluate the correlation between proliferative activity and tissue differentiation or invasive mode at the tumor-stromal border. No statistical difference was detected between the PCNA labelling index (PI) and the tissue differentiation of both OSCC and CAC. A significant difference was demonstrated between PI and invasive mode in OSCC, suggesting that the invasive mode at the tumor-stromal border relate to the degree of the malignancy of carcinoma tissues. However, no significance was found between PI and invasive mode of CAC. In addition, no difference of PI was demonstrated between tissue differentiation or invasive mode, and vascular invasion or lymph node metastasis. Therefore, it seems likely that the invasive mode at the tumor-stromal border in CAC also has no significance in deciding the degree of the malignancy of carcinoma tissues.

Carcinogenic nickel silences gene expression by chromatin condensation and DNA methylation: a new model for epigenetic carcinogens.

A transgenic gpt+ Chinese hamster cell line (G12) was found to be susceptible to carcinogenic nickel-induced inactivation of gpt expression without mutagenesis or deletion of the transgene. Many nickel-induced 6-thioguanine-resistant variants spontaneously reverted to actively express gpt, as indicated by both reversion assays and direct enzyme measurements. Since reversion was enhanced in many of the nickel-induced variant cell lines following 24-h treatment with the demethylating agent 5-azacytidine, the involvement of DNA methylation in silencing gpt expression was suspected. This was confirmed by demonstrations of increased DNA methylation, as well as by evidence indicating condensed chromatin and heterochromatinization of the gpt integration site in 6-thioguanine-resistant cells. Upon reversion to active gpt expression, DNA methylation and condensation are lost. We propose that DNA condensation and methylation result in heterochromatinization of the gpt sequence with subsequent inheritance of the now silenced gene. This mechanism is supported by direct evidence showing that acute nickel treatment of cultured cells, and of isolated nuclei in vitro, can indeed facilitate gpt sequence-specific chromatin condensation. Epigenetic mechanisms have been implicated in the actions of some nonmutagenic carcinogens, and DNA methylation changes are now known to be important in carcinogenesis. This paper further supports the emerging theory that nickel is a human carcinogen that can alter gene expression by enhanced DNA methylation and compaction, rather than by mutagenic mechanisms.

Possible involvement of active oxygen species in selenite toxicity in isolated rat hepatocytes.

Mechanisms of selenite cytotoxicity were examined using isolated rat hepatocytes. When selenite was added to a suspension of rat hepatocytes, intracellular reduced glutathione (GSH) was decreased and the oxygen consumption rate was increased. Subsequently, thiobarbituric acid-reactive substances (TBA-RS) and lactate dehydrogenase (LDH) leakage were increased. A ferric iron chelator, desferrioxamine (DF), and a synthetic superoxide dismutase (SOD) mimic, desferrioxamine manganese (DFMn), reduced the selenite toxicity. These results suggest that superoxide anion and its reactive metabolites such as the hydroxyl radical may be involved in the cytotoxicity of selenite.

99mTc-methylene diphosphonate uptake by ossifications and calcifications of non-osseous metastatic tumors.

Two cases of extra-osseous uptake of 99mTc-methylene diphosphonate (MDP) by non-osseous metastatic tumors are reported. One was a metastasis with ossification in the abdominal wall from carcinoma of the sigmoid colon and the other was a metastasis with calcification from an ovarian carcinoma. The mechanism of extra-osseous uptake of 99mTc-MDP is discussed. Bone scintigraphy can be a potential means to assess tumor spread with ossifications and calcifications.