Yoshiaki Hitomi

Uncoupling protein 2 plays an important role in nitric oxide production of lipopolysaccharide-stimulated macrophages

The expression of uncoupling protein 2 (UCP2) was reduced in macrophages after stimulation with lipopolysaccharide (LPS). The physiological consequence and the regulatory mechanisms of the UCP2 down-regulation by LPS were investigated in a macrophage cell line, RAW264 cells. UCP2 overexpression in RAW264 cells transfected with eukaryotic expression vector containing ucp2 cDNA markedly reduced the production of intracellular reactive oxygen species. Furthermore, in the UCP2 transfectant, nitric oxide (NO) synthesis, inducible NO synthase (NOS II) protein, NOS II mRNA, and NOS II promoter activity were definitely decreased after LPS stimulation compared with those in parental RAW264 or RAW264 cells transfected with the vector alone. Reporter assays suggested that an enhancer element was located in the region of intron 2 of the UCP2 gene and that the UCP2 expression was down-regulated not by the 7.3-kb promoter region but by the 5′ region of the UCP2 gene containing two introns. Deletion of intron 2 resulted in the low transcriptional activities and abolishment of the LPS-associated negative regulation. In addition, the mRNA expression of transfected UCP2 was suppressed in RAW264 cells transfected with expression vector containing UCP2 genomic DNA, but was markedly increased in cells transfected with the vector containing UCP2 intronless cDNA. These findings suggest that the LPS-stimulated signals suppress UCP2 expression by interrupting the function of intronic enhancer, leading to an up-regulation of intracellular reactive oxygen species, which activate the signal transduction cascade of NOS II expression, probably to ensure rapid and sufficient cellular responses to a microbial attack.

Acetaldehyde production by non‐pathogenic Neisseria in human oral microflora: Implications for carcinogenesis in upper aerodigestive tract

Many epidemiological studies have identified chronic alcohol consumption as a significant risk factor for cancer of the upper aerodigestive tract (UAT) in human. Although acetaldehyde, the first metabolite from ethanol by alcohol dehydrogenase (ADH), is regarded as a carcinogen, how systemic production of acetaldehyde particularly affects the UAT remains unclear. In our study, we searched for the regional source of acetaldehyde in UAT, especially the involvement of bacteria in the human normal oral microflora. Here we demonstrate that, among the bacterial species identified from the human oral cavity, genus Neisseria had extremely high ADH activity and produced significant amounts of acetaldehyde when cultured with medium containing ethanol in vitro. The ability to produce acetaldehyde was more than 100‐fold higher than that produced by any other genera we studied. Furthermore, alcohol ingestion influences the bacterial composition of the oral microflora, resulting in an increased proportion of Neisseria. Although Neisseria present in normal oral microflora is generally non‐pathogenic, these findings suggest that this microbe can be a regional source of carcinogenic acetaldehyde and thus potentially play an important role in alcohol‐related carcinogenesis in human UAT. Int. J. Cancer 88:342–350, 2000.

The role of Ets family transcription factor PU.1 in hematopoietic cell differentiation, proliferation and apoptosis

The PU.1 gene encodes an Ets family transcription factor which controls expression of many B cell- and macrophage-specific genes. Expression of the gene is critical for development of lymphoid and myeloid cell lineages, since PU.1-deficient mice exhibit defects in the development of these cell lineages. The PU.1 gene is identical to the Spi-1 gene isolated from common proviral integration sites in Friend virus-induced murine erythroleukemia (MEL), and deregulated expression of the gene is believed to be an essential step of the disease. We recently demonstrated that overexpression of PU.1 inhibits erythroid differentiation of MEL cells induced with the differentiating agent DMSO. We also noticed unexpectedly that overexpression of PU.1 together with DMSO induces marked growth arrest and apoptosis in MEL cells, supporting the notion that some oncogenes induce growth inhibition and apoptosis rather than cell proliferation and transformation under specific circumstances as shown with the c-myc gene. In this review, the role of PU.1 in hematopoietic cell differentiation, proliferation and apoptosis is described and the possible molecular mechanisms of PU.1-induced effects in MEL cells are discussed.

Muscle type-specific response of PGC-1α and oxidative enzymes during voluntary wheel running in mouse skeletal muscle

Aim:  It is generally accepted that endurance exercise increases the expression of peroxisome proliferator‐activated receptor γ coactivator‐1α (PGC‐1α), which governs the expression of oxidative metabolic enzymes. A previous report demonstrated that the regulation of mitochondrial protein expression in skeletal muscles in response to cold exposure depends on muscle fibre type. Cold exposure and endurance exercise are both metabolic challenges that require adjustments in mitochondrial energy metabolism, we hypothesized that the exercise‐induced increase in oxidative enzymes and PGC‐1α expression is higher in fast‐type than in slow‐type muscle.

Genetic variation in hypoxia-inducible factor 1α and its possible association with high altitude adaptation in Sherpas

Hypoxic stress at high altitude requires adaptations in several physiological functions to ensure the optimal oxygenation of all cells. Several lines of evidence suggested that high-altitude native populations such as Sherpas have been genetically adapted to their stressful environment. We investigated the genetic variation in the hypoxia-inducible factor (HIF)-1alpha gene in Sherpas as compared with Japanese, native lowlanders, and found a novel dinucleotide repeat polymorphism in intron 13 of the HIF-1alpha gene. GT15 allele was more frequent in Japanese than in Sherpas with statistical significance, while GT14 allele was significantly more frequent in Sherpas as compared with Japanese. A possible genetic variation in the HIF-1alpha gene might function in adaptation to living at high altitude. Because the activity of HIF-1 is regulated by multiple steps including the transcriptional level, the effect of the polymorphism in intron 13 on the cellular hypoxic responses remains to be elucidated.

Health impact of disaster‐related stress on pregnant women living in the affected area of the Noto Peninsula earthquake in Japan

Aims:  The present study assessed the health impact of stress on women who were pregnant during, or immediately after, a major earthquake and were living in the disaster area. Inherent resistance against the stress induced by the earthquake was also assessed.

Laminin 5 expression protects against anoikis at aerogenous spread and lepidic growth of human lung adenocarcinoma.

Adenocarcinoma of the lung is characterized by frequent aerogenous spread (AE) and advancement along the alveolar wall (BAC growth). To elucidate the mechanism of AE metastasis and BAC growth in human lung adenocarcinoma, we established an in vivo orthotopic animal model and an in vitro culture. Investigation of expression levels of integrins, laminins and Type IV collagens, which are the major regulating molecules for cell attachment and anoikis was carried out and a clear correlation between the expression level of laminin 5 (LN5) and the BAC growth was observed using an orthotopic animal model. Introduction of LN5 cDNA to A549 cells increased anoikis resistance in an expression dependent manner. Cells with LN5 overexpression resisted with anoikis after treatment with PI3K‐Akt and ERK inhibitors. The amount of phosphorylated focal adhesion kinase (FAK) was also higher in LN5 overexpressing cells. Major tyrosine residues of the EGF receptor at 1068, 1086 and 1173, except at 1148, remained phosphorylated only in the LN5 overexpressing cells even without EGF stimulation, that indicates the ligand independent activation of EGF receptor. BAC growth ratio and AE was confirmed to be significantly correlated with LN5 expression in surgically resected human lung adenocarcinomas by immunohistochemistry. Our results indicate that the activation of the EGF receptor by overexpressing LN5‐integrin‐FAK signaling pathway may play a crucial role in BAC growth and AE metastasis in human lung adenocarcinoma.

C-reactive protein is associated with cigarette smoking-induced hyperfiltration and proteinuria in an apparently healthy population

Although cigarette smoking is known to be an important risk factor for renal disease, the mechanism by which smoking induces progressive renal disease in a healthy population has not been established. We hypothesized that oxidative stress (measured as 8-iso-prostaglandin F2α, 8-iso-PGF2a), inflammation (highly sensitive C-reactive protein (CRP), hs-CRP) and nitric oxide may be associated with an alteration in the estimated glomerular filtration rate (eGFR) and proteinuria in otherwise healthy smokers. A total of 649 eligible subjects were classified according to their smoking status. Plasma NOx was measured using ozone-based chemiluminescence, urinary 8-iso-PGF2a was measured using enzyme immunoassay and serum hs-CRP was measured using a latex aggregation nephelometry method. The levels of 8-iso-PGF2a and hs-CRP increased in current smokers (P=0.001 and P=0.029, respectively), although there was not an increase in the NOx level. The prevalence of a high eGFR increased in light smokers (odds ratio (OR) 1.15 (95% confidence interval (CI), 0.61–2.17)) and heavy smokers (OR 2.33 (95% CI, 1.06–5.10)) when compared with non- and past smokers (P for trend=0.024). The multivariable-adjusted mean values of the eGFR in current smokers, reported from the lowest to the highest quintiles of hs-CRP levels, were 82.1, 85.1, 86.4 and 88.5 ml per min per 1.73 m2 (P for trend=0.027). The mean values of proteinuria were 28.6, 34.6, 37.2 and 39.5 mg g−1 creatinine (P for trend=0.003). The correlation coefficient between hs-CRP and eGFR was increased significantly (P=0.03) across non- (r=0.03), past (r=−0.17), light (r=0.13) and heavy smokers (r=0.31). In conclusion, cigarette smoking is a risk factor for renal function alteration in healthy smokers and is characterized by a high eGFR and a high urinary protein associated with an increase in the hs-CRP. This finding suggests that hs-CRP may help mediate the alteration of renal function in smokers.

Acute exercise increases expression of extracellular superoxide dismutase in skeletal muscle and the aorta.

Abstract Exercise dramatically increases oxygen consumption and causes oxidative stress. Superoxide dismutase (SOD) is important in the first-line defence mechanisms against oxidative stress. To investigate the effect of acute exercise on the expression of SOD, we examined the expression of mRNA for three SOD isozymes, in mice run on a treadmill to exhaustion. Six hours after exercise, the expression of extracellular SOD (EC-SOD) mRNA increased significantly in skeletal muscle and persisted for 24 h, whereas no change was observed for cytoplasmic and mitochondrial SOD mRNA. Moreover, acute exercise also induced EC-SOD mRNA in the aorta. These results suggest that a single bout of exercise is enough to augment the expression EC-SOD mRNA in skeletal muscle and the aorta, and may partly explain the beneficial effect of exercise.

Intermittent hypobaric hypoxia increases the ability of neutrophils to generate superoxide anion in humans

1. We investigated the effect of intermittent exposure to hypobaric hypoxia on the ability of neutrophils to generate ·O2–.