Tatsuo Kawashima

Cytotoxicity induced by inhibition of thioredoxin reductases via multiple signaling pathways: Role of cytosolic phospholipase A2α-dependent and -independent release of arachidonic acid

The thioredoxin (Trx) system, comprising Trx, the selenoprotein thioredoxin reductase (TrxR), and NADPH, functions as an antioxidant system. Trx has various biological activities including growth control and anti‐apoptotic properties, and the Trx system offers a target for the development of drugs to treat and/or prevent cancer. We evaluated the role of TrxR inhibition in the release of arachidonic acid (AA), cell toxicity, and intracellular signaling pathways in L929 mouse fibrosarcoma cells. Treatment with 1‐chloro‐2,4‐dinitrobenzene (DNCB, an inhibitor of TrxR) under conditions involving limited inhibition of TrxR activity in cells, released AA before causing cytotoxicity. Treatment with an inhibitor of p38 kinase, a downstream enzyme of the apoptosis signal‐regulating kinase 1 pathway, and pyrrophenone (an inhibitor of α‐type cytosolic phospholipase A2, cPLA2α) partially but significantly decreased the DNCB‐induced release of AA and cell death. The responses were much weaker in cPLA2α knockdown L929 cells. Exogenously added AA showed cytotoxicity. DNCB increased intracellular reactive oxygen species (ROS) levels, and butylated hydroxyanisole (an antioxidant) reduced DNCB‐induced ROS formation and cell toxicity but not the phosphorylation of p38 kinase and release of AA. Auranofin, another inhibitor of TrxR having a different formula, released AA resulting in toxicity in L929 cells. DNCB caused the release of AA and cytotoxicity in A549 human lung carcinoma cells, and caused p38 kinase‐dependent toxicity in PC12 rat pheochromocytoma cells. Our data suggest that a dysfunctional Trx system triggers multiple signaling pathways, and that the AA released by cPLA2α‐dependent and ‐independent pathways is important to cytotoxicity. J. Cell. Physiol. 219: 606–616, 2009.

Change in serum marker of oxidative stress in the progression of idiopathic pulmonary fibrosis

BACKGROUND Increased oxidative stress is supposed to be involved in the etiology of idiopathic pulmonary fibrosis (IPF). It was reported that oxidative stress values measured by a spectrophotometric technique (d-ROMs test) were significantly higher in IPF patients than in controls, and were negatively correlated with Forced Vital Capacity (FVC) and Carbon Monoxide Diffusing Capacity (DLCO). However, the relationship between progression of IPF over time and change in serum oxidative stress marker remains unclarified. AIMS This study aimed to investigate the change in serum oxidative stress marker during progression of IPF. SUBJECTS AND METHODS The levels of oxidative stress in blood samples of 43 treatment-naïve IPF patients were measured by the d-ROMs test. FVC and DLCO were measured concurrently. The changes in oxidative stress and pulmonary function were evaluated in 27 untreated patients 6 months later. Oxidative stress levels of 13 patients with acute exacerbation of IPF (AE-IPF) and 30 healthy controls were also evaluated. RESULTS Oxidative stress values [median, interquartile range (IQR); Carratelli units (U.CARR)] were significantly higher in 43 IPF patients than in controls (366, 339-443 vs. 289, 257-329, p < 0.01) and were significantly increased 6 months later in 27 untreated patients (353, 311-398 at baseline to 385, 345-417 at follow-up, p < 0.01). The increase in oxidative stress values (24.0, 6.0-49.0 U.CARR/6 months) was negatively correlated with baseline DLCO (rs = -0.44, p < 0.05) and FVC changes after 6 months (rs = -0.54, p < 0.01). Oxidative stress values were significantly higher in IPF patients with acute exacerbation than in those with stable disease (587, 523-667 vs. 366, 339-443 U.CARR, respectively; p < 0.01). CONCLUSIONS Serum oxidative stress values increased with disease progression in IPF patients.

Indomethacin induces cellular morphological change and migration via epithelial-mesenchymal transition in A549 human lung cancer cells: A novel cyclooxygenase-inhibition-independent effect

Levels of cyclooxygenase (COX)-2 and its metabolite prostaglandin E(2) (PGE(2)) are frequently increased in colon cancer and other cancers including lung cancer. Non-steroidal anti-inflammatory drugs are considered to have chemo-preventive effects on these diseases by reducing the biosynthesis of PGE(2) via their inhibition of COX-2. Although the COX-2/PGE(2) pathway may directly impact on lung carcinogenesis, some population-based cohort studies of NSAIDs showed no significant protective effects. In this study, using human non-small-cell lung cancer A549 cells, we examined the effects of indomethacin, a potent NSAID, on the growth and motility of lung cancer cells. Besides inhibiting PGE(2) production and cellular growth, indomethacin caused drastic morphological changes with a loss of stress fibers in a time- and dose-dependent manner. Interestingly, the change in cellular shape caused by indomethacin was not seen when the cells were treated with aspirin or diclofenac, two other NSAIDs, despite the concentrations used being sufficient to inhibit PGE(2) production. The indomethacin-induced morphological changes in A549 cells were accompanied by a reduction in levels of the adhesion molecule E-cadherin and a component of basal lamina, collagen IV, as well as an increase in the activity of a collagenase, matrix metalloprotease-9. Furthermore, indomethacin-induced shape changes resulted in enhanced motility via regulation of peroxisome proliferator-activated receptor γ. The dual effects of indomethacin, inhibition of cellular growth and enhancement of migration, would explain, to some extent, the difficulty in using this NSAID for lung cancer therapy.

VGCC antibody-positive paraneoplastic cerebellar degeneration presenting with positioning vertigo

A 70-year-old woman developed paraneoplastic cerebellar degeneration (PCD) due to P/Q-type and N-type voltage-gated calcium channel antibodies and small cell lung cancer, the main clinical manifestations of which were severe positioning vertigo and vomiting. Loss of the visual suppression of caloric nystagmus, spontaneous downbeat nystagmus, periodic alternating nystagmus, and positioning vertigo in our patient most probably corresponds to the cerebellar flocculus/paraflocculus lesion caused by PCD.

Expression of High-Affinity IgE Receptor (FcεRI) on Human Alveolar Macrophages from Atopic and Non-Atopic Patients

The expression of high-affinity IgE receptor (FcERI) on alveolar macrophages was examined in order to determine the association of alveolar macrophages with IgE-dependent inflammation. Immunostai

Decrease in uptake of arachidonic acid by indomethacin in LS174T human colon cancer cells; a novel cyclooxygenase-2-inhibition-independent effect.

Nonsteroidal anti-inflammatory drugs (NSAIDs) have chemopreventive activity and may be suitable for treatment of colorectal cancer. A popular and potent NSAID, indomethacin, is known to cause serious side-effects, for this reason its therapeutic usefulness is limited. However, these side-effects are likely to be attributed to the additional effects of indomethacin besides its cyclooxygenase inhibition. In this study, we examined the effect of indomethacin on arachidonic acid uptake using LS174T human colon cancer cells. We here show that treatment of LS174T cells with indomethacin reduced arachidonic acid uptake as well as reduced expressions of fatty acid translocase/CD36 and peroxisome proliferators-activated receptor gamma. Since arachidonic acid is a major substrate of inflammatory mediators such as prostaglandins and leukotrienes, we believe this novel effect of indomethacin may apply to new treatment strategies that aim to suppress these mediators by decreasing the uptake of their substrates, which would eventually inhibit colorectal cancer malignancy.

Evaluation of reactive oxygen metabolites in patients with non-small cell lung cancer after chemotherapy.

BackgroundThe aim of this study was to evaluate the level of reactive oxygen metabolites (ROMs) after chemotherapy in patients with non-small cell lung cancer (NSCLC) and its association with response to treatment.MethodsFifty-eight untreated NSCLC patients and twenty-three healthy subjects were selected for the study. Patients received two courses of platinum-based chemotherapy and were evaluated for oxidative stress and treatment response. As a marker of reactive oxygen species, ROMs levels were measured using the d-ROMs test.ResultsROMs level (mean ± standard deviation) before chemotherapy in NSCLC patients (416 ± 135 U.CARR) was significantly elevated (p = 0.016) compared to normal healthy subjects (320 ± 59 U.CARR). Patients who responded to chemotherapy showed significantly decreased (p = 0.014) ROMs levels after chemotherapy, whereas patients who had stable disease or progressive disease showed no change in ROMs level (p = 0.387).ConclusionsNSCLC patients had significantly elevated ROMs levels before chemotherapy compared with normal healthy subjects. Chemotherapy may suppress ROMs production in responders but not in non-responders. ROMs level may be a predictor of clinical outcome in patients receiving chemotherapy for NSCLC.

Contrary effects of sphingosine-1-phosphate on expression of α-smooth muscle actin in transforming growth factor β1-stimulated lung fibroblasts.

Transforming growth factor-β1 (TGFβ1) plays a pivotal role in fibrosis in various organs including the lung. Following pulmonary injury, TGFβ1 stimulates conversion of fibroblasts to myofibroblasts that are mainly characterized by up-regulation of α-smooth muscle actin (αSMA) expression, and the resulting excess production of extracellular matrix proteins causes fibrosis with loss of alveolar function. The present study was undertaken to define the role of the sphingosine-1-phosphate (S1P) pathway in TGFβ1-induced expression of αSMA in human fetal lung fibroblasts, HFL1 cells. Analysis of mRNA revealed the existence of S1P(1), S1P(2), and S1P(3) receptor mRNAs. Treatment with TGFβ1 increased sphingosine kinase (SphK) activity and S1P(3) receptor mRNA at 24h after stimulation, and pharmacological data showed the involvement of sphingomyelinase, SphK, and S1P(3) receptor in the TGFβ1-induced up-regulation of αSMA with and without serum. Treatment with pertussis toxin and S1P(1) receptor antagonist W146 enhanced αSMA expression by TGFβ1/serum, and S1P decreased and increased αSMA levels with and without serum, respectively. TGFβ1 increased cyclooxygenase-2 expression in a manner dependent on serum and the sphingomyelinase/SphK pathway, and the response was decreased by pertussis toxin. Prostaglandin E(2), formed by TGFβ1/serum stimulation, decreased the TGFβ1-induced expression of αSMA via EP prostanoid receptor. These data suggest that S1P formed by TGFβ1 stimulation has diverse effects on the expression of αSMA, inhibition via the S1P(1) receptor-mediated and serum-dependent expression of cyclooxygenase-2 and the resulting formation of prostaglandin E(2), and stimulation via the S1P(3) receptor in a serum-independent manner.

Lecithinized superoxide dismutase treatment improves steroid-refractory interstitial pneumonia.

Pulmonary fibrosis associated with amyopathic dermatomyositis is known to have a generally aggressive course and is ultimately fatal. We report the case of a 50‐year‐old patient with amyopathic dermatomyositis, who developed progressive interstitial pneumonia that was unresponsive to corticosteroids and multiple immunosuppressive agents, including cyclosporine and tacrolimus hydrate. Five courses of lecithinized superoxide dismutase were administered without adverse effects. Improvements in physiological parameters, such as pulmonary function and exercise tolerance, as well as the serum Krebs von den Lungen 6 level, were observed. This is the first report of a case of steroid‐refractory interstitial pneumonia treated with lecithinized superoxide dismutase.