Yoshinobu Iwasaki

Spiral CT findings in septic pulmonary emboli

OBJECTIVES The aim of the study was to determine the characteristics of septic pulmonary emboli and their prevalence on spiral computed tomographic (CT) scans. METHODS AND MATERIALS We evaluated 65 lesions on spiral CT scans in ten patients with septic pulmonary emboli. Spiral CT scans (10-mm collimation) were obtained at 10-mm intervals from the lung apex to the diaphragm and were compared with posteroanterior chest radiographs obtained within 24 h after CT scanning. RESULTS Only 21 (32%) of the 65 lesions detected on CT scans were also detected on chest radiographs. Peripheral nodules (39 lesions (60%)) were seen in all ten patients, wedge-shaped peripheral lesions (15 lesions (23%)) in nine patients, and infiltrates (11 lesions (17%)) in four patients. Subpleural lesions (45 lesions (69%)) and feeding vessels (35 (54%)) were found in all patients, and cavitary lesions (seven lesions (11%)) were seen in four patients. Subpleural peripheral nodules and wedge-shaped peripheral lesions were seen in nine patients. Thirty-two lesions (49%) ranged in diameter from 10 to 19 mm, and 59 lesions (91%) were less than 30 mm. CONCLUSIONS Spiral CT is useful in detecting septic pulmonary emboli. On spiral CT subpleural peripheral nodules and wedge-shaped peripheral lesions less than 30 mm in diameter are often found in patients with septic pulmonary emboli.

Intracellular Chloride Regulates Cell Proliferation Through the Activation of Stress-Activated Protein Kinases in MKN28 Human Gastric Cancer Cells

Recently, we reported that reduction of intracellular Cl− concentration ([Cl−]i) inhibited proliferation of MKN28 gastric cancer cells by diminishing the transition rate from G1 to S cell‐cycle phase through upregulation of p21, cyclin‐dependent kinase inhibitor, in a p53‐independent manner. However, it is still unknown how intracellular Cl− regulates p21 expression level. In this study, we demonstrate that mitogen‐activated protein kinases (MAPKs) are involved in the p21 upregulation and cell‐cycle arrest induced by reduction of [Cl−]i. Culture of MKN28 cells in a low Cl− medium significantly induced phosphorylation (activation) of MAPKs (ERK, p38, and JNK) and G1/S cell‐cycle arrest. To clarify the involvement of MAPKs in p21 upregulation and cell growth inhibition in the low Cl− medium, we studied effects of specific MAPKs inhibitors on p21 upregulation and G1/S cell‐cycle arrest in MKN28 cells. Treatment with an inhibitor of p38 or JNK significantly suppressed p21 upregulation caused by culture in a low Cl− medium and rescued MKN28 cells from the low Cl−‐induced G1 cell‐cycle arrest, whereas treatment with an ERK inhibitor had no significant effect on p21 expression or the growth of MKN28 cells in the low Cl− medium. These results strongly suggest that the intracellular Cl− affects the cell proliferation via activation of p38 and/or JNK cascades through upregulation of the cyclin‐dependent kinase inhibitor (p21) in a p53‐independent manner in MKN28 cells. J. Cell. Physiol. 223:764–770, 2010.

Chloride ion modulates cell proliferation of human androgen-independent prostatic cancer cell.

In the present study, we investigated if the intracellular Cl^- affects cell growth and cell cycle progression of androgen-independent prostate cancer PC3 cells. PC3 cells cultured in a medium containing 113 mM Cl^- for 96 h grew up 9-fold in cell number, while PC3 cells cultured in an 8 mM-Cl^--containing culture medium showed complete arrest of cell growth even after culture for 96 h. Exposure of cells to the 8 mM-Cl^- culture medium diminished phosphorylation levels of Rb and cdc2, which are respectively key accelerators of transition from G₁ to S phase and G₂ to M phase in cell cycle progression. Culturing cells in the 8 mM-Cl^--containing culture medium upregulated the protein expression level of p21 (a CDK inhibitor) inhibiting transition of G₁ to S phase, and diminished the incorporation of 5-ethynyl-2′-deoxyuridine (EdU; a thymidine analogue) into DNA. These results suggest that cells cultured in the low Cl^- medium prolonged the duration of all phases of the cell cycle (G₁, S, and G₂/M), thereby abolishing overall cell cycle progression. Effects of culturing cells in the low Cl^- culture medium on cell cycle progression would be mediated via a change in the intracellular Cl^- concentration ([Cl^-]_i), since [Cl^-]_i was decreased under a low Cl^- culture medium. To clarify this possibility, we studied effects of furosemide and bumetanide, Na⁺/K⁺/2Cl^- cotransporter (NKCC) inhibitors, on proliferation of PC3 cells. Furosemide and bumetanide decreased [Cl^-]_i and cell growth of PC3 cells. These results suggest that a change in [Cl^-]_i would play a critical role in this growth mechanism.

ROLE OF INTERCELLULAR ADHESION MOLECULE 1 IN ACUTE LUNG INJURY INDUCED BY CANDIDEMIA

Candidemia, a complication often affecting immunocompromised patients, is a common cause of acute lung injury. Yeast-phase Candida albicans has been shown to express a protein that is antigenically and structurally related to Mac-1. C. albicans is reported to stimulate intercellular adhesion molecule 1 (ICAM-1)expression on endothelial cells. In this study, the authors examined the role of ICAM-1 in acute lung injury induced by candidemia. The authors cultured rat pulmonary artery endothelial cells (RPAEC)and investigated the effect of anti-ICAM-1antibodies on adhesion of C. albicans to RPAEC. In addition, the authors administered anti-ICAM-1 antibodies to rats to examine the effect of the antibodies on experimentally induced candidemia. Survival rates, lung wet-to-dry (W/D) weight ratios, bronchoalveolar lavage (BAL) fluid, histopathological findings, and colony-forming units (CFUs)of lung C. albicans were examined. The adherence of C. albicans to RPAEC was significantly decreased by anti-ICAM-1 antibodies. Anti-ICAM-1 antibodies significantly increased survival, decreased lung W/D weight ratios, decreased neutrophil counts in the BAL fluid, reduced microscopic lung injury, and decreased the quantity of lung C. albicans. These results indicate that ICAM-1plays a role in adherence of C. albicans to pulmonary vascular endothelial cells, which likely leads to invasion of lung tissue by the organism.Candidemia, a complication often affecting immunocompromised patients, is a common cause of acute lung injury. Yeast-phase Candida albicans has been shown to express a protein that is antigenically and structurally related to Mac-1. C. albicans is reported to stimulate intercellular adhesion molecule 1 (ICAM-1) expression on endothelial cells. In this study, the authors examined the role of ICAM-1 in acute lung injury induced by candidemia. The authors cultured rat pulmonary artery endothelial cells (RPAEC) and investigated the effect of anti-ICAM-1 antibodies on adhesion of C. albicans to RPAEC. In addition, the authors administered anti-ICAM-1 antibodies to rats to examine the effect of the antibodies on experimentally induced candidemia. Survival rates, lung wet-to-dry (W/D) weight ratios, bronchoalveolar lavage (BAL) fluid, histopathological findings, and colony-forming units (CFUs) of lung C. albicans were examined. The adherence of C. albicans to RPAEC was significantly decreased by anti-ICAM-1 antibodies. Anti-ICAM-1 antibodies significantly increased survival, decreased lung W/D weight ratios, decreased neutrophil counts in the BAL fluid, reduced microscopic lung injury, and decreased the quantity of lung C. albicans. These results indicate that ICAM-1 plays a role in adherence of C. albicans to pulmonary vascular endothelial cells, which likely leads to invasion of lung tissue by the organism.

Regulation of Epithelial Sodium Transport via Epithelial Na+ Channel

Renal epithelial Na+ transport plays an important role in homeostasis of our body fluid content and blood pressure. Further, the Na+ transport in alveolar epithelial cells essentially controls the amount of alveolar fluid that should be kept at an appropriate level for normal gas exchange. The epithelial Na+ transport is generally mediated through two steps: (1) the entry step of Na+ via epithelial Na+ channel (ENaC) at the apical membrane and (2) the extrusion step of Na+ via the Na+, K+-ATPase at the basolateral membrane. In general, the Na+ entry via ENaC is the rate-limiting step. Therefore, the regulation of ENaC plays an essential role in control of blood pressure and normal gas exchange. In this paper, we discuss two major factors in ENaC regulation: (1) activity of individual ENaC and (2) number of ENaC located at the apical membrane.

Role of Alveolar Macrophages in Candida-Induced Acute Lung Injury

ABSTRACT Recent studies have shown that alveolar macrophages (AMs) not only act as phagocytes but also play a central role as potent secretory cells in various lung diseases, including pneumonia and acute respiratory distress syndrome. The behavior of AMs during disseminated candidiasis, however, is insufficiently elucidated. This study is the first to report disseminated candidiasis in AM-depleted mice and to analyze the effect of AMs on Candida-induced acute lung injury. While all AM-sufficient mice died by day 2 after infection withCandida albicans, no mortality was observed among AM-depleted mice. Unexpectedly, the CFU numbers of C. albicans isolated from the lungs of AM-depleted mice were significantly higher than those for C. albicans isolated from AM-sufficient mice. The lung wet-to-dry weight ratio was lower for AM-depleted mice than for AM-sufficient mice, although this difference was not significant. We found that bronchoalveolar lavage fluid (BALF) from AM-depleted mice in candidemia contained fewer neutrophils than BALF from AM-sufficient mice. In addition, myeloperoxidase activities in lung homogenates of AM-depleted mice were significantly lower than those in homogenates of AM-sufficient mice. A significant decrease in levels of murine macrophage inflammatory protein 2 (MIP-2), a potent chemoattractant for neutrophils, was noted in lung homogenates from AM-depleted mice compared with levels in homogenates from AM-sufficient mice. Immunohistochemical studies using anti-MIP-2 antibodies revealed that AMs were the cellular source of MIP-2 within the lung during candidemia. We observed that AM depletion decreased levels of AM-derived neutrophil chemoattractant, alleviated acute lung injury during candidemia, and prolonged the survival of mice in candidemia, even though clearance of C. albicans from the lungs was reduced.

Quercetin Stimulates Na+/K+/2Cl− Cotransport via PTK-Dependent Mechanisms in Human Airway Epithelium

We investigated regulatory mechanisms of Cl(-) secretion playing an essential role in the maintenance of surface fluid in human airway epithelial Calu-3 cells. The present study reports that quercetin (a flavonoid) stimulated bumetanide-sensitive Cl(-) secretion with reduction of apical Cl(-) conductance, suggesting that quercetin stimulates Cl(-) secretion by activating an entry step of Cl(-) across the basolateral membrane through Na(+)/K(+)/2Cl(-) cotransporter (NKCC1). To clarify the mechanism stimulating NKCC1 by quercetin, we verified involvement of protein kinase (PK)A, PKC, protein tyrosine kinase (PTK), and cytosolic Ca(2+)-dependent pathways. A PKA inhibitor (PKI-14-22 amide), a PKC inhibitor (Gö 6983) or a Ca(2+) chelating agent did not affect the quercetin-stimulated Cl(-) secretion. On the other hand, a PTK inhibitor (AG18) significantly diminished the stimulatory action of quercetin on Cl(-) secretion without inhibitory effects on apical Cl(-) conductance, suggesting that a PTK-mediated pathway is involved in the stimulatory action of quercetin. The quercetin action on Cl(-) secretion was suppressed with brefeldin A (BFA, an inhibitor of vesicular transport from ER to Golgi), and the BFA-sensitive Cl(-) secretion was not observed in the presence of an epidermal growth factor receptor (EGFR) kinase inhibitor (AG1478), suggesting that quercetin stimulates Cl(-) secretion by causing the EGFR kinase-mediated translocation of NKCC1 or an NKC1-activating factor to the basolateral membrane in human airway epithelial Calu-3 cells. However, the surface density of NKCC1 was not increased by quercetin, but quercetin elevated the activity of NKCC1. These observations indicate that quercetin stimulates Cl(-) secretion by activating NKCC1 via translocation of an NKCC1-activating factor through an EGFR kinase-dependent pathway.

Action of N-acylated ambroxol derivatives on secretion of chloride ions in human airway epithelia.

We report the effects of new N-acylated ambroxol derivatives (TEI-588a, TEI-588b, TEI-589a, TEI-589b, TEI-602a and TEI-602b: a, aromatic amine-acylated derivative; b, aliphatic amine-acylated derivative) induced from ambroxol (a mucolytic agent to treat human lung diseases) on Cl(-) secretion in human submucosal serous Calu-3 cells under a Na(+)/K(+)/2Cl(-) cotransporter-1 (NKCC1)-mediated hyper-secreting condition. TEI-589a, TEI-589b and TEI-602a diminished hyper-secretion of Cl(-) by diminishing the activity of NKCC1 without blockade of apical Cl(-) channel (TEI-589a>TEI-602a>TEI-589b), while any other tested compounds including ambroxol had no effects on Cl(-) secretion. These indicate that the inhibitory action of an aromatic amine-acylated derivative on Cl(-) secretion is stronger that that of an aliphatic amine-acylated derivative, and that 3-(2,5-dimethyl)furoyl group has a strong action in inhibition of Cl(-) secretion than cyclopropanoyl group. We here indicate that TEI-589a, TEI-589b and TEI-602a reduce hyper-secretion to an appropriate level in the airway, providing a possibility that the compound can be an effective drug in airway obstructive diseases including COPD by reducing the airway resistance under a hyper-secreting condition.

Procaterol-stimulated Increases in Ciliary Bend Amplitude and Ciliary Beat Frequency in Mouse Bronchioles

The beating cilia play a key role in lung mucociliary transport. The ciliary beating frequency (CBF) and ciliary bend amplitude (CBA) of isolated mouse bronchiolar ciliary cells were measured using a light microscope equipped with a high-speed camera (500 Hz). Procaterol (aβ 2-agonist) increased CBA and CBF in a dose dependent manner via cAMP. The time course of CBA increase is distinct from that of CBF increase: procaterol at 10 nM first increased CBA and then CBF. Moreover, 10 pM procaterol increased CBA, not CBF, whereas 10 nM procaterol increased both CBA and CBF. Concentration-response studies of procaterol demonstrated that the CBA curve was shifted to a lower concentration than the CBF curve, which suggests that CBA regulation is different from CBF regulation. Measurements of microbead movements on the bronchiole of lung slices revealed that 10 pM procaterol increased the rate of ciliary transport by 37% and 10 nM procaterol increased it by 70%. In conclusion, we have shown that increased CBA is of particular importance for increasing the bronchiolar ciliary transport rate, although CBF also plays a role in increasing it.

INFLUENCE OF DEPLETION OF ALVEOLAR MACROPHAGES ON APOPTOSIS IN CANDIDA-INDUCED ACUTE LUNG INJURY

Apoptosis plays an important role in acute lung injury (ALI), and alveolar macrophages (AMs) are known to secrete proinflammatory cytokines and promote alveolar inflammation. The authors have previously reported that AMs can be depleted by inhalation of 1 mM 2-chloroadenosine (2-CA). In this study, the authors evaluated the effect of AM depletion by 2-CA inhalation on apoptosis in Candida-induced ALI. The results of in situ terminal deoxynucleotidyl transferase–mediated dUTP biotin nick end-labeling (TUNEL) and immunohistochemical studies and measurement of cytokine levels and caspase 3 activities in lung homogenates indicated that the Fas-FasL system and apoptosis of alveolar epithelial cells are suppressed by depletion of AMs by 2-CA inhalation.