Mika Terasaki

Hydrogen therapy attenuates irradiation-induced lung damage by reducing oxidative stress

Molecular hydrogen (H(2)) is an efficient antioxidant that diffuses rapidly across cell membranes, reduces reactive oxygen species (ROS), such as hydroxyl radicals and peroxynitrite, and suppresses oxidative stress-induced injury in several organs. ROS have been implicated in radiation-induced damage to lungs. Because prompt elimination of irradiation-induced ROS should protect lung tissue from damaging effects of irradiation, we investigated the possibility that H(2) could serve as a radioprotector in the lung. Cells of the human lung epithelial cell line A549 received 10 Gy irradiation with or without H(2) treatment via H(2)-rich PBS or medium. We studied the possible radioprotective effects of H(2) by analyzing ROS and cell damage. Also, C57BL/6J female mice received 15 Gy irradiation to the thorax. Treatment groups inhaled 3% H(2) gas and drank H(2)-enriched water. We evaluated acute and late-irradiation lung damage after H(2) treatment. H(2) reduced the amount of irradiation-induced ROS in A549 cells, as shown by electron spin resonance and fluorescent indicator signals. H(2) also reduced cell damage, measured as levels of oxidative stress and apoptotic markers, and improved cell viability. Within 1 wk after whole thorax irradiation, immunohistochemistry and immunoblotting showed that H(2) treatment reduced oxidative stress and apoptosis, measures of acute damage, in the lungs of mice. At 5 mo after irradiation, chest computed tomography, Ashcroft scores, and type III collagen deposition demonstrated that H(2) treatment reduced lung fibrosis (late damage). This study thus demonstrated that H(2) treatment is valuable for protection against irradiation lung damage with no known toxicity.

Role of survivin in acute lung injury: epithelial cells of mice and humans

Survivin, an inhibitor of apoptosis, regulates cell division and is a potential target for anticancer drugs because many cancers express high survivin levels. However, whether survivin would be toxic to human lung cells and tissues has not been determined. This report clarified the involvement of survivin in acute lung injury. We used immunohistochemical analysis, immunoelectron microscopy, and real-time reverse transcription-quantitative polymerase chain reaction to study survivin expression and localization in injured mouse and human lungs. We also used cultured human lung epithelial cells (BEAS-2B and A549) to study survivin cytoprotection. Nuclei and cytoplasm of epithelial cells in day 3 and day 7 models of bleomycin-injured lung showed survivin-positive results, which is consistent with upregulated survivin mRNA expression. These nuclei also evidenced double positive findings for proliferating cell nuclear antigen and survivin. Day 7 models had similar Smac/DIABLO-positive and survivin-positive cell distributions. The cytoplasm and nuclei of epithelial cells in lesions with diffuse alveolar damage manifested strong survivin-positive findings. Bleomycin stimulation in both epithelial cell lines upregulated expression of survivin and apoptosis-related molecules. Suppression of survivin expression with small interfering RNA rendered human lung epithelial cells susceptible to bleomycin-induced damage, with markedly upregulated activation of caspase-3, caspase-7, poly (ADP-ribose) polymerase, and lactate dehydrogenase activity and an increased number of dead cells compared with mock small interfering RNA-treated cells. Overexpression of survivin via transfection resulted in these epithelial cells being resistant to bleomycin-induced cell damage, with reduced activation of apoptosis-related molecules and lactate dehydrogenase activity and fewer dead cells compared with results for mock-transfected cells. Survivin, acting at the epithelial cell level that depends partly on apoptosis inhibition, is therefore a key mediator of cytoprotection in acute lung injury. Understanding the precise role of survivin in normal lung cells is required for the development of therapeutic survivin.

Snail expression and outcome in T1 high-grade and T2 bladder cancer: a retrospective immunohistochemical analysis

BackgroundNeoadjuvant chemotherapy has been shown to have benefit in T1 high-grade or T2 bladder cancer. However, neoadjuvant chemotherapy fails in some patients. Careful patient selection for neoadjuvant chemotherapy is therefore needed. Several reports show that Snail is associated with resistance to chemotherapy. We hypothesized that Snail expression could predict survival in T1 high-grade and T2 bladder cancer patients treated with neoadjuvant chemotherapy.MethodsThe participants were 44 patients with T1 high-grade and T2 bladder cancer receiving neoadjuvant chemotherapy. Immunohistochemical analysis was used to determine Snail expression in specimens of bladder cancer obtained by transurethral resection before neoadjuvant chemotherapy. The relationships between Snail expression and patients’ outcomes were analyzed.ResultsSnail expression was positive in 15 of the 44 patients (34.1%) and negative in 29 (65.9%). Disease-free survival was significantly shorter for the Snail-positive group than for the Snail-negative group (p = 0.014). In addition, disease-specific survival was also significantly shorter for the Snail-positive group than for the Snail-negative group (p = 0.039). In multivariate analysis, Snail expression level was identified as an independent prognostic factor for disease-specific survival (p = 0.020).ConclusionsThe results indicate that Snail expression may predict poor outcome in T1 high-grade and T2 bladder cancer patients treated with neoadjuvant chemotherapy.

The difference of neovascularization in early intra-alveolar fibrosis between nonspecific interstitial pneumonia and usual interstitial pneumonia

Of the idiopathic interstitial pneumonias (IIPs), usual interstitial pneumonia (UIP) and diffuse alveolar damage (DAD) usually have poor prognoses. The prognoses of cryptogenic organizing pneumonia (COP) and nonspecific interstitial pneumonia (NSIP) are usually more favorable. Although several reports have described neovascularization in COP and UIP, this aspect of UIP has not been compared with NSIP. In this study, we evaluated neovascularization in intra‐alveolar fibrotic lesion of cases of fibrosing NSIP (f‐NSIP) (n = 26) and UIP (n = 25). In the f‐NSIP group, a considerable degree of neovascularization was observed compared to the UIP group and bud type intra‐alveolar fibrosis showed a greater degree of neovascularization compared to the mural‐incorporation and obliterative types of intra‐alveolar fibrosis. Real‐time reverse transcription polymerase chain reaction revealed a significantly greater expression of VEGF‐A mRNA in f‐NSIP than in UIP. The expression of matrix metalloproteinase‐2 (MMP‐2) mRNA also showed significantly higher in f‐NSIP than UIP. The greater VEGF‐A and MMP‐2 expression may play a role in the pathogenesis of neovascularization in early intra‐alveolar fibrotic lesions in f‐NSIP.

A mucin-rich variant of salivary duct carcinoma with a prominent mucinous component, a tumor that mimics mucinous adenocarcinoma

The mucin-rich variant of salivary duct carcinoma (mSDC) is a rare type of salivary duct carcinoma. mSDC usually has both conventional SDC and mucinous adenocarcinoma-like areas. This article describes a first case of mSDC in which 95% of the tumor consisted of a mucinous area without no solid conventional SDC, so that the tumor mimicked mucinous adenocarcinoma. A 55-year-old man was evaluated for a 14 mm mass in the left submandibular gland. The tumor showed that floating tumor nests in a prominent mucinous lake. Some floating tumor nests had focal cribriform pattern with comedo necrosis, and all tumor cells had immunoreactivity for androgen receptor, gross cystic disease fluid protein 15, and Her-2/neu. A diagnosis of mSDC was rendered. mSDC with prominent mucinous component sometimes resembles mucinous adenocarcinoma. Identifying specific histological and immunohistochemical features of floating tumor nests in the mucinous area are important for the diagnosis.

Vibrio cholerae Cholix Toxin-Induced HepG2 Cell Death is Enhanced by Tumor Necrosis Factor-Alpha Through ROS and Intracellular Signal-Regulated Kinases

Cholix toxin (Cholix) from Vibrio cholerae is a potent virulence factor exhibiting ADP-ribosyltransferase activity on eukaryotic elongation factor 2 (eEF2) of host cells, resulting in the inhibition of protein synthesis. Administration of Cholix or its homologue Pseudomonas exotoxin A (PEA) to mice causes lethal hepatocyte damage. In this study, we demonstrate cytotoxicity of Cholix on human hepatocytes in the presence of tumor necrosis factor α (TNF-α), which has been reported to play a fatal role in PEA administered to mice. Compared with incubating HepG2 cells with Cholix alone, co-treatment with TNF-α and Cholix (TNF-α/Cholix) significantly enhanced the activation of caspases, cytochrome c release from mitochondria into cytoplasm, and poly-ADP-ribose polymerase (PARP) cleavage, while incubation with TNF-α alone or co-treatment with TNF-α/catalytically inactive Cholix did not. In the early stage of cell death, Cholix increased phosphorylation of mitogen-activated protein kinases (e.g., p38, ERK, JNK) and Akt, which was not affected by TNF-α alone. MAPK inhibitors (SP600125, SB20852, and U0126) suppressed PARP cleavage induced by TNF-α/Cholix. Protein kinase inhibitor Go6976 suppressed JNK phosphorylation and PARP cleavage by TNF-α/Cholix. In contrast, PKC activator PMA in the absence of TNF-α promoted Cholix-induced PARP cleavage. Reactive oxygen species (ROS) inhibitor, N-acetyl cysteine (NAC), suppressed TNF-α/Cholix-induced JNK and ERK phosphorylation, resulting in inhibition of PARP cleavage. These data suggest that ROS and JNK pathways are important mediators of TNF-α/Cholix-induced HepG2 cell death.

Role of α1 and α2 chains of type IV collagen in early fibrotic lesions of idiopathic interstitial pneumonias and migration of lung fibroblasts.

Early fibrotic lesions are thought to be the initial findings of fibrogenesis in idiopathic interstitial pneumonias, but little is known about their properties. Type IV collagen comprises six gene products, α1–α6, and although it is known as a major basement membrane component, its abnormal deposition is seen in fibrotic lesions of certain organs. We studied the expression of type I and III collagen and all α chains of type IV collagen in lung specimens from patients with usual interstitial pneumonia (UIP) or organizing pneumonia (OP) via immunohistochemistry. With cultured lung fibroblasts, we analyzed the expression and function of all α chains of type IV collagen via immunohistochemistry, western blotting, real-time quantitative PCR, and a Boyden chamber migration assay after the knockdown of α1 and α2 chains. Although we observed type I and III collagens in early fibrotic lesions of both UIP and OP, we found type IV collagen, especially α1 and α2 chains, in early fibrotic lesions of UIP but not OP. Fibroblasts enhanced the expression of α1 and α2 chains of type IV collagen after transforming growth factor-β1 stimulation. Small interfering RNA against α1 and α2 chains increased fibroblast migration, with upregulated phosphorylation of focal adhesion kinase (FAK), and adding medium containing fibroblast-produced α1 and α2 chains reduced the increased levels of fibroblast migration and phosphorylation of FAK. Fibroblasts in OP were positive for phosphorylated FAK but fibroblasts in UIP were not. These results suggest that fibroblasts in UIP with type IV collagen deposition, especially α1 and α2 chains, have less ability to migrate from early fibrotic lesions than fibroblasts in OP without type IV collagen deposition. Thus, type IV collagen deposition in early fibrotic lesions of UIP may be implicated in refractory pathophysiology including migration of lesion fibroblasts via a FAK pathway.

Renal Inflammatory Changes in Acute Hepatic Failure-Associated Acute Kidney Injury

Background/Aims: Acute kidney injury (AKI) is a common complication in advanced liver dysfunction. Our aim is to clarify the mechanisms of acute hepatic failure (AHF)-associated AKI. Methods: We examined the mechanisms of AHF-associated AKI, which is characterized by AKI in AHF and hyperbilirubinemia, following DA-to-Lewis rat liver transplantation. Results: During the progression of AHF and hyperbilirubinemia in liver graft rejection, AHF-associated AKI gradually developed by day 11. Degeneration and apoptotic cells were apparent in tubular epithelial cells with bile pigment accumulation and mitochondrial degeneration. Injury of peritubular capillaries (PTCs) was also noted with apoptotic endothelial cells, decreased expression of endothelial nitric oxide synthase, accumulation of α-smooth muscle actin+ pericytes and/or myofibroblasts, and inflammation. Angiogenic factors including vascular endothelial growth factor, angiopoietin-1, and angiopoietin-2 in the cortex were decreased on day 11. In addition, a marked reduction in the velocity of red blood cells in PTCs was evident in vivo. Conclusions: AHF-associated AKI seems to be mediated by renal tubular epithelial cell injury with bile pigment accumulation, impaired microcirculation caused by PTC endothelial cell injury with depletion of endothelial nitric oxide synthase and angiogenic factors, and by a decrease in RBC velocity and renal inflammation. Multiple mechanisms including tubular and PTC injuries and renal inflammation may be involved in the development of AHF-associated AKI.

Localized pulmonary crystal-storing histiocytosis complicating pulmonary mucosa-associated lymphoid tissue lymphoma presenting with multiple mass lesions

Crystal-storing histiocytosis (CSH) is an uncommon finding in lymphoplasmacytic disorders that presents histiocytes with abnormal intralysosomal accumulations of immunoglobulin light chains as crystals of unknown etiology. A 38-year-old woman with antiphospholipid syndrome had a surgical lung biopsy because of multiple lung mass lesions. In a right middle lobe lesion, lymphoplasmacytic cells had a monocytoid appearance, destructive lymphoepithelial lesions, and positive immunoglobulin heavy chain (IGH) gene rearrangements. A right upper lobe lesion manifested proliferating rounded histiocytes with abundant, deeply eosinophilic cytoplasm and negative IGH gene rearrangements. Electron microscopy and mass spectrometry revealed a case of pulmonary CSH: abnormal proliferation of the immunoglobulin κ chain of a variable region that may be crystallized within plasma cells and histiocytes. We report a rare case of localized pulmonary CSH complicating pulmonary mucosa-associated lymphoid tissue lymphoma with multiple mass lesions. We demonstrate advances in the understanding of the pathogenesis of CSH by various analyses of these lesions.

Extranodal Natural Killer/T-Cell Lymphoma, Nasal Type, with Primary Manifestation as an Upper Eyelid Swelling.

Extranodal natural killer/T-cell lymphoma (ENK/TCL) is most often in the nose or the nasopharynx but can present elsewhere. We report a rare case of ENK/TCL that presented as swelling of an upper eyelid without ocular involvement. A 76-year-old man visited our hospital with a swollen lesion of the left upper eyelid which had appeared 2 months earlier. A biopsy of the upper eyelid revealed slight perivascular and periadnexal infiltration of mononuclear cells with dermal edema. Treatment with oral prednisolone at a dosage of 20 mg/day decreased the eyelid swelling. However, 5 months later, exacerbation of the swelling and nasal congestion were observed. A second biopsy of the upper eyelid revealed a diffuse dermal infiltrate composed of mononuclear cells with an angiocentic growth pattern. Immunohistochemical studies and in situ hybridization showed natural killer-lineage antigens (CD56, granzyme B, and T-cell intracellular antigen 1) with expression of Epstein-Barr virus. These findings lead to the diagnosis of ENK/TCL. We treated the patient with radiation therapy (50 Gy) and 3 courses of a regimen including dexamethasone, carboplatin, etoposide, and ifosphamide. This case suggests that ENK/TCL can present with swelling of an upper eyelid as the primary sign of the skin lesion. Swelling of an upper eyelid should be considered in the differential diagnosis of ENK/TCL.