Thermal C-O coupling reactions of Ta methylene clusters [TanCH2]+ (n = 1, 4) with O2.

Cationic tantalum carbenes [TaCH2]+ and [Ta4CH2]+, products of thermal methane dehydrogenation, are collected and stored in a ring-electrode ion trap. In there, potential C-O coupling reactions are probed by exposing the tantalum carbenes to dioxygen in the presence of helium buffer gas, thereby facilitating reactions under-well defined multi-collision conditions at 300 K. Time-of-flight mass spectrometry determines the time-dependent reaction products and a kinetic analysis illustrates significantly different selectivities. The cationic tantalum atom and tetramer systems undergo coupling reactions, which may lead to the formation of value-added products from the oxidation of methane. In particular, the reaction of [Ta4CH2]+ with oxygen demonstrates an increased selectivity towards the formation of a formaldehyde equivalent.

A prospective phase II study of combined androgen blockade in patients with androgen receptor-positive metastatic or locally advanced unresectable salivary gland carcinoma.

Background: There is no standard first-line chemotherapy for recurrent/metastatic (RM) or unresectable locally advanced (LA) salivary gland carcinoma (SGC). Patients and methods: We conducted a single institution, open-label, single arm, phase II trial of combined androgen blockade (CAB) for androgen receptor (AR)-positive SGC. Leuprorelin acetate was administered subcutaneously at a dose of 3.75 mg every 4 weeks. Bicalutamide was administered orally at a daily dose of 80 mg. Patients were treated until progressive disease or unacceptable toxicities. Results: Thirty-six eligible patients were enrolled. Thirty-three patients had RM disease and three patients had LA disease. The pathological diagnoses were salivary duct carcinoma (34 patients, 94%) and adenocarcinoma, NOS (two patients, 6%). The best overall response rate was 41.7% [n = 15, 95% confidence interval (CI), 25.5%-59.2%], the clinical benefit rate was 75.0% (n = 27, 95% CI, 57.8%-87.9%). The median progression-free survival was 8.8 months (95% CI, 6.3-12.3 months) and the median overall survival was 30.5 months (95% CI, 16.8 months to not reached). Additional analyses between treatment outcomes and clinicopathological factors or biomarkers including AR positivity, human epidermal growth factor receptor 2 status, and its complex downstream signaling pathway gene mutations showed no statistically significant differences. Elevated grade 3 liver transaminases and increased serum creatinine were reported in two patients, respectively. Discontinuation of leuprorelin acetate or bicalutamide due to adverse event occurred in one patient. Conclusion: This study suggests that CAB has equivalent efficacy and less toxicity for patients with AR-positive RM or unresectable LA SGC compared with conventional chemotherapy, which warrants further study. Clinical Trial Registration: UMIN-CTR (http://www.umin.ac.jp/ctr/index-j.htm), identification number: UMIN000005703.

In Situ Assembly of Transmembrane Proteins from Expressed and Synthetic Components in Giant Unilamellar Vesicles.

Reconstituting functional transmembrane (TM) proteins into model membranes is challenging due to the difficulty of expressing hydrophobic TM domains, which often require stabilizing detergents that can perturb protein structure and function. Recent model systems solve this problem by linking the soluble domains of membrane proteins to lipids, using noncovalent conjugation. Herein, we test an alternative solution involving the in vitro assembly of TM proteins from synthetic TM domains and expressed soluble domains using chemoselective peptide ligation. We developed an intein mediated ligation strategy to semisynthesize single-pass TM proteins in synthetic giant unilamellar vesicle (GUV) membranes by covalently attaching soluble protein domains to a synthetic TM polypeptide, avoiding the requirement for detergent. We show that the extracellular domain of programmed cell death protein 1, a mammalian immune checkpoint receptor, retains its ligand-binding function at a membrane interface after ligation to a synthetic TM peptide in GUVs, facilitating the study of receptor-ligand interactions.

Intra-arterial chemoradiation therapy with weekly low-dose cisplatin for squamous cell carcinoma of the maxillary sinus.

A new intra-arterial chemoradiation regimen that involves infusing low-dose cisplatin in combination with definitive irradiation was used in 36 patients diagnosed with squamous cell carcinoma of the maxillary sinus. The safety and therapeutic efficacy of this regimen were reviewed retrospectively. An intra-arterial catheter was inserted in a retrograde manner into the target artery via the superficial temporal artery or occipital artery. Intra-arterial infusion was performed using cisplatin at a dose of 20-50mg/m(2) per week for 6-8 weeks. At the same time, sodium thiosulphate was infused as a neutralizing agent. Irradiation was performed at 60Gy in 30 fractions. All 36 patients completed treatment. Grade 3 adverse events occurred in only seven patients (19.4%) and no grade 4 events were noted. As a primary therapy, the complete response rate was 83.3%, the partial response rate was 16.7%, and the overall response rate was 100%. The 2-year local control rate was 63.0%, and the 2-year overall survival rate was 75.5%. The 2-year preservation rate of the hard palate was 97.1%, that of the eyeball was 97.2%, and that of visual function was 94.4%. This treatment regimen can contribute to improving the quality of life of patients without reducing the curability of the therapy.

Nanometer-nanosecond oscillatory expansion and contraction behavior of polymer films induced by 248 nm excimer laser excitation.

Microregional mechanical measurements of a poly[methylmethacrylate) film can be made by a combined probe/measurement laser system, as represented in the picture. The localized nature of the laser systems allow quantification of the degree of expansion and contraction processes, as well as the glass-rubber transition. Such nondestructive techniques lead to the elucidation of polymer morphological dynamics.

[ADOC regimen for unresectable advanced thymic cancer].

Between 1996 and 1998, we treated 6 patients with unresectable and advanced thymic cancer (stages IVa and IVb). All received 50 mg/m2 of cisplatin and 40 mg/m2 of doxorubicin intravenously (i.v.) on day 1,0.6 mg/m2 of vincristine i.v. on day 3, and 700 mg/m2 of cyclophosphamide i.v. on day 4; ADOC regimen, respectively at 3-4 week intervals. Four patients obtained a partial response (PR) after ADOC chemotherapy and the overall clinical response rate was 67%. No life-threatening side effects were noted. In 2 patients, cisplatin plus VP-16 chemotherapy failed to demonstrate any benefits prior to the ADOC regimen. Radiotherapy was initiated after the achievement of PR in the other 2 patients. ADOC chemotherapy appears to be an effective treatment for thymic cancer.

Cyclophosphamide stimulates lung fibroblasts to release neutrophil and monocyte chemoattractants.

Cyclophosphamide is an alkylating antineoplastic agent used in several conditions. However, little is known about the mechanism of its pulmonary toxicity. In the present study, we determined that human lung fibroblasts release activity for neutrophils and monocytes in response to cyclophosphamide in a dose- and time-dependent manner. Checkerboard analysis revealed that both neutrophil and monocyte activities were chemotactic. The release of chemotactic activity was inhibited by lipoxygenase inhibitors and cycloheximide. Molecular-sieve column chromatography revealed that both neutrophil (NCA) and monocyte (MCA) chemotactic activities had multiple peaks. NCA was inhibited by a leukotriene B(4) receptor antagonist and anti-interleukin-8 and anti-granulocyte colony-stimulating factor antibodies. MCA was attenuated by a leukotriene B(4) receptor antagonist and anti-monocyte chemoattractant protein-1 and anti-granulocyte-macrophage colony-stimulating factor antibodies. The concentrations of interleukin-8, granulocyte colony-stimulating factor, monocyte chemoattractant protein-1, and granulocyte-macrophage colony-stimulating factor significantly increased in response to cyclophosphamide. These data suggest that lung fibroblasts may modulate inflammatory cell recruitment into the lung by releasing NCA and MCA in response to cyclophosphamide.

Bleomycin stimulates lung epithelial cells to release neutrophil and monocyte chemotactic activities.

Although bleomycin, an antineoplastic drug, is used in the treatment of a variety of tumors, the mechanisms of bleomycin-induced lung injury and fibrosis are not fully elucidated. We postulated that bleomycin might stimulate A549 cells, a type II pneumocyte cell line, to release neutrophil and monocyte chemotactic activities (NCA and MCA, respectively). To test this hypothesis, A549 cell supernatant fluids were harvested and evaluated for NCA and MCA. A549 cell supernatant fluids showed NCA and MCA in response to bleomycin in a dose- and time-dependent manner (P < 0.05). Checkerboard analysis revealed that both NCA and MCA were predominantly chemotactic. Partial characterization of the released NCA and MCA showed that the activities were partially heat labile, trypsin digested, and predominantly ethyl acetate extractable. Lipoxygenase inhibitors and cycloheximide inhibited the release of chemotactic activities significantly. Molecular-sieve column chromatography revealed that the released activities were heterogeneous. However, low-molecular-weight activity was prominent. Leukotriene B4-receptor antagonist, anti-interleukin-8, anti-granulocyte colony-stimulating factor, and anti-monocyte chemoattractant protein-1 antibodies attenuated the chemotactic activities. Immunoreactive leukotriene B4 receptor, interleukin-8, granulocyte colony-stimulating factor, and monocyte chemoattractant protein-1 significantly increased in supernatant fluids in response to bleomycin. These data demonstrate that bleomycin stimulates type II epithelial cells to release chemotactic activities and plays a role in inflammatory cell recruitment into the lung.

Alveolar type II-like cells release G-CSF as neutrophil chemotactic activity.

We evaluated the potential of A549 cells, an alveolar type II epithelial cell line, to release granulocyte colony-stimulating factor (G-CSF), in addition to interleukin (IL)-8 and leukotriene B4, as neutrophil chemotactic activity (NCA). Human recombinant IL-1beta stimulated A549 cells to release NCA in a time- and dose-dependent fashion. The released NCA was blocked by mouse anti-human G-CSF polyclonal antibody. Molecular-sieve column chromatography revealed that IL-1beta induced the release of a 19- to 20-kDa chemotactic mass that was inhibited by anti-human G-CSF antibody. IL-1beta stimulated the release of G-CSF in a dose-dependent fashion, but the time-dependent profile of G-CSF showed that the concentration of G-CSF declined after 48 h. Tumor necrosis factor (TNF)-alpha, Escherichia coli lipopolysaccharide (LPS), and bradykinin (BK) stimulated A549 cells to release NCA that was inhibited by anti-G-CSF antibody. The release of G-CSF in response to TNF-alpha, LPS, and BK was significantly increased. The similar concentrations of human recombinant G-CSF (10-1,000 pg/ml) as in the supernatant fluid induced neutrophil chemotaxis. G-CSF mRNA was expressed time and dose dependently at 4 h and declined after 4 h in response to IL-1beta as evaluated by RT-PCR. The expression of G-CSF mRNA was also observed by TNF-alpha, LPS, and BK stimulation. These data suggest that type II alveolar epithelial cells may produce G-CSF as NCA and may participate in the regulation of leukocyte extravasation.

Procaterol inhibits IL-1beta- and TNF-alpha-mediated epithelial cell eosinophil chemotactic activity.

Theophylline inhibits eosinophilic infiltration into the bronchial wall. It is unknown whether this is mediated by a cyclic adenosine monophosphate (c-AMP)-dependent reduction in eosinophil chemotactic activity (ECA) from bronchial epithelial cells (BEC). Therefore the effect of a beta2-agonist, procaterol and theophylline on the release of ECA from a BEC line, BEAS-2B was evaluated in response to interleukin (IL)-1beta and tumour necrosis factor-alpha (TNF-alpha). ECA was assessed using a blind-well chemotactic chamber, and the release and gene expression of cytokines were evaluated by means of enzyme-linked immunosorbent assay and reverse transcriptase polymerase chain reaction. IL-1beta and TNF-alpha stimulated the release of ECA from BEAS-2B cells in a dose- and time-dependent manner. Procaterol and theophylline directly inhibited eosinophil migration to IL-1beta and TNF-alpha-conditioned medium. The pretreatment of BEAS-2B cells with the same concentrations of procaterol inhibited the release of ECA in a dose-dependent fashion. Anti-IL-8, anti-regulated on activation, normal T-cell expressed and secreted (RANTES), and anti-granulocyte-macrophage colony-stimulating factor (GM-CSF) inhibited ECA. Procaterol inhibited the release of RANTES, GM-CSF and IL-8 in a dose-dependent fashion. The effect of theophylline was less potent. Procaterol augmented cAMP levels in BEAS-2B cells in a time- and dose-dependent manner. The expression of IL-8, RANTES, and GM-CSF messenger ribonucleic acid was not inhibited by procaterol and theophylline. These data indicate that procaterol and theophylline may directly inhibit eosinophil migration and that procaterol may further inhibit the release of eosinophil chemotactic activity from BEAS-2B cells via a cyclic adenosine monophosphate-dependent mechanism. This warrants further studies on the involvement of bronchial epithelial cells in the anti-inflammatory effects of procaterol and theophylline in patients with asthma.

Human lung fibroblasts release chemokinetic activity for monocytes constitutively.

We determined whether human lung fibroblasts (HLFs) might release mediators that are responsible for monocyte chemokinetic activity (MCA) constitutively. HLF supernatant fluids showed MCA in a time-dependent manner (P < 0.001). Checkerboard analysis of 24- and 72-h supernatant fluids showed that the activity was chemokinetic. Partial characterization of 24- and 72-h supernatant fluids revealed that the mediators released after 24 h were predominantly composed of lipid-soluble activity, and MCA was blocked by lipoxygenase inhibitors. The mediators released after 72 h were predominantly trypsin sensitive and blocked by cycloheximide. Molecular-sieve column chromatography identified four peaks of MCA. A polyclonal antibody to monocyte chemoattractant protein-1 (MCP-1) inhibited MCA by 20% after 24 h and by 40% after 72 h. Granulocyte-macrophage colony-stimulating factor (GM-CSF) and transforming growth factor-beta (TGF-beta) antibodies attenuated MCA released after 72 h by 30 and 10%, respectively. These antibodies inhibited corresponding molecular-weight peaks separated by molecular-sieve column. The concentrations of MCP-1, GM-CSF, and TGF-beta were 4,698 +/- 242, 26.8 +/- 3.8, and 550 +/- 15 pg/ml, respectively. A leukotriene B4 (LTB4)-receptor antagonist attenuated the total MCA and the lowest molecular weight peak of MCA. The concentrations of LTB4 were 153.4 +/- 12.4 (24 h) and 212 +/- 16.6 (72 h) pg/ml. These findings suggest that HLFs may modulate the recruitment of monocytes into the lung by releasing MCP-1, GM-CSF, TGF-beta, and LTB4 constitutively.

Smoke extract stimulates lung fibroblasts to release neutrophil and monocyte chemotactic activities.

Accumulation of monocytes and neutrophils and fibrous distortion of the airway are characteristics of airway disease secondary to smoking. The presence of inflammatory cells and fibrosis correlate, and, therefore, we postulated that lung fibroblasts might release chemotactic activity for neutrophils and monocytes in response to smoke extract. To test this hypothesis, human fetal lung (HFL1) fibroblasts were cultured, and the supernatant fluid was evaluated for neutrophil (NCA) and monocyte (MCA) chemotactic activities with a blind well chamber technique. HFL1 fibroblasts released chemotactic activity in response to smoke extract in a dose- and time-dependent manner (P < 0.05). Checkerboard analysis showed that the activity was predominantly chemotactic. Partial characterization of the released chemotactic activity revealed that the activity was partly heat labile, trypsin sensitive, and ethyl acetate extractable. Lipoxygenase inhibitors and cycloheximide inhibited the release of both NCA and MCA. Molecular-sieve chromatography revealed that NCA and MCA were heterogeneous. NCA was inhibited by anti-human interleukin (IL)-8 and anti-granulocyte colony-stimulating factor antibodies and a leukotriene (LT) B(4)-receptor antagonist. Anti-granulocyte-macrophage colony-stimulating factor (GM-CSF) and anti-monocyte chemoattractant protein (MCP)-1 antibodies and an LTB(4)-receptor antagonist inhibited MCA. Immunoreactive IL-8, granulocyte colony-stimulating factor, GM-CSF, and MCP-1 significantly increased in culture supernatant fluid in response to smoke extract. Finally, smoke extract augmented the expression of mRNAs of IL-8, GM-CSF, and MCP-1. These data demonstrate that lung fibroblasts release NCA and MCA in response to smoke extract and suggest that lung fibroblasts may modulate the inflammatory cell recruitment into the lung.

Bleomycin stimulates lung fibroblasts to release neutrophil and monocyte chemotactic activity.

We determined whether human lung fibroblasts might release chemotactic activity for neutrophils (NCA) and monocytes (MCA) in response to bleomycin. The human lung fibroblasts supernatant fluids were evaluated for chemotactic activity by a blind well chamber technique. Human lung fibroblasts released NCA and MCA in a dose- and time-dependent manner in response to bleomycin. Checkerboard analysis of supernatant fluids revealed that both NCA and MCA were chemotactic. Partial characterization revealed that NCA was partly heat labile, trypsin sensitive, and predominantly ethyl acetate extractable. In contrast, MCA was partly trypsin sensitive and ethyl acetate extractable. The release of chemotactic activity was inhibited by lipoxygenase inhibitors and cycloheximide. Molecular sieve column chromatography revealed that both NCA and MCA had multiple chemotactic peaks. NCA was inhibited by leukotriene B4 receptor antagonist and anti-IL-8 and G-CSF Abs. MCA was attenuated by leukotriene B4 receptor antagonist, and monocyte chemoattractant protein-1, GM-CSF, and TGF-beta Abs. Leukotriene B4 receptor antagonist and these Abs inhibited the corresponding m.w. chemotactic activity separated by column chromatography. The concentrations of IL-8, G-CSF, monocyte chemoattractant protein-1, GM-CSF, and TGF-beta in the supernatant fluids significantly increased in response to bleomycin. These data suggest that lung fibroblasts may modulate inflammatory cell recruitment into the lung by releasing NCA and MCA in response to bleomycin.

Smoke extract stimulates lung epithelial cells to release neutrophil and monocyte chemotactic activity.

Inflammatory cells accumulate within the lungs of cigarette smokers. Current concepts suggest that these cells can induce protease-antiprotease and/or oxidant-antioxidant imbalance(s), which may damage the normal lung alveolar and interstitial structures. Because type II pneumocytes line the alveolar space, and because the inflammatory cells migrate and reside at the alveolus, we postulated that the type II pneumocytes might release chemotactic activity for neutrophils and monocytes in response to smoke extract. To test this hypothesis, A549 cells were cultured and the supernatant fluids were evaluated for the neutrophil and monocyte chemotactic activity (NCA and MCA) by a blind-well chamber technique. A549 cells released NCA and MCA in response to smoke extract in a dose- and time-dependent manner (P < 0.05). Checkerboard analysis showed that the activity was chemotactic. Partial characterization of NCA and MCA revealed that the activity was partly heat labile, trypsin sensitive, and ethyl acetate extractable. Lipoxygenase inhibitors and cycloheximide inhibited the release of NCA and MCA. Molecular sieve column chromatography showed multiple peaks for both NCA and MCA. NCA was inhibited by anti-human-interleukin (IL)-8 antibody, granulocyte colony-stimulating factor (G-CSF) antibody, or leukotriene (LT)B4 receptor antagonist. Monocyte chemoattractant protein (MCP)-1 antibody or LTB4 receptor antagonist inhibited MCA. Immunoreactive IL-8, G-CSF, MCP-1, and LTB4 significantly increased in the supernatant fluids in response to smoke extract. These data suggest that the type II pneumocytes may release NCA and MCA and modulate the inflammatory cell recruitment into the lung.

Pulmonary infection with Mycobacterium avium-intracellulare leads to air trapping distal to the small airways.

To clarify the structure and function of the airways in Mycobacterium avium-intracellulare (MAI) infection, we performed pulmonary function tests and high-resolution computed tomography (HRCT) of the thorax in female patients 61 +/- 9 yr of age (n = 12) with pulmonary MAI infection without predisposing lung disease and compared their data with those of normal female volunteers 54 +/- 8 yr of age (n = 9). We calculated the E/I ratio, i.e., the average ratio of HRCT number at full expiration to that at full inspiration, as an index for the evaluation of air trapping distal to the small airways. Patients showed significant increases in residual volume and slope of phase III (DeltaN2) of the single-breath nitrogen test, and significant decreases in flow at 50 and 25% of FVC, suggesting hyperinflation and obstruction of the small airways. HRCT of patients revealed the small nodules and ectasis of bronchioles and small bronchi located mainly in segments (S) S2, S3, S4, and S5. The E/I ratio was significantly elevated in patients, and especially higher in the upper lung field than in the lower lung field, suggesting air trapping distal to the small airways. The difference of E/I ratio between the upper and lower field is probably related to the segmental distribution of CT abnormalities. These findings suggest that MAI infection can lead to air trapping distal to the small airways.