Cytotoxic effects of psychoactive isobutyrylfentanyl and its halogenated derivatives on isolated rat hepatocytes.

The novel and numerous psychoactive compounds derived from the analgesic prescription drug N-phenyl-N-[1-(2-phenylethyl)piperidin-4-yl]propanamide (fentanyl) have been illegally abused as recreational drugs and caused numerous fatalities. Because some psychoactive/psychotropic drugs are known to be hepatotoxic in humans and experimental animals, the cytotoxic effects and mechanisms of 4-fluoroisobutyrylfentanyl (4F-iBF), 4-chloroisobutyrylfentanyl (4Cl-iBF), and the parent compound isobutyrylfentanyl (iBF) were studied in freshly isolated rat hepatocytes. 4F-iBF caused not only concentration (0-2.0 mM)- and time (0-3 h)-dependent cell death accompanied by the depletion of cellular ATP and reduced glutathione (GSH) and protein thiol levels but also the accumulation of oxidized glutathione. Of these fentanyls examined, 4Cl-iBF/4F-iBF-induced cytotoxicity with the loss of mitochondrial membrane potential at concentrations of 0.5 and 1.0 mM and the production of reactive oxygen species (ROS) at 0.5 mM were greater than those induced by iBF. The pretreatment of hepatocytes with N-acetyl-l-cysteine as a precursor of cellular GSH ameliorated, at least in part, cytotoxicity accompanied by insufficient ATP levels, the loss of mitochondrial membrane potential, and generation of ROS caused by 4Cl-iBF/4F-iBF, whereas pretreatment with diethyl maleate as a GSH depletor enhanced fentanyl-induced cytotoxicity accompanied by the rapid loss of cellular GSH. Taken collectively, these results indicate that the onset of cytotoxic effects caused by these fentanyls is partially attributable to cellular energy stress as well as oxidative stress.

Hepatic and renal toxicities and metabolism of fentanyl analogues in rats.

New synthetic opioids continue to emerge in the illicit market, and among them, fentanyl analogues pose a serious threat to the public health with their abuse and trafficking. We investigated the toxicity of fentanyl analogues on the liver and kidneys mediated by the µ-opioid receptor (MOR). Our study focused on 4-fluoro-isobutyrylfentanyl (4F-iBF), which is classified as a "narcotic" in Japan; structurally similar analogues 4-chloro-isobutyrylfentanyl (4Cl-iBF) and isobutyrylfentanyl (iBF) were also investigated. Rats that were intraperitoneally administered 4F-iBF (5 mg/kg (12.3 µmol/kg)) or iBF (12.3 µmol/kg) displayed hepatic and renal ischemic-like damage, but 4Cl-iBF (12.3 µmol/kg) did milder renal damage only. We found that the agonist activity of 4F-iBF, at MORs was approximately 7.2 times that of 4Cl-iBF, and that pretreatment with MOR antagonist naltrexone (0.8 mg/kg) alleviated liver and kidney injuries caused by 4F-iBF. These results suggested that 4F-iBF might cause ischemic damage to the liver and kidneys, induced by respiratory depression mediated by MORs. Furthermore, to elucidate the metabolism of fentanyl analogues, we investigated the change over time in the amount of 4F-iBF, 4Cl-iBF, iBF (6.15 µmol/kg, respectively), and their respective metabolites in serum after intraperitoneal administration to rats. The results showed that in 24-h post-dose serum, 4Cl-iBF and iBF were substantially eliminated while 4F-iBF remained at about 30% of the maximum level, and each of the N-dephenylethylated metabolites of 4F-iBF, 4Cl-iBF, and iBF was detected in 2-h post-dose serum. The results from this study revealed information on the hepatic and renal toxicities and metabolism related to fentanyl analogues.

Double-filter high-resolution soft x-ray tomographic diagnostic for investigating electron acceleration in TS-6 reconnection merging experiments.

An innovative tangential-view soft x-ray (SXR) tomographic imaging measurement was developed on the TS-6 spherical tokamak merging device as a key diagnostic for investigating the mechanism of electron acceleration. In order to measure SXR with different energy ranges, two micro-channel plates (MCPs) are, respectively, installed in two vacuum chambers, which are equipped with different filters. Especially designed lenses and fiber bundles serve as an optical system to transfer images from phosphor plates of MCPs to a high speed imaging system. This design also enables us to simultaneously measure two images appearing on phosphor plates of MCPs by just one high speed imaging system. The temporal and spatial resolution of this diagnostic can be up to 5 µs and 4 mm, respectively, at present. The tomographic method based on the Phillips-Tikhonov regularization is employed to reconstruct line-integrated images into the local emissivity of SXR, which reflects the spatial distribution of high-energy electrons. Owing to this diagnostic, we successfully measured SXR emitted from the downstream region of magnetic reconnection in TS-6 merging experiments for the first time. The energy range of SXR turned out to be higher than 100 eV but lower than 400 eV.

Structure elucidation of a PDE5 inhibitor detected as an illegal adulteration in a libido-boosting dietary supplement.

A compound with potent inhibitory activity for phosphodiesterase type 5 (PDE5) was identified as an illegal adulteration in a libido-boosting dietary supplement being sold at a store in Tokyo. This compound was identified as 5,6-diethyl-2-{5-[(4-methylpiperazin-1-yl)sulphonyl]-2-propoxyphenyl}pyrimidin-4(3H)-one using liquid chromatography-diode array detector (LC-DAD), liquid chromatography-tandem mass spectrometer (LC-MS), LC-HRMS, nuclear magnetic resonance (NMR), and X-ray crystallography. The IC50 value of the inhibitory activity for PDE5A1 (one of the PDE5 isoforms) was 2.0 nM (sildenafil IC50 value was 4.5 nM). This compound was previously synthesised as a PDE5 inhibitor by Shanghai Institute of Materia Medica. The dietary supplement contained 85 mg of this compound in a capsule, which was about 26% of the capsule content (320 mg).

Continuous organic synthesis in water around micro-orifices after flows.

Water flows through micro-orifices are important because they occur in various fields, such as biology, medical science, chemistry, and engineering. We have reported in previous work that organic matter was generated in micro-orifices after water flowed through the orifice, and we proposed that the organic matter was synthesized from nonorganic materials, including CO2 and N2 dissolved in water from air, and water via the action of hydroxyl radicals produced by the flow through the micro-orifice. In the present study, we examined whether organic materials are produced in the water outside of the orifices in addition to that in the orifice. We used the decrease in water volume to measure the organic synthesis because water should be consumed during the synthesis, and thus the decrease in water volume should reflect the organic synthesis. We let ultrapure water containing dissolved air flow through a micro-orifice as a pre-flow, we stopped the flow, and then we measured the volume of water enclosed in the mount in which the micro-orifice was set over more than 100 h. The volume of water decreased gradually and substantially over time. We used Raman and infrared spectroscopy to analyze the residue obtained by evaporating the water present around the orifice. The residue contained organic matter, including carotenoids, amides, esters, and sugars, which were similar to those found in the membranes generated in the orifice in our previous paper, suggesting that the organic matter was synthesized in a wide region of water around the orifice as well as in the orifice. These results may be relevant to the origins of life and biology, and may lead to the development of a technology for reducing CO2 in air, as well as applications in many scientific and engineering fields.

Clinical analysis of EGFR-positive non-small cell lung cancer patients treated with first-line afatinib: A Nagano Lung Cancer Research Group.

BACKGROUND: In the LUX-Lung 3 and LUX-Lung 6 trials, afatinib improved overall survival in previously untreated patients with EGFR 19del mutated non-small cell lung cancer (NSCLC) compared to chemotherapy. The appropriate management of adverse events and dose reduction of afatinib are important for EGFR-positive NSCLC patients. We conducted a retrospective and observational study of patients treated with first-line afatinib for EGFR-positive NSCLC in Nagano prefecture, Japan, focusing on efficacy and toxicities. METHODS: We retrospectively collected the medical records of NSCLC patients initially treated with afatinib between May 2014 and March 2018. RESULTS: A total of 62 patients with a median age of 67 years and a median body surface area (BSA) of 1.57 m2 were included. The overall response rate was 87.7% and median progression-free survival (PFS) was 15.7 months. The median PFS was similar between standard initial dose (40 mg) and reduced initial doses (30 and 20 mg) (15.7 vs. 14.2 months; P = 0.978). The frequency of dose reduction and the discontinuation rate in the 40 mg daily dose group was higher in patients with BSA < 1.58 m2 (100%) compared to BSA ≥ 1.58 m2 (68.2%) (P = 0.014). The frequency of diarrhea was higher in patients with BSA < 1.58 m2 (93.5%) compared to BSA ≥ 1.58 m2 (71.0%) (P = 0.02). CONCLUSION: In real-world clinical practice, first-line afatinib was well managed and was equally as effective as in previous clinical trials of EGFR-positive NSCLC. BSA is considered a predictive marker for appropriate afatinib dose reduction.

[Evaluation of the Cytotoxicity of Commercially Available Nail Adhesives].

Nail tips are nail art materials that can be attached to the nail with adhesives. Recently, nail/finger injuries related to nail tips have been reported and one of the causes is considered to be the adhesives used for attaching nail tips. The components of nail adhesives are mostly cyanoacrylate, which is also used as an industrial instant adhesive. During curing, cyanoacrylate adhesives release formaldehyde through hydrolysis. When it is marketed as a nail adhesive, there is no regulation regarding its formaldehyde amount nor obligation to indicate its ingredients in Japan. Additionally, a biological safety test is not required for nail adhesives. Thus, because the safety of nail adhesives is inadequately confirmed, it is necessary to investigate their biological safety. Therefore, we purchased 5 commercially available nail adhesives and 1 medical adhesive and examined their formaldehyde content and cytotoxicity. We examined the cytotoxicity of the adhesives in V79 cells by a colony forming assay. In this test, 5 nail adhesives showed higher toxicity than 1 medical adhesive. Formaldehyde concentrations in the extract of adhesives were as follows: 17.5 to 24.2 µg/mL for nail adhesives and 7.4 µg/mL for medical adhesives. Cyanoacetate did not elicit cytotoxicity at the final concentration up to 1000 µM. However, formaldehyde showed cytotoxicity, with an IC50 of 79 µM (2.4 µg/mL). Taken together, the cytotoxicity of nail adhesives could be due to the formaldehyde generated by the hydrolysis of cyanoacrylate. It seems important that nail adhesives will be regulated by obligation and enhanced safety in the future.

Organic compounds generated after the flow of water through micro-orifices: Were they synthesized?

Micro-fluid mechanics is an important area of research in modern fluid mechanics because of its many potential industrial and biological applications. However, the field is not fully understood yet. In previous work, when passing ultrapure water (UPW) in which air was dissolved (UPW*) through micro-orifices, we found that the flow velocity decreased and stopped over time, and membranes were frequently formed in the orifice when the flow stopped. The membrane came from the dissolved air in UPW*, and membrane formation was closely related to electric charges generated in orifices by the flow. In the present paper, we clarified the components of the membrane and suggested a mechanism for membrane formation. We examined the effect of contaminants on the membrane formation and confirmed our previous results. We identified the chemical components of the membrane and those present in the UPW* itself by using an electron probe microanalyzer and found that the proportion of each element differed between the membrane and UPW*. Raman and infrared (IR) spectroscopy showed that the membrane consisted of organic substances such as carotenoids, amides, esters, and sugars. We irradiated UPW* with ultraviolet light to cut organic chains that may be left in UPW* as contaminants. We found a similar membrane and organic compounds as in nonirradiated UPW*. Furthermore, although the UPW that was kept from contact with air after it was supplied from the UPW maker (UPW0) and bubbled with Ar gas (UPW0 bubbled with Ar) formed no membrane, the UPW0 bubbled with CO2 formed thin membranes, and Raman and IR analysis showed that this membrane contained carboxylic acid salts, carotenoids, or a mixture of both. We found that electric grounding of the orifice reduces the probability of membrane formation and that the jets issuing from an aperture bear negative charges, and we assumed that the micro-orifices possess positive charges generated by flows. Consequently, we suggest that organic compounds are synthesized from nonorganic matter in air or CO2 dissolved in water by the action of hydroxyl radicals generated by flows through micro-orifices.

Clinical analysis of patients treated with afatinib for advanced non-small cell lung cancer: A Nagano Lung Cancer Research Group observational study.

BACKGROUND: Afatinib has been available in Japan for the treatment of epidermal growth factor receptor (EGFR)-mutated non-small cell lung cancer (NSCLC) since May 2014. We conducted an observational study in patients treated with afatinib in Nagano prefecture, focusing on response and associated toxicities. METHODS: We analyzed the clinical records of NSCLC patients treated with afatinib between May 2014 and February 2015. RESULTS: The records of a total of 73 patients (27 men, 46 women) with a median age of 69 years (range: 42-85 years) were analyzed. Afatinib was administered to 11 patients as a first-line therapy, but it was predominantly administered as a fifth-line or beyond therapy (32 cases, 43.8%). The overall response rates for afatinib as a first-line therapy and beyond second-line therapy were 80% (95% confidence interval [CI]: 55.2-100.0%) and 27.1% (95% CI: 14.5-39.7%), respectively. The main toxicities grade >3 included diarrhea (8.2%), skin rash (6.8%), nausea (6.8%), and appetite loss (6.8%). A low body surface area (BSA) (<1.5m2) was significantly associated with a higher frequency of diarrhea grade >2, compared with a higher BSA (≥ 1.5m2). Forty-eight patients (63.0%) were treated without a dose reduction of afatinib. CONCLUSIONS: Although the survival benefit with afatinib remains unclear, our observational analysis demonstrated the feasibility of using afatinib for EGFR-mutated NSCLC in clinical practice. In particular, a relatively high level of drug delivery is possible. In addition, a lower BSA may be a predictor of diarrhea in patients treated with afatinib.

Development of a low-energy and high-current pulsed neutral beam injector with a washer-gun plasma source for high-beta plasma experiments.

We have developed a novel and economical neutral-beam injection system by employing a washer-gun plasma source. It provides a low-cost and maintenance-free ion beam, thus eliminating the need for the filaments and water-cooling systems employed conventionally. In our primary experiments, the washer gun produced a source plasma with an electron temperature of approximately 5 eV and an electron density of 5 × 10(17) m(-3), i.e., conditions suitable for ion-beam extraction. The dependence of the extracted beam current on the acceleration voltage is consistent with space-charge current limitation, because the observed current density is almost proportional to the 3/2 power of the acceleration voltage below approximately 8 kV. By optimizing plasma formation, we successfully achieved beam extraction of up to 40 A at 15 kV and a pulse length in excess of 0.25 ms. Its low-voltage and high-current pulsed-beam properties enable us to apply this high-power neutral beam injection into a high-beta compact torus plasma characterized by a low magnetic field.

Photomanipulation of antibiotic susceptibility and biofilm formation of Escherichia coli heterologously expressing photoactivated adenylyl cyclase.

A cyaA-deficient Escherichia coli strain was transformed by a plasmid carrying the gene for BsPAC, a photoactivated adenylyl cyclase identified from a Beggiatoa sp., and was subjected to an antibiotic susceptibility assay and biofilm formation assay under a light or dark condition. Cells expressing BsPAC that were incubated under blue light (470 nm) were more susceptible to fosfomycin, nalidixic acid and streptomycin than were cells incubated in the dark. Cells expressing BsPAC formed more biofilms when incubated under the light than did cells cultured in the dark. We concluded from these observations that it is possible to determine the importance of cAMP in antibiotic susceptibility and biofilm formation of E. coli by photomanipulating the cellular cAMP level by the use of BsPAC. A site-directed mutant of BsPAC in which Tyr7 was replaced by Phe functioned even in the dark, indicating that Tyr7 plays an important role in photoactivation of BsPAC. Results of mutational analysis of BsPAC should contribute to an understanding of the molecular basis for photoactivation of the protein.

Thermal and transport properties of ionic liquids based on benzyl-substituted phosphonium cations.

The physicochemical properties of two novel ionic liquids based on benzyltriethylphosphonium and benzyltributylphosphonium cations are described in this report. It was found that both benzyl-substituted phosphonium cations gave low-melting salts in combination with a bis(trifluoromethylsulfonyl)amide anion. The thermogravimetric analysis suggested that the benzyl-substituted phosphonium ionic liquids showed higher thermal stability than those of not only the alkyl-substituted phosphonium ILs but also the corresponding benzyl-substituted ammonium compounds. The benzyl-substituted phosphonium ionic liquids also exhibited relatively high conductivities when compared to those of the corresponding ammonium compounds. These results indicate an improving effect of introducing a benzyl group into the phosphonium cations on both the thermal stability and the conductivity.

Molecular properties of the glucosaminidase AcmA from Lactococcus lactis MG1363: mutational and biochemical analyses.

The major autolysin AcmA of Lactococcus lactis ssp. cremoris MG1363 is a modular protein consisting of an N-terminal signal sequence, a central enzymatic region (glu(acma) as a glucosaminidase), and a C-terminal cell-recognition domain (LysM123). glu(acma) (about 160 amino acids) belongs to the glycoside hydrolase (GH) 73 family, and the two acidic residues E128 and D153 have been thought to be catalytically important. In this study, amino-acid substitution analysis of AcmA was first carried out in the Escherichia coli system. Point mutations E94A, E94Q, E128A, D153A, and Y191A markedly reduced cell-lytic activity (3.8%, 1.1%, 4.2%, 4.8%, and 2.4%, respectively), whereas E128Q and D153N retained significant residual activities (32.1% and 44.0%, respectively). On the other hand, Y191F and Y191W mutations retained high activities (66.2% and 46.0%, respectively). These results showed that E94 (rather than E128 and D153) and the aromatic residue Y191 probably play important roles in catalysis of AcmA. Together with mutational analysis of another GH73 glucoaminidase Glu(atlwm) from the Staphylococcus warneri M autolysin Atl(WM), these results suggested that the GH73 members cleave a glycosidic bond via a substrate-assisted mechanism, as postulated in the GH20 members. AcmA and Glu(atlwm) were purified from E. coli recombinant cells, and their enzymatic properties were studied.

Phase I study of paclitaxel, carboplatin and UFT in chemo-naive patients with advanced non-small cell lung cancer (NSCLC).

OBJECTIVE: We conducted a phase I study of paclitaxel (PTX), carboplatin (CBDCA), and UFT in chemo-naive patients with advanced non-small cell lung cancer (NSCLC). METHOD: Twenty-one chemo-naive patients with advanced NSCLC were enrolled. The study was conducted as a phase I dose-escalation study of various doses of systemic PTX followed by CBDCA on day 1 and oral UFT (400 mg/m2) on days 1-5 and 8-12, with the cycle repeated at 21-day intervals. At least three patients were enrolled in each step. RESULTS: The main toxicities were neutropenia and paresthesia, but were tolerable and reversible in all cases. Overall response rate was 57% (12 out of 21). The MTD was not reached at the highest dose level after the first cycle. Given previous recommends of PTX at 225 mg/m2 and CBDCA AUC 6 for two-drug therapy, the recommended dose for the phase II study under our regimen was set at PTX 225 mg/m2 on day 1, CBDCA AUC 6 on day 1, and UFT 400 mg/m2 on days 1-5 and 8-12. CONCLUSION: The combination of PTX, CBDCA, and UFT showed promising activity and acceptable toxicity in these chemo-naive patients, supporting the development of this combination as a feasible chemotherapeutic option for advanced NSCLC.

Effect of ion skin depth on relaxation of merging spheromaks to a field-reversed configuration.

The effect of ion skin depth on the relaxation of merging spheromaks to a field-reversed configuration (FRC) is studied experimentally for a wide range of size parameter S* (ratio of minor radius to ion skin depth) from 1 to 7. The two merging spheromaks are observed to relax to an FRC or a new spheromak depending on whether the initial poloidal eigenvalue is smaller or larger than a threshold value. The bifurcation value is found to increase with decreasing size parameter S*, indicating that the low-S* condition provides a wide bifurcated range of relaxation to an FRC. The FRC-style relaxation under the low-S* conditions was accompanied by the suppression of the low-n modes (n is the toroidal mode number) activity. The fast rotations of the modes were followed by suppression of the low-n modes.

Influence of daily low-dose 14-membered-ring macrolide therapy on Helicobacter pylori infection in patients with chronic inflammatory disease of the airway.

Our aim was to evaluate the effects of eradication and the incidence of secondary resistance by long-term low-dose daily 14-membered-ring macrolide therapy on Helicobacter pylori ( H. pylori) infection in patients with chronic lower respiratory tract inflammatory disease. In a retrospective analysis, we studied the seroprevalence of H. pylori IgG in 90 patients with inflammation of the lower respiratory tract (68 had been treated with macrolide and 22 served as controls). Then, in a prospective analysis, we evaluated the eradication effect of macrolide therapy by the decline of IgG values and the (13)C-urea breath test. Only long-term macrolide use significantly affected the seroprevalence of H. pylori IgG. However, macrolide therapy did not reduce the H. pylori IgG values in 24 patients and did not eradicate H. pylori in (13)C-urea breath tests. Chemosensitivity testing was performed on three H. pylori strains obtained by gastric biopsy from patients in whom the disease could not be eradicated. Only one strain demonstrated a resistant character. Daily long-term low-dose 14-membered-ring macrolide therapy for patients with lower respiratory inflammatory disease may not be sufficient to eradicate H. pylori, but some strains do not acquire a resistant nature.

["Tube scratching" in the trachea on nasotracheal intubation: comparison of Macintosh and McCoy laryngoscopes].

BACKGROUND: "Tube scratching" in the trachea, the intratracheal resistance of a tube, on nasotracheal intubation was evaluated using the Macintosh and the McCoy laryngoscope blade. METHODS: Fifty young patients requiring nasotracheal intubation were studied. Following induction of anesthesia, X-ray was taken before and during the use of two types of laryngoscopes. For analysis of the configuration of the upper airway, we drew three straight lines [axis of pharynx (P), larynx (L) and trachea (T)] on each film. "Tube scratching" was evaluated and graded to four by a supporting anesthesiologists. RESULTS: "Tube scratching" in the trachea was observed in 14/25 patients with McCoy laryngoscope and 4/25 patients with Macintosh laryngoscope (P < 0.05). Intubation with McCoy laryngoscope decreased more the angles formed by P and L, and L and P those that with Macintosh laryngoscope (P < 0.01). These data indicate that the lines of P, L and T tend not to align with McCoy laryngoscope, preventing the tracheal tube to be passed smoothly. CONCLUSION: An excessive lifting of the epiglottis, often observed in using McCoy laryngoscope, causes "Tube scratching" in the trachea on nasotracheal intubation.