A concise synthesis of peramine, a metabolite of endophytic fungi

The total synthesis of peramine, a natural product isolated from an endophytic fungi, has been achieved in four steps and 34% overall yield from known compounds. The key step was the one-pot construction of the pyrrolopyrazinone ring from pyrrole amide and propargyl bromide. The preparation of peramine-d4 as an internal standard for quantitative analysis by MS is also described. Peramine, a natural product isolated from an endophytic fungi, was synthesized in four steps by one-pot construction of the pyrrolopyrazinone ring as a key step.</p

Adhesive Sulfabetaine Polymer Hydrogels for the Sandwich Cell Culture

Sandwich culture systems are techniques that cultivate cells by sandwiching them between the top and bottom substrates. Since the substrates can be separated, the system is expected to be applied to the construct layering of patterned cells and to the isolation of stacked cells. In this study, we prepared hydrogels composed of zwitterionic sulfabetaine polymers, poly[2-(2-(methacryloyloxyethyl)dimethylammonio)ethyl-1-sulfate] (PZBMA). The ZBMA homopolymers have been shown to form aggregates in aqueous solutions due to their intermolecular interactions. The water content of the PZBMA hydrogels in water was ∼70% regardless of N,N′-methylenebis(acrylamide), BIS, content as the cross-linker. The results indicated that the intermolecular interaction contributed more to the swelling behaviors than the chemical cross-linker. However, PZBMA hydrogels with 0.1 mol % BIS showed not only high elongation (∼850%) properties but also high adhesiveness and self-healing properties. When this PZBMA hydrogel was impregnated with collagen and subjected to sandwich culture using Madin–Darby canine kidney (MDCK) cells, a three-dimensional morphology of MDCK cell aggregates was constructed. Such a sulfabetaine hydrogel is expected to be developed for regenerative medicine

Adhesive Sulfabetaine Polymer Hydrogels for the Sandwich Cell Culture

Sandwich culture systems are techniques that cultivate cells by sandwiching them between the top and bottom substrates. Since the substrates can be separated, the system is expected to be applied to the construct layering of patterned cells and to the isolation of stacked cells. In this study, we prepared hydrogels composed of zwitterionic sulfabetaine polymers, poly[2-(2-(methacryloyloxyethyl)dimethylammonio)ethyl-1-sulfate] (PZBMA). The ZBMA homopolymers have been shown to form aggregates in aqueous solutions due to their intermolecular interactions. The water content of the PZBMA hydrogels in water was ∼70% regardless of N,N′-methylenebis(acrylamide), BIS, content as the cross-linker. The results indicated that the intermolecular interaction contributed more to the swelling behaviors than the chemical cross-linker. However, PZBMA hydrogels with 0.1 mol % BIS showed not only high elongation (∼850%) properties but also high adhesiveness and self-healing properties. When this PZBMA hydrogel was impregnated with collagen and subjected to sandwich culture using Madin–Darby canine kidney (MDCK) cells, a three-dimensional morphology of MDCK cell aggregates was constructed. Such a sulfabetaine hydrogel is expected to be developed for regenerative medicine

Lansoprazole activates the Nrf2/ARE pathway in RL34 cells.

A: Representative immunoblot for Nrf2 expression levels in the nuclear and cytoplasmic protein lysates obtained from cells treated with 100 μM of lansoprazole for 3 h; α-tubulin and histone H1 were loading controls for the cytoplasmic and nuclear lysates, respectively (left). Densitometric quantification of the representative immunoblots was performed using ImageJ, and values were normalized to each of the loading control protein level (right). B: Representative immunocytochemical images of the subcellular localization of Nrf2. RL34 cells were treated with 100 μM of lansoprazole for 3 h. Cells were fixed in 4% PFA for 15 min, followed by staining with an anti-Nrf2 antibody and an Alexa Fluor 488-conjugated goat anti-rabbit IgG antibody. Counterstaining of the nuclei was performed with Hoechst 33342. The scale bar represents 50 μm. C and D: Relative mRNA expression levels of Nrf2-induced genes HO1, NQO1, and GSTA2 in cells treated with 100 μM lansoprazole for 3 h. RNA levels were normalized to those of β-actin (C) or GAPDH (D). E: Induction of the ARE-dependent luciferase reporter activity by lansoprazole. RL34 cells stably expressing an ARE-reporter gene were treated with 100 μM of lansoprazole for 3 h, and the luciferase activity in the cell lysates was measured. F: Degradation rate of the Nrf2 protein after CHX chase. Cells were treated with 100 μM of lansoprazole for 3 h and were then treated with 10 μM of CHX for the indicated periods. Cell lysates were used for immunoblotting analysis with an anti-Nrf2 and anti-β-actin antibody. Intensities of the Nrf2 bands were quantified and plotted on a semilog graph to obtain half-life values. Abbreviations used in the figure: LPZ, lansoprazole; CHX, cycloheximide. Data represent the mean ± SD of the three independent experiments performed in triplicate. Statistical analysis was performed by using Student’s t-test. *: p vs. DMSO-treated cells.</p

Activation of p38 MAPK is required for the lansoprazole-induced ARE/Nrf2 pathway in RL34 cells.

A: Effect of SB203580 on the induction of Nrf2 protein expression by lansoprazole. RL34 cells were treated with 10 μM of SB203580 (a specific p38 MAPK inhibitor) for 30 min, and then exposed to 100 μM of lansoprazole for an additional 3 h. Whole cell protein lysates were analyzed by immunoblotting with an anti-Nrf2 antibody; β-actin served as a loading control. B: Effect of SB203580 on the induction of the ARE-dependent reporter activity by lansoprazole. RL34 cells stably expressing the ARE-reporter gene were pretreated with 10 μM of SB203580 for 30 min, and then exposed to 100 μM of lansoprazole for an additional 3 h. Luciferase activity in the cell lysates was measured. C and D: Total RNA was isolated from the cells. Relative mRNA expression levels of HO1 were measured by quantitative RT-PCR. RNA levels were normalized to β-actin (C) and GAPDH (D) mRNA level. Data represent the mean ± SD of three independent experiments performed in triplicate. Student’s t-test was employed for statistical analysis. *: p vs. untreated cells. E: Representative immunocytochemical images of the subcellular localization of Nrf2. RL34 cells were treated with 10 μM of SB203580 for 30 min and then exposed to 100 μM of lansoprazole for an additional 3 h. Cells were fixed in 4% PFA for 15 min, followed by a staining with anti-Nrf2 antibody and an Alexa Fluor 488-conjugated goat anti-rabbit IgG antibody; counterstaining of the nuclei was performed with Hoechst 33342. The scale bar represents 50 μm. Abbreviation: LPZ, lansoprazole.</p

Original uncropped images underlying all blots.

This file contains all uncropped blot information. The asterisk indicates a nonspecific protein band. (PDF)</p

Size Specifically High Activity of Ru Nanoparticles for Hydrogen Oxidation Reaction in Alkaline Electrolyte

The hydrogen oxidation reaction (HOR) in alkaline electrolyte was conducted on carbon-supported Ru nanoparticles (Ru/C) of which size was controlled in the range from approximately 2 to 7 nm. The HOR activity of Ru/C normalized by the metal surface area showed volcano shaped dependence on the particle size with a maximum activity at approximately 3 nm. The HOR activity of approximately 3 nm Ru/C was higher than commercially available Pt nanoparticles (ca. 2 nm) supported on carbon. The structural analysis of Ru/C using Cs-corrected scanning transmission electron microscopy with atomic resolution revealed the unique structural change of Ru/C different from Pt/C: Ru nanoparticle structure changed from amorphous-like structure below 3 nm to metal nanocrystallite with roughened surface at approximately 3 nm and then to that with well-defined facets above 3 nm, although Pt/C kept well-defined facets even at approximately 2 nm. It is proposed that the generation of unique structure observed on approximately 3 nm Ru nanoparticles, that is, long bridged coordinatively unsaturated Ru metal surface atoms on its nanocrystallite, is a key to achieve high HOR activity

The primer sequences used for quantitative RT-PCR.

The primer sequences used for quantitative RT-PCR.</p

Activation of p38 MAPK is required for the cytoprotective activity of lansoprazole against cisplatin-induced toxicity.

Effect of SB203580 on the cytoprotective activity of lansoprazole. RL34 cells were treated with 100 μM of lansoprazole for 3 h in the presence or absence of 10 μM of an SB203580 (a specific p38 MAPK inhibitor) pretreatment for 30 min and were then exposed to 20 μM of cisplatin for an additional 24 h. Cell viability was quantified using an MTS assay. Data are expressed as a percentage of viability of nontreated cells. Data represent the mean ± SD of three independent experiments performed in triplicate. Statistical analysis was performed using ANOVA followed by Tukey’s test for multiple comparisons. *: p < 0.01. Abbreviations: LPZ, lansoprazole; MTS, 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt.</p

Lansoprazole suppresses the cisplatin-induced cytotoxicity.

Cell viability of cisplatin-treated cells exposed to a lansoprazole pretreatment. RL34 cells were pretreated with 100 μM of lansoprazole for 3 h and then exposed to 20 μM of cisplatin for an additional 24 h. HO1 inhibitor SnMP was administered simultaneously with lansoprazole. Cell viability was quantified using an MTS assay. Data are expressed as a percentage of viability when compared with the viability of the DMSO-treated cells. Data represent the mean ± SD of three independent experiments performed in triplicate. Statistical analysis was performed by using ANOVA followed by Tukey’s test for multiple comparisons. *: p < 0.01. Abbreviations: LPZ, lansoprazole; SnMP, tin-mesoporphyrin IX; MTS, 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium,inner salt.</p