The role of the helper lipid dioleoylphosphatidylethanolamine (DOPE) for DNA transfection cooperating with a cationic lipid bearing ethylenediamine

Gene therapy is expected to treat various incurable diseases including viral infections, autoimmune disorders, and cancers. Cationic lipids (CL) have been used as carriers of therapeutic DNAs for gene therapy because they can form a complex with DNA and such a complex can be incorporated into cells and transport the bound DNA to cytosol. The CL/DNA complexes are called lipoplexes and categorized as a non-viral vector. Lipoplexes are often prepared by adding a neutral phospholipid dioleoylphosphatidylethanolamine (DOPE) to CL in order to enhance transfection. However, the role of DOPE is not fully understood. We synthesized a new CL having an ethylenediamine cationic head group, denoted by DA, and found that addition of DOPE to DA achieved a good efficiency, almost in the similar level of commonly used transfection reagent Lipofectamine 2000 (Invitrogen). The composition of DA:DOPE = 1:1 showed the highest efficiency. This lipoplex showed structural transition when pH was changed from 7 to 4, corresponding pH lowering in late endosome, while DOPE itself showed structural transition at more basic pH around 8. The present data showed that the DOPE/DA composition determines the structural transition pH and choosing a suitable pH, i.e., a suitable composition, is essential to increase the transfection efficiency.     

Synthesis and evaluation of N-alkyl-S-[3-(piperidin-1-yl)propyl]isothioureas: High affinity and human/rat species-selective histamine H3 receptor antagonists

S-Alkyl-N-alkylisothiourea compounds containing various cyclic amines were synthesized in the search for novel nonimidazole histamine H₃ receptor (H₃R) antagonists. Among them, four N-alkyl S-[3-(piperidin-1-yl)propyl]isothioureas 18, 19, 22, and 23 were found to exhibit potent and selective H₃R antagonistic activities against in vitro human H₃R, but were inactive against in vitro human H₄R. Furthermore, three alkyl homologs 18–20 showed inactivity for histamine release in in vivo rat brain microdialysis, suggesting differences in antagonist affinities between species. In addition, in silico docking studies of N-[4-(4-chlorophenyl)butyl]-S-[3-piperidin-1-yl)propyl]isothiourea 19 and a shorter homolog 17 with human/rat H₃Rs revealed that structural differences between the antagonist-docking cavities of rat and human H₃Rs were likely caused by the Ala122/Val122 mutation.     

Real-Time Visualization of Neuronal Activity during Perception

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Excitotoxicity-induced immediate surge in hippocampal prostanoid production has latent effects that promote chronic progressive neuronal death

Excitotoxicity is involved in neurodegenerative conditions. We investigated the pathological significance of a surge in prostaglandin production immediately after kainic acid (KA) administration [initial phase], followed by a sustained moderate elevation in prostaglandin level [late phase] in the hippocampus of juvenile rats. Numerous pyknotic hippocampal neurons were observed 72 h after KA treatment; this number remained elevated on days 10 and 30. Gross hippocampal atrophy was observed on days 10 and 30. Pre-treatment with indomethacin ameliorated neuronal death on days 10 and 30, and prevented hippocampal atrophy on day 30. Microglial response was moderated by the indomethacin pre-treatment. Blockade of only late-phase prostaglandin production by post-treatment with indomethacin ameliorated neuronal death on day 30. These findings suggest a role for initial-phase prostaglandin production in chronic progressive neuronal death, which is exacerbated by late-phase prostaglandin production. Blockade of prostaglandin production has therapeutic implications in preventing long-term neurological sequelae following excitotoxic brain damage.     

Formation and Some Properties of the Acid Phosphatase in Aspergillus terreus

Substrate specificity of purified preparations of phytase from Asp, terreus was examined. The enzyme showed broad specificity. It was found that Asp, terreus produced only one kind of acid phosphatase and it had phytase activity.

Effective materials for the enzyme formation were examined. The formation of the enzyme occurred only during times that mycelia was in contact with inositol.

By differential centrifugation and electron-microscopic autoradiography, it was determined that inositol was incorporated into the mycelia and that it was located at almost the same point as where the active enzyme was located.     

Candidusin A and B: New p-Terphenyls with Cytotoxic Effects on Sea Urchin Embryos

Three fungal trichothecenes, verrucarin A, roridin A and 8-β-hydroxyroridin E, were isolated as callus-initiating promoters from Myrothecium sp. 301. These trichothecenes promoted callus induction, synergistically coupled with a low concentration of 2,4-dichlorophenoxyacetic acid.     

The Formation of Higher Alcohols in the Fermentation of Amino Acids by Yeast

We have found that some straight-chained α-amino acids are converted by yeast to the alcohols with correspondingly longer carbon chains in the alcoholic fermentation contrary to F. Ehrlich’s scheme, i.e., isobutyl alcohol from alanine and active amyl alcohol from α-amino-n-butyric acid or threonine.

In this report, we confirmed this fact in the alcoholic fermentation of many aliphatic amino acids by 2 yeast strains using gas chromatography. Moreover, n-propyl alcohol was proved to come from α-amino-n-butyric acid or threonine. Small quantities of n-propyl, isobutyl, active amyl and isoamyl alcohols were found in all the fermented solutions. There was some difference in the composition of higher alcohols of the alcoholic solutions fermented by different yeasts.     

Chemical and Physicochemical Properties of Phytase from Aspergillus terreus

Some chemical and physicochemical properties of the purified phytase preparation produced by Asp. terreus were investigated. From the results of the examination of amino acid analysis, it was suggested that there existed some components other than amino acids in the purified enzyme. Examination of the neutral sugar analysis, therefore, was made by gaschromatography, and it was found that the purified enzyme preparation contained mannose, galactose and a small amount of inositol.

The molecular weight of the enzyme was found to be 214,000 by the Archibald method, and 2.2~2.3×10⁵ by gel-filtration on a Sephadex G–200 column. It was found that by guanidine hydrochloride or by urea, the purified enzyme preparation was dissociated into only one kind of subunit. The native enzyme was supposed to be a homohexamer of the subunits whose molecular weight is 37,000.     

Metabolisms of Nucleosides in Bacteria

In this study, the authors developed a simplified method for the separation and the quantitative determination of nucleosides and bases, using paper electraophoretic technique. By this method, nucleosides and bases were well separated and determined in a fairly short time. Thus, this method was expected to be as accurate as the published methods and was believed to be a better method for both quantitative determination and detection of the nucleosides and bases in a large number of samples.