Abstracts of papers (2009)

Last Update: 12/03/2009

Abstracts of papers (2009)

[2009-1] Yoshida, M. et al., Biochem Biophys Res. Commun. 378, 313-318 (2009)

The toxicity of acrolein was compared with that of reactive oxygen species using a mouse mammary carcinoma FM3A cell culture system. Complete inhibition of cell growth was accomplished with 10 [micro]M acrolein, 100 [micro]M H2O2, and 20 [micro]M H2O2 plus 1 mM vitamin C, which produce ·OH, suggesting that toxicity of acrolein is more severe than H2O2 and nearly equal to that of ·OH, when these compounds were added extracellularly. Acrolein toxicity was prevented by N-acetyl-L-cysteine and N-benzylhydroxylamine, and attenuated by putrescine and spermidine. Toxicity of H2O2 was prevented by glutathione peroxidase plus N-acetyl-L-cysteine, pyruvate, catalase, and reduced by polyphenol, and toxicity of ·OH was prevented by glutathione peroxidase plus N-acetyl-L-cysteine, pyruvate, catalase and reduced by N-acetyl-L-cysteine. The results indicate that prevention of cell toxicity by N-acetyl-L-cysteine was more effective with acrolein than with ·OH. Protein and DNA synthesis was damaged primarily by acrolein and reactive oxygen species, respectively.

[2009-2] Yoshida, M. et al., Atherosclerosis 203, 557-562 (2009)

We found previously that increased levels of polyamine oxidase (PAO) [acetylpolyamine oxidase (AcPAO) plus spermine oxidase (SMO)], and acrolein (CH2=CHCHO) are good markers of stroke. We then investigated whether silent brain infarction (SBI) can be detected by measuring acrolein, PAO, or other biomarkers. Several biomarkers were measured in the plasma of 53 normal subjects and 44 subjects with SBI. It was found that the levels of protein-conjugated acrolein (PC-Acro), interleukin-6 (IL-6) and C-reactive protein (CRP) were significantly higher in SBI than in normal subjects. PAO was slightly higher in SBI than in normal subjects. Since the probability of SBI was increased with age, values were analyzed including age as a factor. When the combined measurements of PC-Acro, IL-6 and CRP were evaluated together with age using a receiver operating characteristic curve, SBI was indicated with 89% sensitivity and 91% specificity. The results indicate that measurement of PC-Acro together with IL-6 and CRP makes it possible to identify SBI with high sensitivity and specificity.

[2009-3] Terui, Y. et al., J. Bacteriol. 191, 5348-5357 (2009)

Proteins whose synthesis is enhanced by polyamines at the level of translation were identified with a polyamine-requiring mutant cultured in the presence of 0.1% glucose and 0.02% glutamate at 42 °C. Polyamines had a greater effect on cell growth at 42 °C than at 37 °C. At 42 °C, the synthesis of RpoE (sigma24) and StpA, which are involved in the transcription of a number of heat shock response genes, was stimulated by polyamines at the level of translation. In the rpoE and stpA mRNAs, a Shine-Dalgarno (SD) sequence is located at 13 and 12 nucleotides, respectively, upstream of the initiation codon AUG. When the SD sequences were moved to the more common position 7 nucleotides upstream of the initiation codon AUG, the degree of polyamine stimulation was reduced, although the level of RpoE and StpA synthesis was markedly increased. The mechanism underlying polyamine stimulation of RpoE synthesis was then studied. Polyamine stimulation of RpoE synthesis was reduced by changing the bulged-out structure in the initiation site of rpoE mRNA, although the level of RpoE synthesis increased. A selective structural change of this bulged-out region induced by spermidine at 42 °C was observed by circular dichroism. Polyamine stimulation of fMet-tRNA binding to ribosomes at 42 °C also disappeared by changing the bulged-out structure in the initiation site of rpoE mRNA. The results suggest that polyamines enhance the synthesis of RpoE by changing the bulged-out structure in the initiation site of rpoE mRNA.

[2009-4] Nishimura, K. et al., Int. J. Biochem. Cell Biol. 41, 2251-2261 (2009)

In Escherichia coli, several proteins whose synthesis is enhanced by polyamines at the level of translation have been identified. We looked for proteins that are similarly regulated in eukaryotes using a mouse mammary carcinoma FM3A cell culture system. Polyamine deficiency was induced by adding an inhibitor of ornithine decarboxylase, alpha-difluoromethylornithine, to the medium. Proteins enhanced by polyamines were determined by comparison of protein levels in control and polyamine-deficient cells using two-dimensional gel electrophoresis, and were identified by Edman degradation and/or LC/MALDI-TOF/TOF tandem mass spectrometry. Polyamine stimulation of the synthesis of these proteins at the level of translation was confirmed by measuring levels of the corresponding mRNAs and proteins, and levels of the [35S]methionine pulse-labeled proteins. The proteins identified in this way were T-complex protein 1, beta subunit (Cct2); heterogeneous nuclear ribonucleoprotein L (Hnrpl); and phosphoglycerate mutase 1 (Pgam1). Since Cct2 was most strongly enhanced by polyamines among three proteins, the mechanism of polyamine stimulation of Cct2 synthesis was studied using NIH3T3 cells transiently transfected with genes encoding Cct2-EGFP fusion mRNA with normal or mutated 5'-untranslated region (5'-UTR) of Cct2 mRNA. Polyamines most likely enhanced ribosome shunting on the 5'-UTR of Cct2 mRNA.

[2009-5] Saiki, R. et al., Stroke 40, 3356-3361 (2009)

Background and Purpose - We recently found that increases in plasma levels of protein-conjugated acrolein and polyamine oxidases, enzymes that produce acrolein, are good markers for stroke. The aim of this study was to determine whether the level of protein-conjugated acrolein is increased and levels of spermine and spermidine, the substrates of acrolein production, are decreased at the locus of infarction.
Methods - A unilateral infarction was induced in mouse brain by photoinduction after injection of Rose Bengal. The volume of the infarction was analyzed using the public domain National Institutes of Health image program. The level of protein-conjugated acrolein at the locus of infarction and in plasma was measured by Western blotting and enzyme-linked immunosorbent assay, respectively. The levels of polyamines at the locus of infarction and in plasma were measured by high-performance liquid chromatography.
Results - The level of protein-conjugated acrolein was greatly increased, and levels of spermine and spermidine were decreased at the locus of infarction at 24 hours after the induction of stroke. The size of infarction was significantly decreased by N-acetylcysteine, a scavenger of acrolein. It was also found that the increases in the protein-conjugated acrolein, polyamines, and polyamine oxidases in plasma were observed after the induction of stroke.
Conclusions - The results indicate that the induction of infarction is well correlated with the increase in protein-conjugated acrolein at the locus of infarction and in plasma.

[2009-6] Masuko, T. et al., J. Pharmacol. Exp. Ther. 331, 522-530 (2009)

Tosyl-polyamine derivatives such as N-{4-[4-(guanidinobutylamino)-butylamino]butyl}-4-methylbenzenesulfonamide trihydrochroride (TsHSPMG) have been found to strongly inhibit macroscopic currents through heteromeric N-methyl-D-aspartate (NMDA) receptors (NR1/NR2A, NR1/NR2B) and Ca2+-permeable alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (homomeric glutamate receptor 1) receptors expressed in Xenopus laevis oocytes on voltage-clamp recording. In the present study, it was found that the inhibition of NMDA receptor activity induced by tosyl-polyamine derivatives was voltage-dependent. Some mutations located in the intracellular region of the channel pore, such as NR1 E621Q and NR2B W607L, reduced the inhibition by tosyl-polyamine derivatives, suggesting that tosyl-polyamine derivatives penetrate deeply into the channel pore of NMDA receptors. The neuroprotective effects of tosyl-polyamine derivatives against cell injury caused by NMDA were investigated in cultured rat hippocampal neurons. Addition of 1 [micro]M TsHSPMG to medium ablated the neurotoxicity induced by NMDA, and a similar effect was observed with 30 [micro]M memantine. The neuroprotective effects of tosyl-polyamine derivatives on NMDA-induced seizures in mice were also assayed. Intracerebroventricular or intravenous injection of TsHSPMG (0.1 or 0.5 mg/kg) decreased the seizures induced by intraperitoneal injection of NMDA in mice. These findings indicate that tosyl-polyamine derivatives exhibit neuroprotective effects not only in primary cultured neurons but also in mice.

[2009-7] Uemura, T. et al., Int. J. Biochem. Cell Biol. 41, 2538-2545 (2009)

We proposed that a group of genes whose expression is enhanced by polyamines at the level of translation in Escherichia coli and mammalian cells be referred to as a "polyamine modulon". In Saccharomyces cerevisiae, proteins whose synthesis is enhanced by polyamines at the level of translation were searched for using a polyamine-requiring mutant of S. cerevisiae deficient in ornithine decarboxylase (YPH499 Deltaspe1). Addition of spermidine to the medium recovered the spermidine content and enhanced cell growth of the YPH499 Deltaspe1 mutant by 3-5-fold. Under these conditions, synthesis of COX4, one of the subunits of cytochrome C oxidase (complex IV), was enhanced by polyamines about 2.5-fold at the level of translation. Accordingly, the COX4 gene is the first member of a polyamine modulon in yeast. Polyamines enhanced COX4 synthesis through stimulation of the ribosome shunting of the stem-loop structures (hairpin structures) during the scanning of the 5'-untranslated region (5'-UTR) of COX4 mRNA by 40S ribosomal subunit-Met-tRNAi complex.