Expression analysis of cellulose synthases that comprise the Type II complex in hybrid aspen.

Gene duplication in plants occurs via several different mechanisms, including whole genome duplication, and the copied genes acquire various forms and types. The cellulose synthase (CesA) family functions in cellulose synthesis complex (CSC) formation, which is involved in the synthesis of primary and secondary cell walls in plants. In the genome of Populus, 17 CesA have been annotated, and some of them appeared through whole genome duplication. The nucleotide sequence of the duplicated genes changed during subsequent evolution, and functional differentiation of genes might have occurred. To gain insight into the evolutionary fate of the duplicated CesA, expression analysis with quantitative reverse transcription polymerase chain reactions and promoter-reporter assays was performed on three duplicated gene pairs whose products have been reported to form a single CSC. Changes in expression of each gene at different developmental stages were detected and divergent expression patterns in different organs and tissues observed between the gene pairs. Among the tested genes, expression of PttCesA3-C was apparently lower than that of its counterpart, PttCesA3-D. The results suggest that the six CesA are approaching sub-functionalisation or non-functionalisation. Furthermore, the level of functionalisation may vary among the three pairs of genes, and functional specialisation of each CesA should have been achieved, at least partially, through differences in expression of genes.

Development of apertureless near-field scanning optical microscope tips for tip-enhanced Raman spectroscopy.

In this study, we suggested two types of novel metallized tip for the apertureless near-field scanning optical microscope probe. The first is a silver nanorod immobilized tip and the other is a double metallized probe. We calculated the electric field enhancement factor and the electric field distribution of a single sphere, aggregated spheres, an ellipse and a nanorod by the finite-differential time-domain method to improve the silver nanosphere immobilized tip developed in our previous studies. The enhanced field of the nanorod is localized at the external surfaces. The simulation results of the nanorod revealed that the position of the maximum peak is shifted to a longer wavelength and that its electric field enhancement factor increases as the aspect ratio increases. Thus, we developed the silver nanorod immobilized tip, and the tip-enhanced Raman spectrum of rhodamine 6G molecule on the substrate could be measured by the tip though it could not be detected by the previous nanosphere immobilized tip. Further, the finite-differential time-domain calculation predicted that the double metallized tips considerably enhance the electric field and that its enhancement factor in the longer wavelength region (500-600 nm) does not decrease when the tip is rounded. The results show that the proposed metallized tips were useful for the apertureless near-field scanning optical microscope system.

Characterization of factors stimulating differentiation of mouse myeloid leukemia cells from a Yoshida sarcoma cell line cultured in serum-free medium.

A clone, YS-T22, of cells from Yoshida sarcoma cell line, YSSF-212T, grown in "serum-free" culture medium produced factors stimulating differentiation of mouse myeloid leukemia cells (M1) to macrophages and granulocytes. The formation of macrophages and granulocytes was accompanied by induction of phagocytosis, locomotive activity, and lysosomal enzyme activities. The rates of induction of these differentiated phenotypes were proportional to the concentration of the factor added and the period of treatment. The factor stimulating differentiation of M1 cells was a heat-labile, nondialyzable proteinaceous substance that was inactivated by trypsin but not by ribonuclease or glycosidases. On diethylaminoethyl cellulose chromatography, the factor stimulating differentiation of M1 cells from conditioned medium of YS-T22 cells was eluted in various fractions with or without activity of the colony-stimulating factor.

Cleavages of aromatic ring and beta-O-4 bond of synthetic lignin (DHP) by lignin peroxidase.

Lignin peroxidase from a white-rot basidiomycete, Phanerochaete chrysosporium, catalyzed cleavages of the aromatic ring and the beta-O-4 bond of a synthetic lignin, a dehydrogenation copolymer (DHP) of coniferyl alcohol and a (beta-O-4)-(beta-beta) lignin substructure model trimer.

Aromatic ring cleavage of 4,6-di(tert-butyl)guaiacol, a phenolic lignin model compound, by laccase of Coriolus versicolor.

It was found that 2,4-di(tert-butyl)-4-(methoxycarbonylmethyl)-2-buten-4-ol ide (II) was formed as an aromatic ring cleavage product of a phenolic lignin model compound, 4,6-di(tert-butyl)guaiacol (I), by laccase of Coriolus versicolor. Based on isotopic experiments with 18O2 and H2 18O, the mechanism of formation of II from I is discussed.

Regiospecific oxygenations during ring cleavage of a secondary metabolite, 3,4-dimethoxybenzyl alcohol catalyzed by lignin peroxidase.

Enzymatic oxidation of veratryl alcohol yielded a new ring cleavage product (delta-lactone) in addition to the two known gamma-lactone products. The experiment with 18O-enriched water and dioxygen clearly showed that one oxygen atom each from water and dioxygen is specifically incorporated into the cleavage product at the original C3 or C4 position of 3,4-dimethoxybenzyl alcohol. A new type of reaction mechanism proposed for the ring cleavage of this compound is rationally explained in good accord with the one-electron transfer mechanism.

Changes in quinone profiles of hot spring microbial mats with a thermal gradient

The respiratory and photosynthetic quinones of microbial mats which occurred in Japanese sulfide-containing neutral-pH hot springs at different temperatures were analyzed by spectrochromatography and mass spectrometry. All of the microbial mats that developed at high temperatures (temperatures above 68 degreesC) were so-called sulfur-turf bacterial mats and produced methionaquinones (MTKs) as the major quinones. A 78 degreesC hot spring sediment had a similar quinone profile. Chloroflexus-mixed mats occurred at temperatures of 61 to 65 degreesC and contained menaquinone 10 (MK-10) as the major component together with significant amounts of either MTKs or plastoquinone 9 (PQ-9). The sunlight-exposed biomats growing at temperatures of 45 to 56 degreesC were all cyanobacterial mats, in which the photosynthetic quinones (PQ-9 and phylloquinone) predominated and MK-10 was the next most abundant component in most cases. Ubiquinones (UQs) were not found or were detected in only small amounts in the biomats growing at temperatures of 50 degreesC and above, whereas the majority of the quinones of a purple photosynthetic mat growing at 34 degreesC were UQs. A numerical analysis of the quinone profiles was performed by using the following three parameters: dissimilarity index (D), microbial divergence index (MDq), and bioenergetic divergence index (BDq). A D matrix tree analysis showed that the hot spring mats consisting of the sulfur-turf bacteria, Chloroflexus spp., cyanobacteria, and purple phototrophic bacteria formed distinct clusters. Analyses of MDq and BDq values indicated that the microbial diversity of hot spring mats decreased as the temperature of the environment increased. The changes in quinone profiles and physiological types of microbial mats in hot springs with thermal gradients are discussed from evolutionary viewpoints.

Purification and partial sequences of Aralia cordata cinnamyl alcohol dehydrogenase.

Cinnamyl alcohol dehydrogenase (CAD) (EC 1.1.1.195) from a dicot, Aralia cordata, was purified to homogeneity and its properties were characterized. The enzyme shows a preference for cinnamyl alcohols and cinnamyl aldehydes as substrates. The M(r) is estimated at 72,000. The enzyme is composed of two heterogeneous subunits of slightly different sizes, and it differs from the bean enzyme in the size of subunits. Partial amino acid sequencing of the purified enzyme was carried out both from the N-terminus and using selected peptides obtained by cyanogen bromide cleavage.

A case of undifferentiated glioma in a 70-year-old woman.

The present case involved a 70-year-old woman who was diagnosed with a right cerebral hemorrhage. Excisional surgery of the hematoma was performed. Grossly, a whitish, solid tumor (1 x 1 x 0.8 cm in size) was recognized in the hematoma. Histologically, the tumor was composed of large, polygonal cells and small undifferentiated cells in a jumbled architectural arrangement with a cartilage component. The large, polygonal cell component was conspicuous and somewhat rhabdoid in appearance and appeared to be an astrocytic tumor showing glial differentiation. The small, undifferentiated cell component resembled tumor cells of a primitive neuroectodermal tumor (PNET). Clinical follow-up of the patient for 2 months after the first operation revealed recurrence with rapid growth. A second operation was performed, but the patient died 8 months after the first operation (2 months after the second). Immunohistochemically, the tumor cells suggesting glial differentiation were positive for glial fibrillary acidic protein (GFAP), S-100, neuron-specific enolase (NSE), and vimentin. PNET-like components in the primary tumor were positive for NSE, GFAP, and S-100, and weakly positive for vimentin and synaptophysin. Each tumor cell was negative for epithelial membrane antigen (EMA), keratin, desmin, actin, myoglobin, neurofilament (NF), and MIC2 protein. The recurrent tumor revealed predominantly PNET-like components; however, only a few tumor cells were positive for GFAP. This appearance suggested that this brain tumor might originate from a common multipotential stem cell. Considering its histopathological and immunohistochemical characteristics, the primary tumor was finally regarded as an undifferentiated glioma with dedifferentiation of the glial component in the recurrent tumor.

Comparisons of the pharmacokinetics and the leukopenia and thrombocytopenia grade after administration of irinotecan and 5-fluorouracil in combination to rats.

BACKGROUND: Irinotecan (CPT-11) has been used recently for the treatment of several cancers in combination with 5-fluorouracil (5-FU). Preliminary data on this combination therapy in humans demonstrated no drug interactions between CPT-11 (or its metabolite, SN-38) and 5-FU however, because there is so little information on the combination, the possibility for an interaction still exists. MATERIAL AND METHODS: CPT-11 and 5-FU were injected intravenously into rats and the pharmacokinetics of CPT-11 and SN-38 and alternations in blood cell count were investigated. RESULTS: In the group of rats administered 5-FU prior to CPT-11, the area under the concentration-time curve (AUC) of CPT-11 was approximately eight-fold larger compared with the group administered CPT-11 prior to 5-FU. On the other hand, the grade of leukocytopenia or thrombocytopenia was not significantly different among the two groups. CONCLUSION: In rats, the conversion of CPT-11 into SN-38 is possibly delayed by prior administration of 5-FU.

Effect of diltiazem and thromboxane A2 synthetase inhibitor (OKY-046) on vessels following experimental subarachnoid hemorrhage.

In order to examine the functional changes in the vascular smooth muscle, the effects of a thromboxane A2 synthetase inhibitor (OKY-046) and a calcium channel blocker (diltiazem) on vessels following subarachnoid hemorrhage, and the contractile activity of cerebral vessels with various vasoactive agents, were investigated by studying isometric tension recordings in rings of cat basilar artery. The maximum contractile activities of the vessels in response to noradrenalin and adrenaline during the course of subarachnoid hemorrhage were significantly less than those in the control group. On the other hand, the maximum contractile activity of the vessels in response to prostaglandin F2 alpha on the seventh day following subarachnoid hemorrhage was significantly augmented compared with that in the control group. A significant decline in the relaxation of responsiveness to diltiazem during the course of subarachnoid hemorrhage was observed compared with that of diltiazem in the control group. This responsiveness to vasoactive agents was not influenced by the application of OKY-046. The present study reveals functional changes in vascular smooth muscle exposed to subarachnoid hemorrhage in response to vasoactive agents and a calcium entry blocker. Thromboxane A2 may not be a significantly influential factor in the present results. It is suggested that cerebral vasospasm may well be related to functional changes of the arterial wall, which appear to be associated with derangement of the mechanisms of smooth muscle constriction and dilatation based on organic changes.

[Meningoencephalitis in Kawasaki disease].

Kawasaki disease (KD) is a syndrome characterized by various degrees of vasculitis in small-and medium-sized arteries. We discussed the characteristic manifestations and prognosis of 5 KD patients (male 3, female 2) with meningoencephalitis in the acute stage. The incidence was 3.7% (5 of 138 patients) in our institute. The age of onset was between 3 months and 15 months. The clinical manifestations included disturbance of consciousness and seizures; disturbance of consciousness developed in all patients in the early acute stage. The duration was between 2 and 11 days; seizures developed as status convulsions in two. Electroencephalograms (EEG) demonstrated certain abnormalities in 2 of 4 patients studied. Computed-tomographic scanning (CT scan) revealed fluid collection in the frontal extracerebral space, and monocyte-predominant pleocytosis was observed in the cerebrospinal fluid (CSF) in all 5 patients studied. The age of onset in the 5 KD patients was significantly earlier than that in non-complicated cases (n = 138, P less than 0.01). Blood hemoglobin (P less than 0.05) and hematocrit (P less than 0.05), serum total protein (P less than 0.01), and serum albumin levels (P less than 0.01) were also significantly lower. Moreover the period until CRP values turned negative was significantly longer (P less than 0.05) and erythrocyte sedimentation rate was significantly lower (P less than 0.01) in these 5 patients. It was considered that meningoencephalitis in KD may develop in cases having more severe and prolonged inflammatory changes; the clinical findings revealed a serious form of KD. We suggest that this might because by vasculitis of small arteries, arterioles, capillaries, and venules, which consists of infiltration of lymphocytes and large mononuclear cells, and edema. There was no neurological sequela in 4 of the 5 patients. However, one patient was found to have hearing difficulty 3 years after the onset. Therefore, the prognosis of meningoencephalitis in KD was considered to be generally favorable.

[Antitumor effect of FU-O-G, new antitumor agent, following long term administration].

FU-O-G, the O-glucuronide methylester of 5-fluorouracil (5-FU), is a compound with a unique chemical structure. It is an antitumor agent of the prodrug type which exerts its activity by enzymatically liberating 5-FU in tumor tissues. It has been reported that the antitumor activity of this compound is superior to those of 5-FU and Tegafur (FT-207) in the treatment of various transplantable tumors. In this study, long-term administration of FU-O-G to mice bearing relatively slow-growing tumors such as Lewis lung carcinoma, mammary tumor FM3 A and hepatoma MH-134 was carried out. Consequently, FU-O-G was shown to be remarkably effective against those tumors, whereas 5-FU and FT-207 were hardly effective. Long-term daily administration was shown to be more effective than intermittent dosage in the treatment of Lewis lung carcinoma. In combination therapies of FU-O-G with other antitumor drugs, FU-O-G exhibited a synergistic effect against Lewis lung carcinoma when combined with Carboquone (CQ) or Nimustine hydrochloride (ACNU).