Diatoms and the ecological conditions of their growth in sea ice in the arctic ocean.

A summer field survey off Point Barrow, Alaska, revealed that Arctic sea ice develops a growth of phytoplanktonic diatoms. The diatoms are found in a brine solution in microfissures between ice crystals on the underside of the ice. The chlorophyll content of this layer is 100 times more than that of the surrounding sea waters; this has led to a hypothesis that a considerable fraction of the primary production of the Arctic Sea may be carried out in sea ice, especially during the spring and early summer months.

Evaluation of ethanol washing on dioxins-polluted soil and sediment based on adsorption relationships.

Multi-stage ethanol washing on dioxins-polluted soil and sediment were performed. The results indicated the existence of limit washing concentration (LWC), where no more dioxins were removed from the soil or the sediment by further washing. In each stage, dioxins concentration in the soil, sediment and ethanol could be described satisfactory by the Freundlich equation. The Freundlich capacity factor, K(ef) correlated with the LWC which was estimated to be ca. 1000 pmol g(-1) in the case of soil, and about 150 pmol g(-1) in the case of sediment. Organic contents in the soil and sediment affected the Freundlich intensity parameter, n(-1) but not K(f). A model, which enables the calculation of removal efficiency of PCDD/DFs at each stage using K(f), n, and initial PCDD/DFs concentration, is presented.

High expression of ABCG2 induced by EZH2 disruption has pivotal roles in MDS pathogenesis.

Both proto-oncogenic and tumor-suppressive functions have been reported for enhancer of zeste homolog 2 (EZH2). To investigate the effects of its inactivation, a mutant EZH2 lacking its catalytic domain was prepared (EZH2-dSET). In a mouse bone marrow transplant model, EZH2-dSET expression in bone marrow cells induced a myelodysplastic syndrome (MDS)-like disease in transplanted mice. Analysis of these mice identified Abcg2 as a direct target of EZH2. Intriguingly, Abcg2 expression alone induced the same disease in the transplanted mice, where stemness genes were enriched. Interestingly, ABCG2 expression is specifically high in MDS patients. The present results indicate that ABCG2 de-repression induced by EZH2 mutations have crucial roles in MDS pathogenesis.

A convenient evaluation of the stereoselectivity of lipase-catalyzed hydrolysis of tri-O-acylglycerols on a chiral-phase liquid chromatography.

A general method was developed using chiral-phase chromatography in order to evaluate the stereoselectivities of lipases-catalyzed hydrolysis of tri-O-acylglycerols independent of acyl groups. 1,2-Di-O-acyl-sn-glycerols or its enantiomer 2,3-di-O-acyl-sn-glycerols in the enzymatic reaction mixtures were derivatized to the key compound, 1,2-di-O-benzoyl-3-O-tert-butyldimethylsilyl-sn-glycerol 2 (+) or its enantiomer 2' (-), respectively. The enantiomers were separated on a chiral-phase HPLC, and the method was highly sensitive to determine the stereoselectivities of lipases.

Determination of the lipase stereoselectivities using circular dichroism (CD); lipases produce chiral di-O-acylglycerols from achiral tri-O-acylglycerols.

A general method was described to determine the optical purity of 1,2 (or 2,3)-di-O-acylglycerols via a key compound, 3 (or dibenzoyl-sn-glycerol (3 or 3'). The chiral di-O-acylglycerols were first silylated and the acyl groups were removed by the Grignard degradation to 3 (or 1) O-tert-butyldimethylsilyl-sn-glycerol and subsequent benzoylation lead to the key compound 3 or 3' without racemization. The optical purity was determined from the strong exciton Cotton effect of 3 (+) or 3' (-) at 238 nm in the concentration of ca. 1 mM. The method was successfully applied to determine the stereoselectivities of lipases (EC 3.1.1.3) from three origins, bacteria, mammal and fungus such as Pseudomonas (AP, 89% optical purity, sn-1 preference), porcine pancreatin (PPL, 9.3% optical purity, sn-3 preference) and Candida (CC, sn-2 preference) using tripalmitin. The similar studies were extended to tri-O-benzoylglycerol (6) and tri-O-(cyclohexanecarbonyl)glycerol (5). All the enzymes showed high stereoselectivities with tri-O-benzoylglycerol. PPL and AP showed high and low stereoselectivities with tri-O-(cyclohexanecarbonyl)glycerol, while low and high stereoselectivities with tri-O-palmitoylglycerol, respectively. The results show that the stereoselectivities are ruled by the origins of lipases and acyl groups. The structures of the recognition site might be associated with enantioselectivities of the enzymes.

Colonic carcinoma resembling submucosal tumor: report of a case and review of the literature.

Submucosal tumor-like colorectal carcinoma, most of whose surface is covered with normal mucosa, is very rare. We report a case of colonic carcinoma resembling submucosal tumor. A 54-yr-old man visited our institution for an evaluation of a positive fecal occult blood test. Colonoscopic examination revealed a small, mainly red polypoid lesion with a central deep ulceration and many white spots in the sigmoid colon. Indigocarmine staining demonstrated that the white spots were faint shallow depressions. Magnifying colonoscopic examination showed that the lesion surface, except for the ulceration and the depressions, was covered with normal mucosa. Although the tumor was small, we strongly suspected its malignancy due to a deep ulceration. As we could not excise it endoscopically, we performed sigmoidectomy. The lesion was 12 mm in size. Histologic examination revealed that the lesion was a moderately differentiated adenocarcinoma that was mainly covered with normal mucosa, that carcinoma was exposed only at the ulceration and the depressions on the surface, and that it had expanded to the muscularis propria. Together with considerations from the literature, this type of colorectal carcinoma is supposed to be invasive and surgical resection should be considered, no matter how small it may be.

Two forms of expression and genomic structure of the human heterogeneous nuclear ribonucleoprotein D-like JKTBP gene (HNRPDL).

The human DNA- and RNA-binding protein JKTBP is a member of a 2xRNA-binding domain (RBD)-glycine family of heterogeneous nuclear ribonucleoproteins that are involved in mRNA biogenesis. Northern and Western blottings revealed that mRNAs of approx. 1.4 and 2.8kb and proteins of approx. 38 and 53kDa were present in HL-60 cells and various tissues. Cloning and characterization of a previously unknown cDNA for the 2.8kb mRNA indicated that the cDNA encodes a 420 amino acid JKTBP polypeptide. Isolation and characterization of the genomic DNA showed that the gene (HNRPDL) had nine exons and had two separate transcription start sites for the two transcripts. The features of the 5' flanking sequences of these sites showed that the gene is a housekeeping gene. Fluorescence in situ hybridization mapped the gene to 4q13-q21. From its gene organization, the JKTBP seems to be most closely related to hnRNP D/AUF1.

Functional expression from cloned cDNAs of glutamate receptor species responsive to kainate and quisqualate.

The complete amino acid sequences of two mouse glutamate receptor subunits (GluR1 and GluR2) have been deduced by cloning and sequencing the cDNAs. Xenopus oocytes injected with mRNA derived from the GluR1 cDNA exhibit current responses both to kainate and to quisqualate as well as to glutamate, whereas oocytes injected with mRNA derived from the GluR2 cDNA show little response. Injection of oocytes with both the mRNAs produces current responses larger than those induced by the GluR1-specific mRNA and the dose-response relations indicate a positively cooperative interaction between the two subunits. These results suggest that kainate and quisqualate can activate a common glutamate receptor subtype and that glutamate-gated ionic channels are hetero-oligomers of different subunits.

Establishment of feeder-independent cloned caprine trophoblast cell line which expresses placental lactogen and interferon tau.

A feeder-independent cloned trophoblast cell line, HTS-1, was established from a mature placenta of Shiba goat (Capra hircus). During the growth phase, single HTS-1 cells exhibited ruffled membranes or lamellipodia often accompanied by elongated cell shape, indicating highly motile nature of the cells. At or near confluence, HTS-1 cells formed monolayers with few sign of cellular overlapping. Binucleate cells were found at a high frequency especially in the peripheral regions of monolayers. In small colonies and the monolayers, majority of HTS-1 cells assumed polygonally shaped cobble-stone like morphology characteristic to epithelial cells, although considerable variations in cellular morphology were observed despite of repeated cloning. Time-lapse video recordings of HTS-1 cells during culture revealed that not only the small colonies but also the monolayers near or at confluence were remarkably motile, often causing extreme elongation of the cells within them. The extremely plastic nature of HTS-1 cells in vitro is likely to be the reflection of the extraordinary capacity of caprine trophoblast cells to be stretched to extreme thinness in vivo as shown by electron microscopy. HTS-1 cells cultured on matrigel are highly invasive, and express MT1-MMP which, in the mouse, has been known to be expressed at the invasive edge of trophoblast both in vitro and in vivo. HTS-1 cells express placental lactogen (PL) and interferon-tau (IFNtau), as confirmed by immunocytochemistry, Western blotting and RT-PCR analysis. Both PL and IFNtau expression in the cells appeared to be down-regulated by cell-cell contact. In the medium conditioned by HTS-1 cells, the presence of secretory form of PL and IFNtau was confirmed by Western blotting. The HTS-1 cell line will serve as a useful in vitro model for the analysis of the molecular and/or cellular mechanisms underlying synepitheliochorial placentation in bovidae animals.

Cloning and functional expression of a cDNA encoding the mouse beta 2 subunit of the kainate-selective glutamate receptor channel.

The primary structure of the mouse glutamate receptor beta 2 subunit has been deduced by cloning and sequencing cDNA. The beta 2 subunit has structural characteristics common to the subunits of glutamate-gated ion channels. Expression of the cloned cDNA in Xenopus oocytes yields functional glutamate receptor channels selective for kainate.

Investigation of the heterogeneity of heterogalactan from the fruit bodies of Fomitopsis pinicola, by employing concanavalin A-Sepharose affinity chromatography.

A heterogalactan was isolated from the hot water extract of fruit bodies of Fomitopsis pinicola by a combination of fractionation procedures including precipitation with ethanol and with Cetavlon, and chromatography on columns of DEAE-cellulose and Sephadex G-100. Despite its apparent homogeneity on gel filtration, zone electrophoresis, sedimentation equilibration, and immunodiffusion analyses, the neutral component of heterogalactan was further fractionated into unbound, weakly bound, and strongly bound forms by affinity chromatography on a column of concanavalin A-Sepharose CL 4B. The former two polysaccharides fractions eluted with 0.1 M phosphate buffer (pH 7.0) were found to be a fucogalactan and a mannofucogalactan, respectively. A more tightly bound fraction (mannofucogalactan) was subsequently eluted with 0.1 M glucose in 1 M NaCl. The results of methylation, complete Smith degradation, and proton and 13C NMR spectroscopic analyses indicated that the three kinds of heterogalactans are all highly branched polysaccharides containing a framework of (1 leads to 6)-linked alpha-D-galactopyranosyl residues, the C-2 positions of which are substituted in different proportions with either single L-fucopyranosyl residues or disaccharide units of 3-O-alpha-D-mannopyranosyl-L-fucopyranose residues.

Fluorimetric determination of diarrhetic shellfish toxins in scallops and mussels by high-performance liquid chromatography.

The fluorimetric determination of okadaic acid (OA) and dinophysistoxin-1 (DTX-1), the principal toxins of diarrhetic poisoning, is reported. The digestive glands of mussels or scallops were homogenized with 2-propanol. OA and DTX-1 were extracted from the homogenate, with hexane-ethyl acetate and labelled with 2,3-(anthracenedicarboximido)ethyl trifluoromethanesulfonate in dry acetonitrile. After cleaning up by passage through a short silica gel column, the fluorescent derivatives were determined by HPLC. The derivatives were at first separated on a Develosil Ph-5 column, and only the target fraction obtained was introduced into a Develosil ODS K-5 column by a valve-switching device. Both toxins were determined in the range 2.5-500 pg, and the detection limits were 0.8 pg (OA) and 1.3 pg (DTX-1) with a signal-to-noise ratio of 3.

No association between MTHFR gene polymorphism and diabetic nephropathy in Japanese type II diabetic patients with proliferative diabetic retinopathy.

The development of diabetic nephropathy shows marked variation among individuals. Not only hyperglycemia, but also genetic factors may contribute to the development of diabetic nephropathy. Methylenetetrahydrofolate reductase (MTHFR) is involved in remethylation of homocysteine to methionine. Decreased activity of MTHFR which can result in hyperhomocysteinemia may lead to cerebrovascular disease and coronary artery disease. Recently, a common C to T mutation at nucleotide position 677 of the MTHFR gene (MTHFR677CT) has been reported to be correlated with hyperhomocysteinemia and the severity of coronary artery disease as macroangiopathy. In the present study, we recruited 173 of Japanese type II diabetic patients with proliferative diabetic retinopathy who would be exposed to long-term hyperglycemia, and examined the contribution of the MTHFR gene polymorphism to the development of diabetic nephropathy as microangiopathy. The frequency of the mutated allele was 43.3% in patients with nephropathy (n = 105) versus 41.9% in those without nephropathy (n = 68). The genotype frequencies were +/+, 16.2%; +/-, 54.3%; -/-, 29.5% in patients with nephropathy versus +/+, 13.2%; +/-, 57.4%; -/-, 29.4% in those without nephropathy (+ indicates the presence of the mutation). The MTHFR genotype and allele frequencies were not significantly different between patients with and without nephropathy. Therefore, we conclude that the MTHFR gene polymorphism is not associated with the development of diabetic nephropathy in Japanese type II diabetic patients.

Fluorometric determination of total and bound sulfite in wine by N-(9-acridinyl)maleimide.

N-(9-Acridinyl)maleimide (NAM) reacts with sulfite in wine and gives strong fluorescent derivatives that lead to highly sensitive fluorometry of both total and bound sulfite in wine. Values of free and bound sulfite in wine determined by the NAM method and the modified Rankine method agreed. Sulfite was determined in < 200 microL wine within 2 h.

[Prevalence of penicillin-insensitive Streptococcus pneumoniae and its response to oral beta lactam antibiotics at a primary pediatric office].

Penicillin (PC) resistance of Streptococcus pneumoniae was tested by oxacillin disk method (Bauer-Kirby method) of the strains collected at the primary pediatric office. The rate of oxacillin resistance of S. pneumoniae was 36.4% in 1990, 41.4% in 1991, and 51.9% in 1992, respectively. The efficacy of oral antibiotics in the treatment of PC-insensitive S. pneumoniae infections was also studied retrospectively in 234 cases. Treatment failure rate was 17.7% in the amoxicillin group, 8.7% in the cefpodoxime proxetil group, while it was 42.9% in the cefixime group. These differences were statistically significant. From these data prevalence of PC-insensitive S. pneumoniae is very high in Japanese children, and amoxicillin and cefpodoxime proxetil can be used for the treatment of outpatients with PC-insensitive S. pneumoniae infections.

[Therapeutic evaluation of combination therapy using C-425, human native immunoglobulin liquid preparation for i.v. administration, and antibiotics in severe and/or refractory infections in pediatrics].

A newly developed human immunoglobulin liquid preparation for intravenous injection was studied for efficacy, safety, and usefulness in treating severe and/or refractory infections in children receiving antibiotic treatment. It is suggested that C-425 is a useful intravenous preparation of human immunoglobulin for the treatment of severe and/or refractory infections in pediatrics. C-425 was administered to 87 inpatients with severe and/or refractory infections at 23 institutions nationwide. The Committee selected 61 cases for the present analysis. Physicians in charge judged clinical efficacy of C-425 to be "excellent" in 23 cases (40.4%), "good" in 24 (42.1%), "fair" in 7 (12.3%), "poor" in 3 (5.3%), and "unknown" in 4. The efficacy rate was calculated at 82.5% when the "excellent" and "good" cases were combined, and 94.7% when the "fair" cases were also included. According to the Committee's judgement, the efficacy of C-425 was "excellent" in 27 cases (44.3%), "good" in 18 (29.5%), "fair" in 7 (11.5%), and "poor" in 9 (14.8%). The efficacy rate was 73.8% when the "excellent" and "good" cases were combined. The rate increased to 85.2% when the "fair" cases were added. Organisms were identified in 31 cases, and the time course was followed in 19 instances. Organisms were eliminated in 12 cases (63.2%), decreased in number in 2 (10.5%), and persisted in 5 (26.3%). Eradication rate was 63.2%. One of the 87 patients died of fulminant hepatitis 2 days after the end of the treatment. The remaining 86 cases were analyzed for the safety of C-425. A skin rash was observed in one case. Laboratory examination revealed increase in transaminase levels in a total of 8 cases; both in GOT and GPT in 5, in GOT alone in 2, and in GPT alone in 1. These findings were not clinically important.

[Influence of dexamethasone on the clinical course of bacterial meningitis in children. Especially on secondary fever. Experiences in 27 institutions].

Of pediatric patients with purulent meningitis seen at the institutions listed in the title page of this paper between 1986 and 1994, 93 patients treated with antibiotics and dexamethasone (DXM) were compared with 91 patients treated with antibiotics alone. The patients receiving antibiotics with dexamethasone achieved overall improvement in inflammatory symptoms and signs and cerebrospinal fluid findings and became afebrile significantly earlier than those receiving antibiotics alone. However, some of the patients became febrile again. The secondary fever rate for the DXM group was much higher than that for the antibiotic alone group (p < 0.0001). In most of the rebounded cases, the body temperature rose above 38 degrees C and remained elevated for 2-4 days. Cerebrospinal fluid (CSF) was cultured daily in 54 and 32 patients receiving antibiotics with and without DXM, respectively. Although this study was not a controlled study in a strict sense, these patients compared. In both groups, the CSF became mostly culture-negative within 48 hours. In a few patients receiving DXM, however, it became culture-negative after 72 hours or longer. DXM caused an adverse effect in a patient with meningitis caused by Streptococcus pneumoniae. The adverse effect was mild gastrointestinal bleeding, which recovered spontaneously. From the findings described above, the use of DXM combined with antibiotic therapy was considered to accelerate the relief from fever and improvement of inflammatory symptoms and signs and CSF findings. The body temperature rose again in more than half of the patients receiving DXM, but fell to normal spontaneously without treatment. The elevation doubtlessly could not be distinguished from recurrence of the meningitis itself or complications. It seems to be likely that no treatment but careful observation is required even if the fever recurs as far as the CSF findings showed favorable progress with excelluent general conditions. When DXM is given, it is essential that CSF tests and culture are repeated during the early stages and the progress is monitored carefully.

[Trend of bacterial meningitis in children over a 14 year period (1981 through 1994) in Japan--an analysis based on studies in 27 institutions].

We observed 266 children with purulent meningitis in 27 institutions in Japan during the 14 years from 1981 on dividing these years into 3 periods, 1981-1985, 1986-1990 and 1991-1994, and studied the trend of causative organisms identified in 254 among the 266 patients. Their ages were less than 3 months after birth in 50 children and 3 months or older in 216: there were 141 boys and 125 girls. The causative organisms were H. influenzae in 134 patients and S. pneumoniae in 50, most of them being aged 3 months or older. Next to the above bacteria ranked S. agalactiae in 29 and E. coli in 12, many of the patients were aged less than 3 months. Staphylococcus spp. was found in 7 patients and about 70% of them were aged 3 months or older. L. monocytogenes was found in 4 patients and N. meningitidis in 3 and they were aged 3 months or older in both patient groups. S. pyogenes, Enterococcus spp., Peptostreptococcus spp., P. Mirabilis and Enterobacter spp. were detected each in 1 patient. The causative organism was unknown in 21 patients and there was no double infection. H. influenzae were detected in 18 patients in 1981-1985 period (36.7%), in 56 in 1986-1990 (54.9%) and in 60 in 1991-1994 (63.8%) showing an increasing tendency, but S. pneumoniae exhibited neither an increasing nor decreasing tendency. There was a decreasing tendency with S. agalactiae and E. coli, but the details were not clear because there were few patients aged less than 3 months. Although the period of coexistence of 4 main bacterial species was not made clear in this study. Listeria is considered to develop mainly in the early childhood, and we believe that the conventional way of using a cephem preparation and ampicillin combined for patients under 6 years need not be altered. However, panipenem (phonetic) is likely to be effective for insensible S. pneumoniae for the time being.

[Clinical evaluation of a new macrolide antibiotic, azithromycin, in the pediatric field].

Azithromycin (AZM) preparations in fine granules and capsules were evaluated in 36 pediatric patients with various infections. In patients with pneumonia caused by Moraxella catarrhalis, Haemophilus influenzae or Mycoplasma pneumoniae, bronchitis, pharyngitis, scarlet fever, whooping cough, or campylobacter enteritis, AZM was found effective in 94.4% (34/36). As for the bacteriological efficacy of AZM, all of 12 strains identified were found eradicated by the treatment. Plasma T 1/2(24 approximately 48 hrs.) of AZM in fine granules, given two patients at 10 mg/kg body weight once daily for 3 days, were 41.5 and 51.4 hours, while AUC0 approximately infinity was 7.45 and 13.44 mg.hr/ml. The rates of AZM recovered in the urine samples from two pediatrics patients in the first 81 hours of treatment, when it is given in fine granules at 10 mg/kg body weight once daily for 3 days, were 6.27% and 11.0%. Data from 43 patients were included for drug safety evaluation. Neither adverse reactions nor abnormal laboratory changes were observed. In conclusion, AZM was found useful in treatment of pediatric infections.

[Clinical evaluation of cefozopran in the pediatric field].

A new parenteral cephem, cefozopran (CZOP), was evaluated for its safety and efficacy in 21 children with acute infections. A mild side effect or an abnormality in laboratory tests was observed in one case each, but the safety of CZOP was otherwise observed. CZOP was effective in all of the 18 bacterial infections tested including pneumonia, cellulitis and urinary tract infections, and all the causative organisms were eradicated. Serum half-lives of CZOP were 1.5-1.8 hours and the urinary excretion rates were 63-96% in the first 5-8 hours after administration. These data suggest that CZOP is safe and effective in children with susceptible bacterial infections.