A case of hypereosinophilia with digital ischaemia successfully treated with intravenous prostaglandin E1.

In the therapeutic management of eosinophilic disorder, it is important to prevent hypereosinophilia (HE)-related organ damage even in the process of diagnosis. We describe here a unique clinical and histopathological findings of the patient with HE accompanied with digital ischaemia. Treatment with intravenous prostaglandin E1 was essential for digital ischaemia in our case while benralizumab, humanized monoclonal antibody against interleukin-5 receptorα, did not affect. Our case suggests an earlier intervention for digital ischaemia in the therapeutic management of eosiniphilic disorder.

Estimation of annual pollutant loadings in two small catchments and examination of their differences caused by regional properties.

A series of runoff surveys was conducted for more than one year in two small catchments of the Kamo River basin (75.4 km2) and the Takano River basin (66.8 km2) in Kyoto, Japan, which adjoin each other, and may have the same precipitation pattern. The investigation consisted of a high-frequency periodic survey, a long-term regular survey and a storm event survey. The survey results were compared with the regional properties of the basins, and the following results were obtained. (1) Pollutant loadings were successfully estimated as two portions of base discharge and storm events discharge from the survey results. (2) Estimated annual loading of the sites was 2.9-4.5, 1.3-1.8, 17-27, 1.3-2.2, 0.076-0.97 t/km2/y, respectively for COD(Mn), DOC, SS, TN and TP. (3) 52-53% of the whole flow, which was caused by rainfall events, conveyed 81-87, 68-73, 92-95, 64-67, 76-81% of the whole loading, respectively for COD(Mn), DOC, SS, TN and TP. (4) Differences of regional properties in two basins cause different runoff patterns, but the differences in runoff patterns also depend on the rainfall patterns. In general, a more urbanized basin receives early and strong influence of precipitation on the storm event runoff.

Analysis of parameter variations in L-Q equations for river runoff processes from the viewpoint of spatial and temporal conditions.

This study aims to find out variation characteristics of the parameters of 'a' and 'b' in L = a x Q(b), an empirical equation for run-off loading (L) and flow rate (Q), by evaluating the effects of flow conditions and regional properties of the watersheds on the values. We selected the Kamo River basin (155 km2) as a study field, and conducted various kinds of investigations, such as 80 day high frequency observations, continuous monitoring for more than 2 years, storm event surveys, and simultaneous surveys of 39 stations. Then, we obtained 7-170 data in each of 39 sampling stations. The main results obtained are as follows: (1) L-Q equation with a range of 'a' can express L-Q relation in most of the WQIs (water quality indices); (2) 'a' receives temporal (flow condition) effects more in SS, VSS and Al, while it receives regional effect more in inorganic carbon, TN, Ca and Fe; (3) both of flow change in storm events, and base flow levels affect the L-Q relation, and their effects can classify the WQIs into several groups; (4) the effects of regional properties were obviously observed in 'a', and quantitatively evaluated, especially for density of population.

A cold-adapted endo-arabinanase from Penicillium chrysogenum.

Previously, three arabinan-degrading enzymes were isolated from Penicillium chrysogenum 31B. Here we describe another arabinan-degrading enzyme, termed Abnc, from the culture filtrate of the same organism. Analysis of the reaction products of debranched arabinan by high-performance anion-exchange chromatography (HPAEC) revealed that Abnc cleaved the substrate in an endo manner and that the final major product was arabinotriose. The molecular mass of Abnc was estimated to be 35 kDa by SDS-PAGE. Enzyme activity of Abnc was highest at pH 6.0 to 7.0. The enzyme was stable up to 30 degrees C and showed optimum activity at 30 to 40 degrees C. Compared with a mesophilic counterpart from Aspergillus niger, Abnc exhibited a lower thermal stability and optimum enzyme activity at lower temperatures. Production of Abnc in P. chrysogenum was found to be strongly induced by arabinose-containing polymers and required a longer culture time than did other arabinanase isozymes in this strain.

The inhibition of human factor Xa by plasminogen activator inhibitor type 1 in the presence of calcium ion, and its enhancement by heparin and vitronectin.

Plasminogen activator inhibitor type 1 (PAI-1), a member of serine proteinase inhibitor superfamily, is known to inhibit thrombin in the presence of either heparin or vitronectin. We analyzed possible inhibitory activity of PAI-1 on human factor Xa. PAI-1 inhibited factor Xa in the presence of calcium ion (Ca2+), whereas no inhibition was observed in the absence of Ca2+. Half maximal enhancement by Ca2+ was obtained at 0.8 mM. An equimolar complex formation between factor Xa and PAI-1 in the presence of Ca2+ was observed by SDS polyacrylamide gel electrophoresis. Both unfractionated heparin and vitronectin enhanced the inhibition only in the presence of Ca2+. Apparent second-order rate constant (ki) for the inhibition of factor Xa by PAI-1 at 5 mM Ca2+ was 1.6 x 10(4) M-1 s-1, and was enhanced 3-fold by 2 u/ml of heparin (4.6 x 10(4) M-1 s-1) and 10-fold by 100 nM vitronectin (1.6 x 10(5) M-1 s-1), respectively. The interaction between Ca(2+)-bound factor Xa and PAI-1 could be important from the view of PAI-1 neutralization and enhancement of fibrinolysis.

Preparation and characterization of spherical polymer packings from polybutadiene for size-exclusion chromatography.

Porous polymer spherical particles for column packings in nonaqueous size-exclusion chromatography (SEC) were prepared from 1,2-syndiotactic polybutadiene by suspension and evaporation method. The polymer microbeads obtained were crosslinked by radical reaction between 2-vinyl groups in polybutadiene with ultraviolet radiation, to render them insoluble. These microbeads have wider chromatographic separation width than polystyrene column packings. In addition, the polybutadiene microbeads did not show the excessive retention observed with commercial polystyrene columns for polycyclic aromatic compounds. Therefore, a close correlation between the elution volume and M, for polycyclic aromatic compounds was observed with polybutadiene microbeads columns.

Early skin toxicity predicts better outcomes, and early tumor shrinkage predicts better response after cetuximab treatment in advanced colorectal cancer.

Cetuximab-containing treatments for metastatic colorectal cancer have been shown to have higher overall response rates and longer progression-free and overall survival than other systemic therapies. Cetuximab-related manifestations, including severe skin toxicity and early tumor shrinkage, have been shown to be predictors of response to cetuximab. We hypothesized that early skin toxicity is a predictor of response and better outcomes in patients with advanced colorectal carcinoma. We retrospectively evaluated 62 patients with colorectal adenocarcinoma who had unresectable tumors and were treated with cetuximab in our institution. Skin toxicity grade was evaluated on each treatment day. Tumor size was evaluated using computed tomography prior to treatment and 4-8 weeks after the start of treatment with cetuximab.Patients with early tumor shrinkage after starting treatment with cetuximab had a significantly higher overall response rate (P = 0.0001). Patients with early skin toxicity showed significantly longer overall survival (P = 0.0305), and patients with higher skin toxicity grades had longer progression-free survival (P = 0.0168).We have shown that early tumor shrinkage, early onset of skin toxicity, and high skin toxicity grade are predictors of treatment efficacy and/or outcome in patients with advanced colorectal carcinoma treated with cetuximab.

Tissue-type plasminogen activator is involved in the process of neuronal death induced by oxygen-glucose deprivation in culture.

Effect of tissue-type plasminogen activator (tPA) on oxygen-glucose deprivation (OGD) was studied in cultured cortical neurons prepared from tPA gene knockout (tPA-KO) and wild-type (Wt) mice. Three hours of OGD induced 45% and 23% of neuronal death in Wt and tPA-KO mice, respectively. Neuronal death in tPA-KO mice was increased to 42% by additional tPA. Six hours of OGD induced 80% and 40% of neuronal death in Wt and tPA-KO mice, respectively, whereas the addition of tPA increased to 62% in tPA-KO mice. These results suggest that tPA is directly involved in the process of neuronal death induced by ischemia-mimic stress without involving vascular or circulatory components.

Conformational change in DNA induced by cationic bilayer membranes.

The effect of synthetic cationic lipids on the structure of DNA was studied. The fluorescence enhancement of ethidium bromide on intercalation into DNA was suppressed by the addition of bilayer-forming lipids, but not by micellar ones. Results on the fluorescence depolarization index suggest that ethidium bromide is not released from DNA by lipids intercalated into DNA. CD spectra of the DNA-lipid complexes revealed that the structure of DNA was changed only by bilayer-forming lipids at temperatures lower than their Tc values. Thus, the conformation of DNA is forced to change by cationic lipids forming the rigid bilayer membrane so that ethidium bromide fluorescence might be reduced, and the conformation can be controlled by selection of the appropriate lipid and temperature.

Induction of thromboxane synthase and prostaglandin endoperoxide synthase mRNAs in human erythroleukemia cells by phorbol ester.

The effects of 12-O-tetradecanoyl-phorbol-13-acetate (TPA) on the mRNA levels of two enzymes, thromboxane synthase (TXS) and prostaglandin endoperoxide synthase (PES), responsible for the synthesis of thromboxane A2 from arachidonic acid, were studied in human erythroleukemia (HEL) cells by RNA blot analysis. TPA induced both TXS and PES mRNAs in HEL cells in a dose-dependent manner at 36 h. The half-maximal and maximal effects for the induction of both mRNAs were at approximately 3 x 10(-9) M and at 10(-8) M, respectively. TXS and PES mRNA levels increased in a time-dependent fashion by TPA, and reached to 7- and 3.5-fold of the control, respectively after 48 h of TPA treatment. These results suggest that expression of TXS and PES genes in HEL cells were simultaneously stimulated by TPA.

Characterization of a novel monoclonal antibody that senses nitric oxide-dependent activation of soluble guanylate cyclase.

Two monoclonal antibodies (mAbs) against bovine lung soluble guanylate cyclase (sGC) were prepared and characterized. mAb 3221 recognized both the alpha- and beta-subunits of sGC and had greater binding affinity to the enzyme in the presence of NO. mAb 28131 recognized only the beta-subunit and its affinity did not change with NO. Neither mAb cross-reacted with particulate GC. Cultured Purkinje cells from rats were treated with S-nitroso-N-acetylpenicillamine, an NO donor, and examined by immunocytochemical methods. The immunoreactivity associated with mAb 3221 increased with the cGMP content in a crude extract of cerebellum and the NO2 generated in the culture medium increased.

Activated protein C attenuates coagulation-associated over-expression of fibrinolytic activity by suppressing the thrombin-dependent inactivation of PAI-1.

Several activated coagulation factors have been reported to enhance fibrinolysis by neutralizing plasminogen activator inhibitor type 1 (PAI-1) activity. We evaluated the physiological relevance of this mechanism using the euglobulin clot lysis time (ECLT) assay in the presence and absence of Ca2+, which is controlled by PAI-1 and mimics physiological thrombolysis. We found that the ECLT (18.5 +/- 0.6 h) was shortened by Ca2+ (5 mm) (6.6 +/- 0.1 h). A significant difference was observed in thrombin generation by the presence of Ca2+ in the euglobulin fraction. Prothrombin was almost fully converted to thrombin within 15 min in the presence of Ca2+, whereas essentially no conversion was observed without Ca2+. The presence of activated protein C (aPC) suppressed thrombin generation, and attenuated the shortening of ECLT in a dose-dependent manner, an effect enhanced by phospholipid and protein S. In the absence of Ca2+, aPC did not prolong the ECLT. After addition of biotin-labeled recombinant PAI-1 to the euglobulin fraction, PAI-1 was cleaved to lower molecular weight forms only in the presence of Ca2+. This cleavage did not occur in the presence of aPC, suggesting that thrombin was the catalyst for PAI-1 cleavage. The cleavage and inactivation of PAI-1 by generated thrombin is proposed to be responsible for the shortening of ECLT by Ca2+ and for coagulation-associated over-expression of fibrinolysis. Under such conditions, aPC appeared to suppress thrombin generation and to normalize highly activated fibrinolysis.

Correlations between platelet aggregation, fibrinolysis, peripheral and central serotonergic measures in subhuman primates.

We have studied the relationships between whole blood and plasma serotonin (5-hydroxytryptamine, 5-HT), its major metabolite 5-hydroxyindoleacetic acid (5-HIAA) and serum lipids, platelet aggregation in the whole blood and in the platelet-rich plasma (PRP), and some fibrinolytic parameters in monkeys. Plasma 5-HT was found to be positively related to 5-HT- and ADP-induced platelet aggregation, tissue plasminogen activator (tPA) activity, serum cholesterol and LDL-cholesterol, whereas 5-HT in cerebrospinal fluid correlated inversely with serum cholesterol. Plasminogen activator inhibitor (PAI) activity was positively related to LDL. Euglobulin clot lysis time was related to both tPA and PAI activities. The significance of these findings and the possible role of 5-HT in atherogenesis and hemostasis are discussed.

The immunosuppressive mode of action of mizoribine.

Mizoribine (MIZ) suppressed the mitogen response and mixed lymphocyte reaction (MLR) significantly at doses of 100 micrograms/ml and 10 micrograms/ml in a dose-response analysis. The 50% inhibition dose (ID50) was between 10 micrograms/ml and 1.0 microgram/ml, both in the mitogen response and MLR. In a kinetic study of the MLR, the degree of suppression with MIZ at a given dosage was essentially the same as the degree of suppression observed in the dose-response analysis when MIZ was added to MLR cultures from day 0 to day 4. In addition, MLR was more susceptible to the suppressive activity of MIZ at 100 micrograms/ml when MIZ was added near the peak of lymphocyte proliferation. This experiment also showed that MLR suppression induced by MIZ at 10 micrograms/ml was reversible and MLR activity had completely recovered 6-8 hr after its removal. MIZ had no inhibitory action on MLR-derived cytotoxic cells or the effector phase of cell mediated lymphocytotoxicity. These results clearly demonstrate that MIZ suppresses lymphoproliferation, but has no effect on the recognition phase or effector phase of cytotoxic lymphocytes.

Rapid, specific and active site-catalyzed effect of tissue-plasminogen activator on hippocampus-dependent learning in mice.

In the present study we trained tissue-plasminogen activator (tPA)-knockout (tPA -/-) and wild-type (tPA +/+) male mice in step-down inhibitory avoidance learning, a hippocampus-dependent task. tPA -/- displayed significantly shorter latencies to step down at 90 min, one, two and seven days after training indicating the learning deficit in these animals (P < 0.05 vs tPA +/+). The locomotor activity, the level of anxiety in an elevated-plus maze, as well as the pain threshold did not differ between the two strains of mice. The learning disability of tPA -/- was overcome by more intense training. The learning deficit was also partially restored by limited intrahippocampal delivery of tPA (infused for 2 h before training; P < 0.05 vs control), but not by the delivery of urokinase plasminogen activator, indicating the acute need for tPA in learning. The beneficial effect of tPA was abolished by co-infusion of its inhibitor tPA-STOP, indicating that the facilitatory effect of tPA on learning requires a proteolytic step. However, tPA activity in the hippocampus was not indispensable for effective memory retrieval in tPA-infused tPA -/- mice. Thus, rapid, specific and proteolytic action of tPA facilitates hippocampus-dependent learning, but not retrieval of previously acquired information.

Human thrombin and calcium bound factor Xa significantly shorten tPA-induced fibrin clot lysis time via neutralization of plasminogen activator inhibitor type 1 activity.

Employing a fibrin clot lysis assay, we reassessed the significance of the neutralization of plasminogen activator inhibitor type 1 (PAI-1) activity by thrombin and factor-Xa in fibrinolysis. When PAI-1 enriched fibrin clots were formed using increasing concentrations of thrombin (0.1, 0.5, 1.0 IU/ml), their lysis times became shorter (43.8 +/- 4.9, 25.7 +/- 3.7, 13.9 +/- 0.8 h respectively). Times were shortened further by either heparin (43.9 +/- 11.0, 12.1 +/- 2.6, 3.6 +/- 0.2 h respectively) or vitronectin (17.0 +/- 1.6, 1.9 +/- 0.7, 0.9 +/- 0.0 h respectively). Factor-Xa together with Ca++ shortened the clot lysis time further. Fibrin autography revealed that both enzymes dose dependently interfered with complex formation between tPA and PAI-1, making large amounts of tPA remaining to be free form. The mechanism seems to play a role in the coagulation associated enhancement of fibrinolysis.

Complete nucleotide sequence of a thermophilic alpha-amylase gene: homology between prokaryotic and eukaryotic alpha-amylases at the active sites.

The nucleotide sequence of a thermophilic, liquefying alpha-amylase gene cloned from B. stearothermophilus was determined. The NH2-terminal amino acid sequence analysis of the B. stearothermophilus alpha-amylase confirmed that the reading frame of the gene consisted of 1,644 base pairs (548 amino acids). The B. stearothermophilus alpha-amylase had a signal sequence of 34 amino acids, which was cleaved at exactly the same site in E. coli. The mature enzyme contained two cysteine residues, which might play an important role in maintenance of a stable protein conformation. Comparison of the amino acid sequence inferred from the B. stearothermophilus alpha-amylase gene with those inferred from other bacterial liquefying alpha-amylase genes and with the amino acid sequences of eukaryotic alpha-amylases showed three homologous sequences in the enzymatically functional regions.

Kinetic basis for the donor nucleotide-sugar specificity of beta1, 4-N-acetylglucosaminyltransferase III.

The kinetic basis of the donor substrate specificity of beta1, 4-N-acetylglucosaminyltransferase III (GnT-III) was investigated using a purified recombinant enzyme. The enzyme also transfers GalNAc and Glc moieties from their respective UDP-sugars to an acceptor at rates of 0.1-0.2% of that for GlcNAc, but Gal is not transferred at a detectable rate. Kinetic analyses revealed that these inefficient transfers, which are associated with the specificity of the enzyme, are due to the much lower V(max) values, whereas the K(m) values for UDP-GalNAc and UDP-Glc differ only slightly from that for UDP-GlcNAc. It was also found that various other nucleotide-Glc derivatives bind to the enzyme with comparable affinities to those of UDP-GlcNAc and UDP-Glc, although the derivatives do not serve as glycosyl donors. Thus, GnT-III does not appear to distinguish UDP-GlcNAc from other structurally similar nucleotide-sugars by specific binding in the ground state. These findings suggest that the specificity of GnT-III toward the nucleotide-sugar is determined during the catalytic process. This type of specificity may be efficient in preventing a possible mistransfer when other nucleotide-sugars are present in excess over the true donor.

Changes in the central and peripheral serotonergic system in rats exposed to water-immersion restrained stress and nicotine administration.

The effects of water-immersion restraint stress (WS) on chronically nicotine-administered rats were studied in the blood and various regions of the brain. Serotonin (5-HT) levels increased in the hypothalamus, hippocampus, cortex and cerebellum following the administration of nicotine. 5-HT levels increased in all the brain regions following stress. Nicotine decreased stress-induced increased levels of 5-HT in the hippocampus and cerebellum. Nicotine administration alone increased 5-hydroxyindole acetic acid (5-HIAA) levels in the hippocampus and cerebellum. Stress alone also increased 5-HIAA levels in all the brain regions. In the cortex, 5-HT and 5-HIAA levels further increased following the administration of a combination of stress and nicotine compared to rats given stress alone. In the blood as well as in all the brain regions, except the cerebellum, stress or nicotine administration did not affect tryptophan levels. Stress given to nicotine-administered rats resulted in a decrease in tryptophan levels in the blood and plasma. Although 5-HT and 5-HIAA levels were not influenced by stress and/or nicotine administration, the 5-HIAA/5-HT ratio increased in the blood and plasma of rats administered with nicotine and exposed to stress. The effects of nicotine on the serotonergic system depend upon the kind of stress given together with the organs and brain regions involved.

Predictability of renal allograft prognosis during rejection crisis by ultrasonic Doppler flow technique.

Using the ultrasonic Doppler technique, renal blood flow was measured in 67 patients who underwent living related renal transplantation from January, 1976 to December, 1979. In 58 of 67 cases, 81 acute and 9 chronic rejection episodes occurred. In the initial stage of acute rejection, there are no particular changes in the pattern of systolic blood flow and by contrast marked changes of diastolic flow. The disappearance of the diastolic phase is indicative of an advanced stage of rejection, the reappearance indicative of recovery from rejection, and persistent loss accompanied by changes of systolic flow indicative of an unfavorable prognosis of rejection. In chronic rejection, there are rapid changes of neither systolic nor diastolic flow though the acceleration time in the systolic phase lengthens gradually. The ultrasonic Doppler flow technique for blood flowmetry of a transplanted kidney is a useful means of knowing the prognosis of rejection and provides an index for corticosteroid bolus therapy.