Ulcerative colitis complicated by gastroduodenal lesions

The glycosylated multivalent three-domain Kunitz inhibitor TFPI is a natural inhibitor of tissue factor-FVIIa complex in the presence of FXa. TFPI has an experimental antithrombotic capacity indistinguishable from LMWH in a prophylactic dose, regardless of glycosylation and of the third domain. An inherited equilibrium between antithrombosis and haemorrhage exists. The aim of the study was to evaluate whether a two-domain non-glycosylated TFPI (117QTFPI1-161) has a bleeding potential in a rat gastric mucosa model. Groups; placebo, LMWH (tinzaparin) 60 and 250 anti-Xa IU/kg and 117 QTFPI1-161 1.0 and 10.0 mg/kg, given i.v. (bolus injection), randomised double dummy design. All actively treated groups significantly prolonged both the bleeding volume (493-984 microliters) and the bleeding time (10-20 min) compared to placebo (41 microliters, 2 min). It was not possible to distinguish a difference between the lower dose of LMWH and 117QTFPI1-161 in either parameter (p = 0.23-0.71). The two doses of 117QTFPI1-161 caused elevation of plasma-TFPI, 18 and 150 times baseline value. Both LMWH doses (0.6-3.2 anti-Xa IU/ml) and both 117QTFPI1-161 doses (0.2-2.7 anti-Xa IU/ml), caused significant effect in the anti-Xa assay, however 117QTFPI1-161 significantly less. Only the largest dose of 117QTFPI1-161 caused significant prolongation in the APTT assay (34 s). Both doses of LMWH caused significant prolongation (60-300 s). LMWH was the only substance to prolong the dilute-PT assay. Non-glycosylated two-domain 1.0 mg/kg TFPI, yielding supraphysiological plasma concentration, has an experimental haemorrhagic potential indistinguishable from LMWH in a prophylactic dose. The effect mediated by this type of TFPI could primarily be due to an inhibition of FXa.     

Crystal structure of NMC-4 fab anti-von Willebrand factor A1 domain

We have solved the crystal structure of the Fab fragment of NMC-4, a mouse monoclonal antibody that binds to the A1 domain of von Willebrand factor (vWF). Two Asp and three Tyr residues in the complementarity determining regions 1 and 3 of the heavy chain exhibited a spatial orientation suggestive of a dominant role in establishing contact with the antigen. A cluster of Asp and Tyr residues occurs also in a region of the platelet glycoprotein (GP) Ib alpha amino terminal domain known to be critically involved in vWF binding. Thus, the structural information obtained with NMC-4 may prove relevant to understand the stereochemical bases of the GP Ib alpha-vWF interaction essential for thrombus formation at sites of vascular lesion.     

Simultaneous analysis of 2-aminopyridine-derivatized neutral and sialylated oligosaccharides from human serum in the negative-ion mode by sonic spray ionization ion trap mass spectrometry

Neutral and acidic (sialylated) 2-aminopyridine-derivatized (PA) oligosaccharides were analyzed by using reversed-phase high-performance liquid chromatography/ion trap mass spectrometry (RP-HPLC/IT MS) with a sonic spray ionization (SSI) source. Under the RP-HPLC separation using a buffer of 1 mM ammonium acetate (pH4.3) at a flow rate of 0.2 mL/min, both PA-oligosaccharides in the negative-ion mode showed a comparable degree of ionization efficiency, differing from that of the positive-ion mode, which exhibits a wide gap between their ionization efficiencies. In addition, the ion intensities of both PA-oligosaccharides were higher in the negative-ion mode than in the positive-ion mode. These results strongly suggest that the negative-ion mode of SSI-MS is suitable for simultaneous analysis of neutral and acidic (sialylated) oligosaccharides in RP-HPLC/MS. In the present study, RP-HPLC/SSI-IT MS in the negative-ion mode was used in the analysis of PA-oligosaccharides from human serum and its usefulness was investigated. As a result, 32 neutral and sialylated PA-oligosaccharides from human serum were identified with differentiating isomeric oligosaccharides and relatively quantified by a single HPLC/MS run. This method is useful for simple and rapid analysis of the overall distribution of neutral and sialylated oligosaccharides in a complex sample such as serum.     

A novel glycerol kinase from Flavobacterium meningosepticum: characterization, gene cloning and primary structure

A thermostable glycerol kinase (FGK) was purified 34-fold to homogeneity from Flavobacterium meningosepticum. The molecular masses of the enzyme were 200 kDa by gel filtration and 50 kDa by SDS-PAGE. The Km for glycerol and ATP were 0.088 and 0.030 mM, respectively. The enzyme was stable at 65 degrees C for 10 min and at 37 degrees C for two weeks. The enzyme gene was cloned into Escherichia coli and its complete DNA was sequenced. The FGK gene consists of an open reading frame of 1494-bp encoding a protein of 498 amino acids. The deduced amino acid sequence of the gene had 40-60% similarity to those of glycerol kinases from other origins and the amino acid sequence of the putative active site residue reported for E. coli GK is identical to the corresponding sequence of FGK except for one amino acid residue.     

Neoadjuvant intraarterial chemotherapy for ten cases of inflammatory breast cancer by Seldinger's methods

The conventional methods (CM) of intraarterial (IA) chemotherapy for inflammatory breast cancer were catheterizations from superior epigastric and brachial artery under general anesthesia. Since 1997, we selected the Seldinger's methods (SM) for ten cases of the disease to control local effects and simplify the technique. Complications of the SM were slight, and side effects were equal to those with CM. The postoperative pathological findings of the SM showed the direct effects of chemotherapy to tumor cells (degenerative and necrotic changes) as compared with the embolism-like effects of CM. But the overall histological effects of chemotherapy by SM were almost equal to those for CM. The strong points of the SM were as follows: 1) More selective IA chemotherapy is available, 2) one can find the passage of the aim vessels and no trouble related to catheter, 3) general anesthesia is not necessary, and the techniques are simple, 4) the wounds are not remarkable. The disadvantages are as follows: 1) Patients must rest the day of IA chemotherapy, 2) in 20% of the procedures, one can not search the vessels, and 3) in 4 cases complications of stiffness of Mj or Mn pectral muscles were found. In future, we expect more effective results by dose escalation or combination chemotherapy.