Deamination and reductive deamination of (2-amino-5, 6-dimethylbenzimidazolyl)cobamide. Preparation of (2-hydroxy-5, 6-dimethylbenzimidazolyl)cobamide and (5, 6-dimethyl-[2(-14)C]-benzimidazolyl)cobamide

(2-Amino-5, 6-dimethylbenzimidazolyl)-cobamide (III) is transformed to (2-hydroxy-5, 6-dimethylbenzimidazolyl) cobamide (IV) by nitrous acid. Exchange of the NH2-group by hydrogen with nitrous acid/hypophosphorous acid yields vitamin B12 (I). This reaction completes a cycle vitamin B12 (I)----[carboxy(2-cyanoamino-4,5-dimethylphenyl)amino]cobamide+ ++ (II)----(2-amino-5,6-dimethylbenzimidazolyl)cobamide (III)----vitamin B12 (I), which allows chemical 14C-labelling of vitamin B12. In this procedure cyanogen bromide, which is necessary for the first step, was labelled with [14C] cyanide. By the following reactions a vitamin B12 was formed in which C-2 of the 5, 6-dimethylbenzimidazole moiety is labelled.     

Activity of β-galactosidase in homogenates and isolated microvilli fraction of jejunal mucosa from suckling rats

1. beta-Galactosidase activity was studied in homogenates and isolated microvilli fraction of jejunal mucosa from 14-day-old suckling rats. o-Nitrophenyl beta-d-galactoside served as the substrate. 2. The microvilli fraction contains about one-third of the total activity of the original homogenate. 3. The pH optimum of the beta-galactosidase was 3.5 in the total homogenate and supernatant fraction, whereas in the microvilli fraction the maximum activity was at pH5.5. 4. This work gives further support to the view that two beta-galactosidases exist in the jejunal mucosa.     

Endovascular brachytherapy of transjugular intrahepatic portosystemic shunt

Purpose: To evaluate the technical feasibility and efficacy of endovascular brachytherapy with Iridium-192 in the prevention of restenosis caused by neointimal hyperplasia of transjugular intrahepatic portosystemic shunt (TIPS).Materials and Methods: The endovascular brachytherapy with high dose rate automatic afterloading system was performed in six patients with recurrent of stenosis of TIPS. We used a single dose fraction of 12 Gy delivered at 3 millimeter (mm) from the source axis to the stenotic vessel segment in five patients with spiral Z-stent, and 15 Gy at 5 mm in one patient with Wallstent.Results: Follow-up time ranged from 148 to 639 days. In one patient, restenosis occurred in the treated vessel segment, diagnosed 71 days after endovascular brachytherapy by doppler ultrasound. All other patients were, during the follow-up time, without restenosis in the irradiated vessel segment. Radiation-associated side effects were not observed.Conclusions: Endovascular brachytherapy of TIPS is technically feasible and may be done as a part of the percutaneous revision of the shunt. This pilot study may be the largest experience of treating TIPS restenosis in humans to date. For definitive conclusions, a lot of studies are needed.     

Antibody labeling with copper-67 using the bifunctional macrocycle 4-[(1,4,8,11-tetraazacyclotetradec-1-yl)methyl]benzoic acid.

The high kinetic stability of the Cu2+ complex of the chelator 4-[(1,4,8,11-tetraazacyclotetradec-1-yl)-methyl]benzoic acid was demonstrated at physiological pH as well as under acidic conditions. The chelating agent was conjugated to AB35, a monoclonal antibody directed against CEA, without a significant loss of immunoreactivity. The conjugate could, under optimal labeling conditions, be labeled with 67Cu in acetate buffer with a full occupancy of ligands within 20 min. This radiolabeled conjugate showed no transfer of radiocopper to serum proteins in human serum over 7 days. The biodistribution in tumor-bearing mice was measured and compared to that of iodinated AB35. Tumor uptake was high with 15 +/- 3% ID (injected dose)/g after 24 h and 32 +/- 7% ID/g after 96 h for the 67Cu-labeled antibody and 13 +/- 4% ID/g after 24 h and 14 +/- 2% ID/g after 96 h for the 125I-labeled antibody. Whereas radioactivity in normal organs decreased with time after 24 h, increased residence time was shown up to 4 days with the 67Cu-labeled AB35.     

Stability of metal ion complexes formed with methyl phosphate and hydrogen phosphate

 The acidity constants of methyl phosphoric acid, CH₃OPO(OH)₂, and orthophosphoric acid, HOPO(OH)₂, and the stability constants of the 1 : 1 complexes formed between Mg²⁺, Ca²⁺, Sr²⁺, Ba²⁺, Mn²⁺, Co²⁺, Ni²⁺, Cu²⁺, Zn²⁺, or Cd²⁺ and methyl phosphate, CH₃OPO₃ ^2–, or hydrogen phosphate, HOPO₃ ^2–, were determined by potentiometric pH titration in aqueous solution (25  °C;I = 0.1 M, NaNO₃). On the basis of previously established log K versus pK _a straight-line plots for the complexes of simple phosphate monoesters and phosphonate derivatives, R-PO₃ ^2–, where R is a noncoordinating residue, it is shown that the stability of the M(CH₃OPO₃) complexes is solely determined (as one might expect) by the basicity of the –PO₃ ^2– residue. It is emphasized that the mentioned reference lines may also be used to reveal increased complex stabilities, for example, for certain complexes formed with 8-quinolyl phosphate the occurrence of 7-membered chelates can be proven in this way; the same procedure is also applicable to complexes of nucleotides, etc. The M(HOPO₃) complexes are slightly more stable (on average by 0.08 log unit) than it is expected from the basicity of HPO₄ ^2–; this observation is attributed to a more effective solvation, including hydrogen bonding, than is possible with CH₃OPO₃ ^2– species. Received: 9 November 1995 / Accepted: 5 February 1996     

How Soluble GARP Enhances TGFβ Activation

GARP (glycoprotein A repetitions predominant) is a cell surface receptor on regulatory T-lymphocytes, platelets, hepatic stellate cells and certain cancer cells. Its described function is the binding and accommodation of latent TGFβ (transforming growth factor), before the activation and release of the mature cytokine. For regulatory T cells it was shown that a knockdown of GARP or a treatment with blocking antibodies dramatically decreases their immune suppressive capacity. This confirms a fundamental role of GARP in the basic function of regulatory T cells. Prerequisites postulated for physiological GARP function include membrane anchorage of GARP, disulfide bridges between the propeptide of TGFβ and GARP and connection of this propeptide to α_vβ₆ or α_vβ₈ integrins of target cells during mechanical TGFβ release. Other studies indicate the existence of soluble GARP complexes and a functionality of soluble GARP alone. In order to clarify the underlying molecular mechanism, we expressed and purified recombinant TGFβ and a soluble variant of GARP. Surprisingly, soluble GARP and TGFβ formed stable non-covalent complexes in addition to disulfide-coupled complexes, depending on the redox conditions of the microenvironment. We also show that soluble GARP alone and the two variants of complexes mediate different levels of TGFβ activity. TGFβ activation is enhanced by the non-covalent GARP-TGFβ complex already at low (nanomolar) concentrations, at which GARP alone does not show any effect. This supports the idea of soluble GARP acting as immune modulator in vivo.     

Microbiological processes at the interface of aerobic and anaerobic waters in the deep-water zone of the Black Sea

Chemical and key microbiological processes (assimilation of carbon dioxide, oxidation and formation of methane, and sulfate reduction) occurring at the boundary between the aerobic-anaerobic interface in the deep-water zone of the Black Sea were investigated. Measurements were taken at depths from 90 to 300 m at intervals of 5-10 m. The integral rate of the dark assimilation of carbon dioxide varied from 120 to 207 mg C/(m2 day) with a maximum at the boundary of cyclonic currents. The organic matter (OM) formed from methane comprised less than 5% of the OM formed from carbon dioxide. A comparison between the rates of methane oxidation and methane production suggests that methane that is oxidized at depths from 100 to 300 m was formed in deeper water horizons. The maximum rate of sulfate reduction (1230 mg S/(m2 day)) was observed in the western halistatic region, and the minimum rate (490 mg S/(m2 day)), in the eastern halistatic region. The average rate of hydrogen sulfide production measured at three deep-sea stations amounted to 755 mg S/(m2 day), or 276 g S/(m2 year).