Enantioselective hydroxylation of 4-alkylphenols by vanillyl alcohol oxidase

Vanillyl alcohol oxidase (VAO) from Penicillium simplicissimum catalyzes the enantioselective hydroxylation of 4-ethylphenol, 4-propylphenol, and 2-methoxy-4-propylphenol into 1-(4'-hydroxyphenyl)ethanol, 1-(4'-hydroxyphenyl)propanol, and 1-(4'-hydroxy-3'-methoxyphenyl)propanol, respectively, with an ee of 94% for the R enantiomer. The stereochemical outcome of the reactions was established by comparing the chiral GC retention times of the products to those of chiral alcohols obtained by the action of the lipases from Candida antarctica and Pseudomonas cepacia. Isotope labeling experiments revealed that the oxygen atom incorporated into the alcoholic products is derived from water. During the VAO-mediated conversion of 4-ethylphenol/4-propylphenol, 4-vinylphenol/4-propenylphenol are formed as side products. With 2-methoxy-4-propylphenol as a substrate, this competing side reaction is nearly abolished, resulting in less than 1% of the vinylic product, isoeugenol. The VAO-mediated conversion of 4-alkylphenols also results in small amounts of phenolic ketones indicative for a consecutive oxidation step. Copyright 1998 John Wiley & Sons, Inc.     

The nucleotide metabolism in lactate perfused hearts under ischaemic and reperfused conditions

It was examined whether lactate influences postischaemic hemodynamic recovery as a function of the duration of ischaemia and whether changes in high-energy phosphate metabolism under ischaemic and reperfused conditions could be held responsible for impairment of cardiac function. To this end, isolated working rat hearts were perfused with either glucose (11 mM), glucose (11 mM) plus lactate (5 mM) or glucose (11 mM) plus pyruvate (5 mM). The extent of ischaemic injury was varied by changing the intervals of ischaemia, i.e. 15, 30 and 45 min. Perfusion by lactate evoked marked depression of functional recovery after 30 min of ischaemia. Perfusion by pyruvate resulted in marked decline of cardiac function after 45 min of ischaemia, while in glucose perfused hearts hemodynamic performance was still recovered to some extent after 45 min of ischaemia. Hence, lactate accelerates postischaemic hemodynamic impairment compared to glucose and pyruvate. The marked decline in functional recovery of the lactate perfused hearts cannot be ascribed to the extent of degradation of high-energy phosphates during ischaemia as compared to glucose and pyruvate perfused hearts. Glycolytic ATP formation (evaluated by the rate of lactate production) can neither be responsible for loss of cardiac function in the lactate perfused hearts. Moreover, failure of reenergization during reperfusion, the amount of nucleosides and oxypurines lost or the level of high-energy phosphates at the end of reperfusion cannot explain lactate-induced impairment. Alternatively, the accumulation of endogenous lactate may have contributed to ischaemic damage in the lactate perfused hearts after 30 min of ischaemia as it was higher in the lactate than in the glucose or pyruvate perfused hearts. It cannot be excluded that possible beneficial effects of the elevated glycolytic ATP formation during 15 to 30 min of ischaemia in the lactate perfused hearts are counterbalanced by the detrimental effects of lactate accumulation.     

Cyclodextrin encapsulation to prevent the loss of l-menthol and its retention during drying

The taste and flavor of spray-dried powdered products are the most important quality factors. In the present study, molecular encapsulation in cyclodextrin was applied to prevent the loss of a hydrophobic flavor compound (l-menthol) during the drying of a droplet. beta-Cyclodextrin appeared to be a better encapsulant for menthol than alpha- and gamma-cyclodextrin. The retention of menthol increased with increasing concentration of both cyclodextrin and maltodextrin. A simple mathematical model is proposed for estimating the flavor retention. The theoretical results by this model estimated well the final retention of menthol encapsulated in a blend of beta-cyclodextrin and maltodextrin.     

Antibodies to citrullinated proteins and differences in clinical progression of rheumatoid arthritis

Antibodies to citrullinated proteins (anti-cyclic-citrullinated peptide [anti-CCP] antibodies) are highly specific for rheumatoid arthritis (RA) and precede the onset of disease symptoms, indicating a pathogenetic role for these antibodies in RA. We recently showed that distinct genetic risk factors are associated with either anti-CCP-positive disease or anti-CCP-negative disease. These data are important as they indicate that distinct pathogenic mechanisms are underlying anti-CCP-positive disease or anti-CCP-negative disease. Likewise, these observations raise the question of whether anti-CCP-positive RA and anti-CCP-negative RA are clinically different disease entities. We therefore investigated whether RA patients with anti-CCP antibodies have a different clinical presentation and disease course compared with patients without these autoantibodies. In a cohort of 454 incident patients with RA, 228 patients were anti-CCP-positive and 226 patients were anti-CCP-negative. The early symptoms, tender and swollen joint count, and C-reactive protein level at inclusion, as well as the swollen joint count and radiological destruction during 4 years of follow-up, were compared for the two groups. There were no differences in morning stiffness, type, location and distribution of early symptoms, patients' rated disease activity and C-reactive protein at inclusion between RA patients with and without anti-CCP antibodies. The mean tender and swollen joint count for the different joints at inclusion was similar. At follow-up, patients with anti-CCP antibodies had more swollen joints and more severe radiological destruction. Nevertheless, the distribution of affected joints, for swelling, bone erosions and joint space narrowing, was similar. In conclusion, the phenotype of RA patients with or without anti-CCP antibodies is similar with respect to clinical presentation but differs with respect to disease course.     

Giant membrane vesicles as a model to study cellular substrate uptake dissected from metabolism

In order to use giant vesicles for substrate uptake studies in metabolically important tissues, we characterized giant vesicles isolated from heart, liver, skeletal muscle and adipose tissue. We investigated which cell types and which plasma membrane regions are involved in giant vesicle formation and we examined the presence of transporters for metabolic substrates. Analysis of giant vesicles with markers specific for distinct cell types and distinct domains of the plasma membrane reveals that the plasma membrane of parenchymal cells, but not endothelial cells, are the source of the vesicle membranes. In addition, plasma membrane regions enriched in caveolae and involved in docking of recycling vesicles from the endosomal compartment are retained in giant vesicles, indicating that KCl-induced alterations in recycling processes are involved in giant vesicle formation. Giant vesicles contain vesicular lumen consisting of the soluble constituents of the cytoplasm including, fatty-acid binding proteins. Furthermore, giant vesicles isolated from heart, liver, skeletal muscle and adipose tissue are similar in size (10–15 m) and shape and do not contain subcellular organelles, providing the advantage that substrate fluxes in the different organs can be studied independently of the surface/volume ratio but most importantly in the absence of intracellular metabolism.     

Feasibility of tailored treatment based on risk stratification in patients with early rheumatoid arthritis

Introduction

Personalized medicine is the holy grail of medicine. The EULAR recommendations for the management of rheumatoid arthritis (RA) support differential treatment between patients with baseline characteristics suggestive of a non-poor prognosis (non-PP) or poor prognosis (PP) (presence of autoantibodies, a high inflammatory activity and damage on radiographs). We aimed to determine which prognostic risk groups benefit more from initial monotherapy or initial combination therapy.

Methods

508 patients were randomized to initial monotherapy (iMono) or initial combination therapy (iCombo). Disease outcomes of iMono and iCombo were compared within non-PP or PP groups as determined on baseline characteristics

Results

PP patients treated with iCombo after three months more often achieved ACR20 (70% vs 38%, P <0.001), ACR50 (48% vs 13%, P <0.001) and ACR70 response (24% vs 4%, P <0.001) than those treated with iMono, and had more improvement in HAQ (median decrease 0.75 vs 0.38, P <0.001). After 1 year, differences in ACR20 response and DAS-remission remained; PP patients treated with iCombo (vs iMono) had less radiographic progression (median 0.0 vs 1.5, P =0.001).Non-PP patients treated with iCombo after three months more often achieved an ACR response (ACR20: 71% versus 44%, P <0.001; ACR50: 49% vs 13%, P <0.001; ACR70: 17% vs 3%, P =0.001) than with iMono, and functional ability showed greater improvement (median decrease in HAQ 0.63 vs 0.38, P <0.001). After 1 year, differences in ACR20 and ACR50 response remained; radiographic progression was comparable between the groups.Non-PP and PP patients responded equally well to iCombo in terms of improvement of functional ability, with similar toxicity.

Conclusions

Since PP and non-PP patients benefit equally from iCombo through earlier clinical response and functional improvement than with iMono, we conclude that personalized medicine as suggested in the guidelines is not yet feasible. The choice of treatment strategy should depend more on rapid relief of symptoms than on prognostic factors.

Trial registration

Netherlands Trial Register NTR262 (registered 7 September 2005) and NTR265 (8 September 2005).

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-014-0430-3) contains supplementary material, which is available to authorized users.     

Bioaccumulation of selected metals in the gill,liver and muscle tissue of rednose labeo Labeo rosae from two impoundments on the Olifants River,Limpopo river system,South Africa

Metal concentrations in the gill, muscle and liver tissues of Labeo rosae from two impoundments, Loskop and Flag Boshielo dams on the Olifants River, were evaluated in 2011 to detect patterns in metal associations between tissues and impoundments. Elevated concentrations of Ba, Zn, B, Al, Si and Fe, relative to a pristine site in the catchment, were found in the muscle, liver and gill tissues at both impoundments. Molybdenum concentrations were exceptionally high in all tissues at Loskop Dam and in liver at Flag Boshielo Dam. No definite pattern in the ratio metal concentrations within, or between, fish tissues was identified. The expected trend, liver > gills > muscle, was found at both impoundments, but was less prominent at Loskop Dam. Metal concentrations in muscle of Loskop Dam fish were significantly higher than in those at Flag Boshielo Dam. The inverse was true for liver. The long-term impact of elevated metal concentrations on fish health at both impoundments raises concern.     

Human recombinant thiamine triphosphatase: purification, secondary structure and catalytic properties

Thiamine triphosphate (ThTP) is found in most living organisms and it may act as a phosphate donor for protein phosphorylation. We have recently cloned the cDNA coding for a highly specific mammalian 25 kDa thiamine triphosphatase (ThTPase; EC 3.6.1.28). As the enzyme has a high catalytic efficiency and no sequence homology with known phosphohydrolases, it was worth investigating its structure and catalytic properties. For this purpose, we expressed the untagged recombinant human ThTPase (hThTPase) in E. coli, produced the protein on a large scale and purified it to homogeneity. Its kinetic properties were similar to those of the genuine human enzyme, indicating that the recombinant hThTPase is completely functional. Mg2+ ions were required for activity and Ca2+ inhibited the enzyme by competition with Mg2+. With ATP as substrate, the catalytic efficiency was 10(-4)-fold lower than with ThTP, confirming the nearly absolute specificity of the 25 kDa ThTPase for ThTP. The activity was maximum at pH 8.5 and very low at pH 6.0. Zn2+ ions were inhibitory at micromolar concentrations at pH 8.0 but activated at pH 6.0. Kinetic analysis suggests an activator site for Mg2+ and a separate regulatory site for Zn2+. The effects of group-specific reagents such as Woodward's reagent K and diethylpyrocarbonate suggest that at least one carboxyl group in the active site is essential for catalysis, while a positively charged amino group may be involved in substrate binding. The secondary structure of the enzyme, as determined by Fourier-transform infrared spectroscopy, was predominantly beta-sheet and alpha-helix.     

Endovascular brachytherapy in coronary arteries: the Rotterdam experience

Purpose: The use of endovascular coronary brachytherapy to prevent restenosis following percutaneous transluminal coronary angioplasty (PTCA) began in April 1997 at the Department of Interventional Cardiology of the Thoraxcenter at the University Hospital of Rotterdam. This article reviews the more than 250 patients that have been treated so far.Methods and Materials: The Beta-Cath System (Novoste), a manual, hydraulic afterloader with 12 90Sr seeds, was used in the Beta Energy Restenosis Trial (BERT-1.5, n=31), for compassionate use (n=25), in the Beta-Cath System trial (n=27) and in the Beta Radiation in Europe (BRIE, n=14). Since the Beta-Cath System has been commercialized in Europe, 57 patients have been treated and registered in RENO (Registry Novoste). In the Proliferation Reduction with Vascular Energy Trial (PREVENT), 37 patients were randomized using the Guidant-Nucletron remote control afterloader with a 32P source wire and a centering catheter. Radioactive 32P coated stents have been implanted in 102 patients. In the Isostent Restenosis Intervention Study 1 (IRIS 1), 26 patients received a stent with an activity of 0.75-1.5 μCi, and in the IRIS 2 (European 32P dose response trial), 40 patients were treated with an activity of 6-12 μCi. In two consecutive pilot trials, radioactive stents with non-radioactive ends (cold-end stents) and with ends containing higher levels of activity (hot-end stents) were implanted in 21 and 17 patients, respectively.Results: In the BERT-1.5 trial, the radiation dose, prescribed at 2 mm from the source train (non-centered), was 12 Gy (10 patients), 14 Gy (10 patients) and 16 Gy (11 patients). At 6-month follow-up, 8 out of 28 (29%) patients developed restenosis. The target lesion revascularization rate (TLR) was 7 out of 30 (23%) at 6 months and 8 out of 30 (27%) at 1 year. Two patients presented with late thrombosis in the first year. For compassionate use patients, a restenosis rate (RR) of 53% was observed. In the PREVENT trial, 34 of 37 patients underwent an angiographic 6-month follow-up. The doses prescribed at 0.5 mm depth into the vessel wall were 0 Gy (8), 28 Gy (9), 35 Gy (11) and 42 Gy (8). TLR was 14% in the irradiated patients and 25% in the placebo group. One patient developed late thrombosis. In the IRIS 1 trial, 23 patients showed an RR of 17% (in-stent). In the IRIS 2 trial, in-stent restenosis was not seen in 36 patients at 6-month follow-up. However, a high RR (44%) was observed at the stent edges.Conclusions: The integration of vascular brachytherapy in the catheterization laboratory is feasible and the different treatment techniques that are used are safe. Problems, such as edge restenosis and late thrombotic occlusion, have been identified as limiting factors of this technique. Solutions have been suggested and will be tested in future trials.