RESUMEN

DNA has emerged as a highly versatile construction material for nanometer-sized structures and sophisticated molecular machines and circuits. The successful application of nucleic acid based systems greatly relies on their ability to autonomously sense and act on their environment. In this feature article, the development of DNA-based strategies to dynamically control protein activity via oligonucleotide triggers is discussed. Depending on the desired application, protein activity can be controlled by directly conjugating them to an oligonucleotide handle, or expressing them as a fusion protein with DNA binding motifs. To control proteins without modifying them chemically or genetically, multivalent ligands and aptamers that reversibly inhibit their function provide valuable tools to regulate proteins in a noncovalent manner. The goal of this feature article is to give an overview of strategies developed to control protein activity via oligonucleotide-based triggers, as well as hurdles yet to be taken to obtain fully autonomous systems that interrogate, process and act on their environments by means of DNA-based protein control.

Asunto(s)

RESUMEN

An analytical model of a single-atom electron source is presented, where electrons are created by near-threshold photoionization of an isolated atom. The model considers the classical dynamics of the electron just after the photon absorption, i.e. its motion in the potential of a singly charged ion and a uniform electric field used for acceleration. From closed expressions for the asymptotic transverse electron velocities and trajectories, the effective source temperature and the virtual source size can be calculated. The influence of the acceleration field strength and the ionization laser energy on these properties has been studied. With this model, a single-atom electron source with the optimum electron beam properties can be designed. Furthermore, we show that the model is also applicable to ionization of rubidium atoms, and thus also describes the ultracold electron source, which is based on photoionization of laser-cooled alkali atoms.

RESUMEN

We present a detailed description of measurements of the effective temperature of a pulsed electron source, based on near-threshold photoionization of laser-cooled atoms. The temperature is determined by electron beam waist scans, source size measurements with ion beams, and analysis with an accurate beam line model. Experimental data is presented for the source temperature as a function of the wavelength of the photoionization laser, for both nanosecond and femtosecond ionization pulses. For the nanosecond laser, temperatures as low as 14 ± 3 K were found; for femtosecond photoionization, 30 ± 5 K is possible. With a typical source size of 25 µm, this results in electron bunches with a relative transverse coherence length in the 10⁻4 range and an emittance of a few nm rad.

RESUMEN

The study of structural dynamics of complex macromolecular crystals using electrons requires bunches of sufficient coherence and charge. We present diffraction patterns from graphite, obtained with bunches from an ultracold electron source, based on femtosecond near-threshold photoionization of a laser-cooled atomic gas. By varying the photoionization wavelength, we change the effective source temperature from 300 K to 10 K, resulting in a concomitant change in the width of the diffraction peaks, which is consistent with independently measured source parameters. This constitutes a direct measurement of the beam coherence of this ultracold source and confirms its suitability for protein crystal diffraction.

RESUMEN

With the development of ultrafast electron and X-ray sources it is becoming possible to study structural dynamics with atomic-level spatial and temporal resolution. Because of their short mean free path, electrons are particularly well suited for investigating surfaces and thin films, such as the challenging and important class of membrane proteins. To perform single-shot diffraction experiments on protein crystals, an ultracold electron source was proposed, based on near-threshold photoionization of laser-cooled atoms, which is capable of producing electron pulses of both high intensity and high coherence. Here we show that high coherence electron pulses can be produced by femtosecond photoionization, opening up a new regime of ultrafast structural dynamics experiments. The transverse coherence turns out to be much better than expected on the basis of the large bandwidth of the femtosecond ionization laser pulses. This surprising result can be explained by analysis of classical electron trajectories.

RESUMEN

OBJECTIVE: The primary objective was to compare the antioxidant activity of micronised fenofibrate 200 mg to 400 IU of vitamin E and placebo, on the LDL and VLDL particles of patients with type 1 diabetes. The secondary objective was to investigate the possible synergy between micronized fenofibrate and vitamin E and to compare the efficacy of these treatments on lipids. METHODS: A double-blind, placebo-controlled trial in which patients were randomised into three treatment groups after a wash-out period of 8 weeks: the placebo group (Pla/Pla-group) in which patients received placebo during two consecutive periods of 8 weeks, the vitamin E group (Vit E/Vit E-group) in which patients received Vitamin E during two consecutive periods, and the fenofibrate/Vitamin E group (Fen/Fen + Vit E-group) in which patients received fenofibrate during the first period, followed by fenofibrate and vitamin E during the consecutive period. Blood samples taken at each visit, were analysed for routine biochemistry, blood lipids and copper mediated lipid peroxidation in vitro. RESULTS: The lag time of the non-HDL lipoprotein oxidation increased in the group given fenofibrate. The lag-time increased further when fenofibrate and vitamin E were given in association. (This reached significance in the intention-to-treat population, P = 0.034). The AUC of TBARS formation in the Vit E/Vit E group decreased after the first period, but this effect was not enhanced by continuing vitamin E for another 8 weeks. The AUC of TBARS formation did not change significantly after the administration of fenofibrate. Only after the second period, when vit E was given in association, the AUC of TBARS formation decreased significantly (P = 0.004). Fenofibrate caused a significant decrease in total and LDL-cholesterol and triglycerides (P < 0.05). In contrast, vitamin E had no effect on lipids. CONCLUSIONS: The combination of micronized fenofibrate 200 mg/day and vitamin E 400 IU/day tended to increase the resistance of non-HDL lipoproteins to copper-mediated oxidation, shown by a prolongation of the lag-time. Vitamin E administration only, decreased the oxidation of non-HDL lipoproteins shown by a reduction of TBARS formation. This protective effect of vitamin E tended to be amplified by micronized fenofibrate.

Asunto(s)

RESUMEN

Mycophenolate mofetil (MMF) is one of the new immunosuppressive drugs used in renal transplantation. MMF inhibits the de novo purine synthesis. Since this purine synthesis in lymphocytes entirely depends on the de novo pathway, MMF is considered to cause a selective inhibition of T- and B lymphocytes. Recently, 4 transplant patients out of 30 developed a severe anemia in the early post-transplantation period. Their immediate post-transplantation immunosuppression consisted of corticosteroids, cyclosporine and MMF. They all received anti-T-lymphocyte globulin (ATG) as induction treatment or because of rejection. In all 4 patients, iron supplementation and a treatment with erythropoietin were started. Blood loss, deficiencies, hemolysis, drug interactions or viral infections were excluded as causes of the anemia. Bone marrow biopsies were carried out, showing pure red cell aplasia that was ascribed to the use of MMF. Cessation or reduction of MMF was followed by a hematological improvement after 5-9 days. We hypothesized that MMF has a broader antiproliferative effect than its proposed lymphocyte-specific effect.

Asunto(s)

RESUMEN

In erythrocytes from diabetic patients, increased membrane lipid peroxidation might lead to abnormalities in composition and function. To study this relationship, we investigated the effects of a moderate pharmacologic dose of vitamin E for 1 y on erythrocyte membrane peroxidation in vitro and on its fatty acid composition, antioxidant capacity and rheological function. In a random and double-blind manner, type 1 diabetic patients (n = 44) were assigned to the following two groups: Group S received 250 IU (168 mg) d-alpha tocopherol 3 times daily for 1 y. Group P received placebo for 6 mo followed by d-alpha-tocopherol for an additional 6 mo. Variables were monitored every 3 mo. After 3 mo of supplementation, serum vitamin E doubled (P < 0.0005), thiobarbituric acid reactive substances in erythrocyte membranes incubated with tert-butyl hydroperoxide decreased by 25% (P = 0.006) and the lagtime of fluorescence increased from 28 +/- 16 to 41 +/- 28 min (P = 0.028). Patients who did not respond to supplementation (13 of 44) had lower serum lipids (P = 0.017) and body mass index (P = 0.024). We did not detect any significant effects of vitamin E supplementation on membrane lipid composition, antioxidant capacity or blood viscosity. Continuing supplementation for up to 1 y did not further affect serum vitamin E or membrane peroxidation. Stopping supplementation was followed by a return to inclusion values. These results show that the decrease in erythrocyte membrane peroxidation after vitamin E supplementation is moderate, saturable, reversible, restricted to some individuals and has no detectable effect on erythrocyte composition and function.

Asunto(s)

RESUMEN

BACKGROUND: Vitamin E supplementation has been proposed as adjunctive therapy to counteract the increased LDL oxidation in diabetes and thus prevent or delay cardiovascular complications. OBJECTIVE: The objective of this study was to investigate the effect of a moderate pharmacologic dose of vitamin E for

Asunto(s)

RESUMEN

OBJECTIVE: Our aim was to study the relationship between the magnesium status in type 1 diabetic patients and disturbances in nerve conduction velocity. Furthermore we wanted to investigate whether repletion of magnesium depletion could improve the decreased nerve conduction velocity measurements. In a cross-sectional study, 154 type 1 diabetic patients were screened for their erythrocyte magnesium content and an electrophysiological measurement of the peripheral nervous system was carried out. In a subsequent intervention study, out of this screened population, 23 type 1 patients, with disturbed nerve conduction velocity measurements and low erythrocyte magnesium levels [< 2.3 mmol/L) were given oral magnesium supplements, during 1 year. Twenty type 1 patients with identical characteristics served as controls. In the cross-sectional study disturbed nerve conduction velocities were found in the older patients, in patients with a longer duration of diabetes and a worse metabolic control. EMG polyneuropathy signs were significantly more frequent in diabetic patients with low erythrocyte Mg. The intervention study demonstrated that under unchanged metabolic control, supplementation with magnesium could improve nerve conduction, especially in younger patients with a shorter duration of diabetes. Erythrocyte Mg was lower in type 1 diabetic patients with polyneuropathy. Mg supplementation increasing Mg RBC might (possibly?) improve nerve conduction measured by electromyography at least in younger patients with a short duration of diabetes and presenting early signs of the neurological complication.

Asunto(s)

RESUMEN

There is overwhelming evidence that platelets from diabetic individuals are hyperreactive, not only when microvascular complications are apparent, but already at an early stage of the disease. There is still controversy about the question of whether primary hyperreactive platelets may contribute to the origin or progression of microangiopathy or whether diabetic platelet hyperfunctionality is just a logical consequence of a continuous low-grade activation of platelets by contact with a diseased microvascular wall. As a consequence of platelet activation, the outer layer of its phospholipid membrane is more procoagulant than in the quiescent state, stimulating thrombin formation in plasma. This platelet function is called platelet procoagulant activity. We studied platelet prothrombinase activity (PPA), a final pathway platelet procoagulant activity of type 1 diabetic platelets, and looked for an eventual correlation with microvascular disease (background retinopathy) and mean platelet volume (MPV). Stypven clotting times (SCTs), reflecting PPA expression, and MPV of citrated platelet-rich plasma (PRP), were measured in 21 patients with type 1 diabetes-10 with and 11 without background retinopathy-under clinically acceptable metabolic control and compared them to 20 disease-free voluntary controls. We also compared PPA expression and MPV in diabetic individuals with and without retinopathy. With the SCT, a selective test adapted for studying PPA in PRP, we found hyperexpression of PPA in all diabetic patients. We found no difference in MPV between diabetic and control PRP. Comparing patients with and without background retinopathy we found no significant difference in PPA expression. From these results, we suggest that the phospholipid surface of diabetic platelets, more than the surface of normal control platelets, stimulate the expression of PPA. This diabetic platelet coagulant anomaly was not related to an increased platelet mass (higher MPV) nor to the presence of microangiopathy. We conclude that PPA hyperexpression is associated with patients with type 1 diabetes, already occurring in an early stage of the disease, and not necessarily a consequence of early-stage microvascular disease, because the anomaly is also demonstrable, in the same degree, in patients with diabetes without microangiopathy.

Asunto(s)

RESUMEN

Type 1 diabetic patients with low RBC-MG have an increased in vitro oxidizability of LDL and VLDL as compared to matched diabetic controls with normal RBC-MG. In this study the effect of a 10 week intensive oral + i.v. supplementation with Mg was studied in 11 depleted type 1 patients under stable metabolic control. Despite the significant increase of erythrocyte Mg and the higher body stores demonstrated by the Mg retention test, no significant decrease of the oxidizability could be demonstrated after supplementation. Persistent Mg loss in the presence of peripheral hyperinsulinaemia, insufficient and/or too short supplementation and decreased antioxidant defence could be the reasons for this negative result.

Asunto(s)

RESUMEN

Magnesium depletion in diabetic patients is a frequent observation and is involved in the pathogenesis of acute and chronic complications. In this study the effect of a 10 week intensive oral + i.v. supplementation of Mg was studied in 11 depleted IDDM patients with stable metabolic control. Ionized Mg decreased and erythrocyte Mg increased significantly together with an increased storage of Mg in the body demonstrated with a classical retention test. Normalization of the Mg parameters was however not obtained. Lipid parameters and metabolic control remained unchanged.

Asunto(s)

RESUMEN

Diabetic nephropathy is the most important cause of increased morbidity and premature mortality in patients with insulin dependent diabetes mellitus. Several longitudinal studies done in different countries have shown that microalbuminuria strongly predicts the development of diabetic nephropathy in insulin dependent diabetes mellitus. Very few data are available about the situation in Belgium. In a retrospective study we analysed the data of 271 insulin dependent diabetic patients attending the University Hospital of Antwerp during the year 1994. Normoalbuminuria was defined as a urinary albumin excretion rate of < or = 20 micrograms/min, microalbuminuria as 20-200 micrograms/min and macroalbuminuria as > 200 micrograms/min. In our population we find an overall prevalence of microalbuminuria of 14%. Patients with microalbuminuria are characterised by an earlier onset and a longer duration of diabetes when compared with patients with normoalbuminuria. Further we find that they have a worse glycemic control, a higher frequency of raised arterial blood pressure and a higher percentage of proliferative retinopathy. These data are comparable with those from other studies done in different countries. They show that patients with microalbuminuria should be regarded as a high risk group and therefore urinary albumin excretion rate should be monitored routinely. Also they stress the importance to strive to a good metabolic control and to an aggressive treatment of arterial hypertension.

Asunto(s)

RESUMEN

In 1991, Belgium realized, on a national level, a change-over from U40 to U100 insulin. We took advantage of this evolution to investigate the consequences of changing the concentration of insulin. The patients' weight, daily insulin dosis, insulin binding-capacity of plasma and glycated hemoglobin HbA1c were registered before, and after the change of concentration. Overall, none of these parameters underwent an obvious change, except for the percentage of insulin binding that significantly decreased after the adaptation. Especially in the range of 40% or more insulin binding, the decrease becomes very pronounced. In conclusion, changing the insulin concentration from U40 to U100, did not lead to any harmful clinical consequence. On the contrary, a positive influence of this adaptation, in terms of decreased amount of insulin antibodies was suggested. Probably this decrease is not clinically relevant, since neither the glycated hemoglobin, nor the total daily insulin dose underwent a similar change.

Asunto(s)