[Wine consumption and prevention of coronary artery disease]. / Weingenuss und Prävention der koronaren Herzkrankheit.

There is a J-shaped correlation between the amount of alcohol consumed per day and overall mortality risk and an inverse correlation between the amount of alcohol consumed per day and cardiovascular mortality. The evidence is stronger for men than for women. The correlations are independent of the type of alcoholic beverage predominantly consumed. Possible mechanisms explaining the cardioprotective, antiatherosclerotic effects of moderate alcohol consumption are inhibition of platelet aggregation, increase in serum high density lipoprotein (HDL) levels and prevention of diabetes mellitus. The two latter mechanisms can also explain a delayed progression of atherosclerosis due to alcohol consumption. The beneficial effects are counteracted by detrimental effects of alcohol on the incidence of cancer diseases, liver cirrhosis, violence and accidents; therefore, alcohol consumption in general cannot be recommended for prevention of cardiovascular diseases.

Conformational changes of the betaine transporter BetP from Corynebacterium glutamicum studied by pulse EPR spectroscopy.

The betaine transporter BetP from Corynebacterium glutamicum is activated by hyperosmotic stress critically depending on the presence and integrity of its sensory C-terminal domain. The conformational properties of the trimeric BetP reconstituted in liposomes in the inactive state and during osmotic activation were investigated by site-directed spin labeling and electron paramagnetic resonance (EPR) spectroscopy. Comparison of intra- and intermolecular inter spin distance distributions obtained by double electron-electron resonance (DEER) EPR with the crystal structure of BetP by means of a rotamer library analysis suggest a rotation of BetP protomers within the trimer by about 15° as compared to the X-ray structure. Furthermore, we observed conformational changes upon activation of BetP, which are reflected in changes of the distances between positions 545 and 589 of different protomers in the trimer. Introduction of proline at positions 550 and 572, both leading to BetP variants with a permanent (low level) transport activity, caused changes of the DEER data similar to those observed for the activated and inactivated state, respectively. This indicates that not only displacements of the C-terminal domain in general but also concomitant interactions of its primary structure with surrounding protein domains and/or lipids are crucial for the activity regulation of BetP.

[Diabetic foot syndrome-Part 1 : Definition, pathophysiology, diagnostics and classification]. / Diabetisches Fußsyndrom ­ Teil 1 : Definition, Pathophysiologie, Diagnostik und Klassifikation.

There are ca. 8 million persons with diabetes mellitus living in Germany. A late sequelae of diabetes is the diabetic foot syndrome (DFS), the prevalence of which is greatly increasing. It comprises all alterations of the foot as a result of diabetic polyneuropathy as well as microvascular and macrovascular (peripheral arterial occlusive disease, PAOD) alterations. Many of the ca. 250,000 newly diagnosed diabetic foot ulcers per year become chronic wounds. Despite intensive efforts for prevention, early diagnosis and adequate wound care, ca. 13,000 persons with diabetes undergo major limb amputation in Germany every year. With consistent treatment in interdisciplinary centers and by exhausting all possible methods of wound treatment, pressure relief as well as arterial revascularization, the major amputation rate in patients with diabetic foot problems can be reduced by 80%. With a suitable strategy of prevention, the recurrence rate of foot ulcers would be reduced.

[Diabetic foot syndrome-Part 2 : Revascularization, treatment alternatives, care structures, recurrency prophylaxis]. / Diabetisches Fußsyndrom ­ Teil 2 : Revaskularisation, Behandlungsalternativen, Versorgungsstrukturen, Rezidivprophylaxe.

Diabetic foot syndrome (DFS) is the most frequent reason for major amputations in Germany. The majority of foot lesions are triggered by repetitive pressure in diabetic polyneuropathy. Peripheral arterial occlusive disease (PAOD) impairs wound healing and is the main risk factor for amputations. The treatment of wounds and infections as well as timely revascularization are decisive. The use of endovascular and vascular surgical methods depends on the distribution pattern and length of the occlusion processes. Both procedures are complementary. Bypass surgery is of great importance for neuroischemic DFS. Multidisciplinary centers that provide revascularization in DFS can achieve an improvement of arterial blood flow in 90% of the cases and reduce the amputation rate by up to 80%. Due to the high recurrence rate of diabetic foot lesions, measures for secondary prophylaxis are of exceptional importance (podological and orthopedic technical care, foot surgery).

[Minor amputations for diabetic foot syndrome]. / Minoramputationen bei diabetischem Fusssyndrom.

Minor amputations are frequently performed for neuroischemic or neuropathic lesions of the diabetic foot. Depending on the definition used, minor amputations can range from toe to Syme amputations. Minor amputations are often combined with necrosectomy and débridement. For early and optimal rehabilitation, as much vital tissue as possible should be conserved, especially considering the skeletal structures of the foot (borderline amputation). Minor amputations are of utmost importance for the prevention of ascending infections and reduce the duration of clinical and outpatient treatment. Minor amputations should be performed only by experienced surgeons and only if arterial perfusion is sufficient. They should be as tissue-conserving as possible and structured interdisciplinary postoperative care is mandatory. Metabolic control should be optimized. Controversial opinions exist with respect to the use of tourniquets, conservation or resection of cartilage and sesamoid bones, open amputation or primary closure of the wound, interdigital spacer function of toes, aseptic proximal transection of tendons, postoperative wound care, negative-pressure wound treatment and antibiotic therapy. In view of these controversies the most important minor amputation techniques are described and discussed.

[Diagnostics and therapy of the diabetic foot syndrome from a vascular surgery perspective]. / Diagnostik und Therapie des diabetischen Fusssyndroms aus gefässchirurgischer Sicht.

There are more than 6 million diabetic patients in Germany. Due to neuropathic and angiopathic long term damage the number of patients with diabetic foot syndrome has been increasing dramatically over the past years. Despite all efforts for prevention, early diagnosis and adequate therapy, more than 20,000 diabetics undergo major limb amputation in Germany every year. A major portion of these amputations could be avoided if an improvement of the arterial perfusion would be timely considered. By consequent therapy in interdisciplinary centres, and by applying all methods of arterial revascularization, the amputation rate in patients with diabetic foot problems could be reduced by 80%. This article describes the diagnostics and therapy of the diabetic foot syndrome from a vascular surgical point of view. The importance of endovascular, vascular surgical as well as combined (hybrid) procedures of revascularization is emphasized.

[The vascular surgeon's role in interdisciplinary treatment of diabetic foot syndrome]. / Interdisziplinäre Wundzentren in der Behandlung des diabetischen Fusssyndroms. Die Rolle des Gefässchirurgen.

There are more than 6 million diabetes patients in Germany. Due to long-term neuropathic and angiopathic sequelae, the number of patients with "diabetic foot syndrome" has increased dramatically in recent years. Diabetic foot ulcers have become one of the most common pathologies in interdisciplinary wound care centers. Because of its complex pathogenesis, diabetic foot syndrome needs a multidisciplinary therapeutic approach. More than 150,000 diabetics per year develop foot ulcers that often heal slowly and progress into chronic wounds. Despite all efforts at prevention, early diagnosis, and adequate therapy, more than 20,000 diabetics suffer major limb amputation in Germany every year. Applying stringent standards of care in interdisciplinary wound care centers, the amputation rate in patients with diabetic foot syndrome can be reduced to less than 50%. This article describes the complexity of diabetic foot syndrome with respect to pathogenesis, diagnostics, and therapy from a vascular surgeon's point of view. The importance of an interdisciplinary approach is emphasized.

Cathepsin D links TNF-induced acid sphingomyelinase to Bid-mediated caspase-9 and -3 activation.

Acidic noncaspase proteases-like cathepsins have been introduced as novel mediators of apoptosis. A clear role for these proteases and the acidic endolysosomal compartment in apoptotic signalling is not yet defined. To understand the role and significance of noncaspases in promoting and mediating cell death, it is important to determine whether an intersection of these proteases and the caspase pathway exists. We recently identified the endolysosomal aspartate protease cathepsin D (CTSD) as a target for the proapoptotic lipid ceramide. Here, we show that tumor necrosis factor (TNF)-induced CTSD activation depends on functional acid sphingomyelinase (A-SMase) expression. Ectopic expression of CTSD in CTSD-deficient fibroblasts results in an enhanced TNF-mediated apoptotic response. Intracellular colocalization of CTSD with the proapoptotic bcl-2 protein family member Bid in HeLa cells, and the ability of CTSD to cleave directly Bid in vitro as well as the lack of Bid activation in cathepsin-deficient fibroblasts indicate that Bid represents a direct downstream target of CTSD. Costaining of CTSD and Bid with Rab5 suggests that the endosomal compartments are the common 'meeting point'. Caspase-9 and -3 activation also was in part dependent on A-SMase and CTSD expression as revealed in the respective deficiency models. Our results link as novel endosomal intermediates the A-SMase and the acid aspartate protease CTSD to the mitochondrial apoptotic TNF pathway.

New principle for large-scale preparation of purified human pancreas islets.

Because successful human islet transplantation requires large quantities of viable islets that must be separated from the highly immunogenic exocrine tissue and because handpicking is too time-consuming and laborious to be clinically relevant, a new approach for solving this problem has been established in rat models. It is based on the principle that magnetic microspheres (MMSs) coupled to lectins with binding specificity for the exocrine tissue portion are trapped in an electromagnetic field, thus providing effluent islets of a high degree of purity. In this study our aim was to adapt this principle to human islet preparations. In this context our prime interest was focused on a lectin suitable for human pancreatic tissue. Of 19 different lectins tested, only 1, Wisteria floribunda agglutinin (WFA), is suitable, as shown by immunofluorescence, MMS-lectin binding, and magnetic separation.

Projection structure and oligomeric state of the osmoregulated sodium/glycine betaine symporter BetP of Corynebacterium glutamicum.

The high-affinity glycine betaine uptake system BetP, an osmosensing and osmoregulated sodium-coupled symporter from Corynebacterium glutamicum, was overexpressed in Escherichia coli with an N-terminal StrepII-tag, solubilized in beta-dodecylmaltoside and purified by streptactin affinity chromatography. Analytical ultracentrifugation indicated that BetP forms trimers in detergent solution. Detergent-solubilized BetP can be reconstituted into proteoliposomes without loss of function, suggesting that BetP is a trimer in the bacterial membrane. Reconstitution with E.coli polar lipids produced 2D crystals with unit cell parameters of 182A x 154A, gamma=90 degrees exhibiting p22(1)2(1) symmetry. Electron cryo-microscopy yielded a projection map at 7.5A. The unit cell contains four non-crystallographic trimers of BetP. Within each monomer, ten to 12 density peaks characteristic of transmembrane alpha-helices surround low-density regions that define potential transport pathways. Small but significant differences between the three monomers indicate that the trimer does not have exact 3-fold symmetry. The observed differences may be due to crystal packing, or they may reflect different functional states of the transporter, related to osmosensing and osmoregulation. The projection map of BetP shows no clear resemblance to other secondary transporters of known structure.

A comparison of the use of two immunomagnetic microspheres for secondary purification of pancreatic islets.

Immunomagnetic cell separation has been shown to be a highly attractive alternative to density-dependent methods for islet purification. There are two types of beads, magnetic inducible microspheres (MIMS) and Dynabeads, in this context. The aim of this study was to compare the two beads and ligands using the same method of purification. Two batches of collagenase were used. Using either monoclonal antibodies or lectins with a specificity for rat acinar tissue, the beads were used to immunomagnetically label pancreatic digest before magnetic separation. The results showed that both MIMS coated with a lectin and Dynabeads coated with an antibody removed 80% of the acinar contamination with a 70% islet yield. However, the MIMS were significantly less effective with the second enzyme, which produced larger acinar particles. In this study, although the MIMS produced the least nonspecific islet trapping, the Dynabeads coated with Leicester Department of Surgery no. 10 antibody were found to be the most efficient particle for immunomagnetic islet purification.

Characterization of ultrasmall magnetite [correction of paramagnetic magnetite] particles as superparamagnetic contrast agents in MRI.

RATIONALE AND OBJECTIVES: Very small dextran-coated magnetite particles were developed. These particles can be used either as immunospecific contrast agents for MRI by coupling to antibodies or as an interstitial contrast agent. METHODS: The particles were synthesized from iron chloride/dextran solutions. Size was evaluated by electron microscopy and photon correlation spectroscopy. The iron concentration was determined by x-ray spectroscopy. T1 and T2 values as well as relaxivities RI and R2 were evaluated with a clinical MR scanner at 1.5 T. Biocompatibility assays were performed with the cell line U937 in methylcellulose cultures. RESULTS: Superparamagnetic, dextran-coated magnetite particles with a hydrodynamic diameter of 10 nm were developed. The iron core size was 7 nm; R1,7 L/mmol x s; and R2, 19 L/mmol x s. These particles are smaller than those currently available commercially and therefore show a smaller R1 to R2 ratio. Biocompatibility tests have shown no toxic side effects so far. CONCLUSIONS: Ultrasmall magnetite particles with a dextran coating were developed; the physical properties of these particles evaluated in vitro are described in this study.

Attenuation control of ilvBNC in Corynebacterium glutamicum: evidence of leader peptide formation without the presence of a ribosome binding site.

The ilvBNC operon of Corynebacterium glutamicum encodes acetohydroxy acid synthase and isomero-reductase, which are key enzymes of L-isoleucine, L-valine and L-leucine syntheses. In this study we identified the transcript initiation site of ilvBNC operon 292 nucleotides in front of the first structural gene, and detected the formation of a short transcript from the leader region in addition to the full-length transcript of the operon. This identifies the control of ilvBNC transcription by an attenuation mechanism involving antitermination. Mutations in the leader region were made and their effect on the operon expression in ilvB'lacZ fusions was quantified. Although a presumed leader-peptide-coding region is only one nucleotide away from the transcript initiation site determined, there is clear evidence to support the formation of this leader peptide: (i) the substitution of initiation codon ATG of the peptide by AGG reduced lacZ expression of the appropriate fusion construct to 19%; (ii) the replacement of three subsequent Val codons by Ala codons resulted in the loss of Val-dependent expression; and (iii) a leader peptide LacZ fusion resulted in active beta-galactosidase. Based on these results, it is concluded that transcription of ilvBNC is controlled by a translational-coupled attenuation mechanism. The absence of a ribosome binding site for leader peptide formation means that additional mechanisms may contribute to the transcription control at the decoding initiation step in the leader peptide formation.

Ceramide as an activator lipid of cathepsin D.

We have identified the aspartic protease cathepsin D as a novel intracellular target protein for the lipid second messenger ceramide. Ceramide specifically binds to and induces CTSD proteolytic activity. A-SMase deficient cells derived from Niemann-Pick patients show decreased CTSD activity that was reconstituted by transfection with A-SMase cDNA. Ceramide accumulation in cells derived from A-ceramidase defective Farber patients correlates with enhanced CTSD activity. These findings suggest that A-SMase-derived ceramide targets endolysosomal CTSD.