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Biomedical subjects

W Kramer

Publications and source records attributed to W Kramer.

At least 127 records · Page 7Linked to original sources
PubMed1988

Oligonucleotide-directed construction of mutations: a gapped duplex DNA procedure without enzymatic reactions in vitro.

The gapped duplex DNA approach to oligonucleotide-directed construction of mutations (Kramer et al. 1984, Nucl. Acids Res. 12, 9441-9456) has been developed further. A procedure is described that makes in vitro DNA polymerase/DNA ligase reactions dispensable. Direct transfection of host bacteria with gdDNA molecules of recombinant phage M13 plus mutagenic oligonucleotide results in marker yields in excess of 50% (gap size 1640 nucleotides). An important feature incorporated into the mutagenic oligonucleotide is the presence of one or two internucleotidic phosphorothioate linkages immediately adjacent to the 5'-terminus. Automated preparation and biochemical properties of such compounds are described as well as their performance in oligonucleotide-directed mutagenesis. A systematic study of the following parameters influencing marker yield is reported: Gap size, length of oligonucleotide, chemical nature of oligonucleotide termini and heatshock temperature during transformation.

Base Sequence↗
PubMed1988

Direct photoaffinity labelling of binding proteins for beta-lactam antibiotics in rabbit intestinal brush border membranes with [3H]benzylpenicillin.

Brush border membrane vesicles from rabbit small intestine were used to study the intestinal uptake system for beta-lactam antibiotics. Benzylpenicillin inhibited the H+-dependent uptake of alpha-aminocephalosporins in a concentration-dependent manner suggesting a common transport system for alpha-aminocephalosporins and benzylpenicillin. Benzylpenicillin is therefore a suitable probe to characterize this transport system. Irradiation of [3H]benzylpenicillin using light sources having their maximum of radiation at 300 or 254 nm resulted in a covalent incorporation of radioactivity into penicillin binding proteins as was shown with serum albumin. Hence [3H]benzylpenicillin can be used for direct photoaffinity labeling of penicillin binding proteins in different cells and tissues. In brush border membrane vesicles from rabbit small intestine predominantly a membrane polypeptide with an apparent molecular weight of 127,000 was labeled by [3H]benzylpenicillin. Competition labeling experiments demonstrated that beta-lactam antibiotics--penicillins and cephalosporins--specifically interact with this protein, whereas amino acids, sugars or bile acids had no effect on the labeling pattern. Compounds which decreased the labeling of the 127,000 molecular weight membrane polypeptide also inhibited the H+-dependent uptake of the alpha-aminocephalosporin cephalexin into intestinal brush border membrane vesicles. These results suggest that a polypeptide of molecular weight 127,000 in the brush border membrane from rabbit small intestine is a constituent of a common transport system responsible for the uptake of orally effective beta-lactam antibiotics and dipeptides. beta-Lactam antibiotics which are not absorbed from the small intestine also bind from the luminal site to this transport system, but are not transported across the brush border membrane.

Affinity Labels↗
PubMed1988

Characterization of the transport system for beta-lactam antibiotics and dipeptides in rat renal brush-border membrane vesicles by photoaffinity labeling.

The uptake of the alpha-aminocephalosporin cephalexin into brush-border membrane vesicles from rat renal cortex was independent on an inward H+-gradient in contrast to the intestinal transport system. The transport system could be irreversibly inhibited by photoaffinity labeling. Two binding polypeptides for beta-lactam antibiotics and dipeptides with apparent molecular weights 130,000 and 95,000 were identified by photoaffinity labeling with [3H]benzylpenicillin and N-(4-azido[3,5-3H]benzoyl) derivatives of cephalexin and glycyl-L-proline. The uptake of cephalexin and the labeling of the respective binding proteins was inhibited by beta-lactam antibiotics and dipeptides as with intestinal brush-border membranes. These data indicate that the transport systems for beta-lactam antibiotics and dipeptides in the brush-border membrane from rat kidney and small intestine are similar but not identical.

Affinity Labels↗
PubMed1988

Direct photoaffinity labeling of the putative sulfonylurea receptor in rat beta-cell tumor membranes by [3H]glibenclamide.

The oral antidiabetic sulfonylurea [3H]glibenclamide specifically binds to plasma membranes from a rat beta-cell tumor indicating a receptor for sulfonylureas in these membranes. Irradiation of [3H]glibenclamide at 254 or 300 nm in the presence of albumin resulted in covalent labeling of the albumin molecule. Direct photoaffinity labeling of beta-cell membranes with [3H]glibenclamide resulted in the covalent modification of two membrane polypeptides with apparent molecular masses 140 and 33 kDa. The extent of labeling of the 140 kDa polypeptide was specifically decreased by sulfonylureas. This suggests that a membrane polypeptide of 140 kDa is a component of the sulfonylurea receptor in the beta-cell membrane.

ATP-Binding Cassette Transporters↗
PubMed1988

Identification and comparison of bile acid-binding polypeptides in ileal basolateral membrane.

Bile acid-binding polypeptides were examined using basolateral membrane vesicles and enterocytes isolated from rat ileum. The uptake of a photolabile taurocholate derivative, (7,7,-azo-3 alpha,12 alpha-dihydroxy-5 beta[3 beta-3H]cholan-24-oyl)-2- aminoethanesulfonate,7,7-azo-TC, in ileal vesicles preloaded with paraaminohippurate (PAH) was stimulated with respect to uptake in unpreloaded vesicles. The PAH-transstimulated uptake of 7,7-azo-TC was inhibited by taurocholate and vice versa. Irradiation of membrane vesicles in the presence of 7,7-azo-TC irreversibly inhibited PAH-transstimulated taurocholate uptake. Photoaffinity labeling of basolateral membrane vesicles directly with [3H] 7,7-azo-TC and separation of proteins by SDS-PAGE revealed incorporation of radioactivity into several polypeptides. Photoaffinity labeling of vesicles in the presence of taurocholate inhibited the labeling of 54,000 and 59,000 mol. wt. polypeptides. The efflux of taurocholate from ileal enterocytes was cis-inhibited by 7,7-azo-TC and transstimulated by PAH. Irradiation of enterocytes in the presence of 7,7-azo-TC inhibited taurocholate efflux greater than the presence of 7,7-azo-TC in the dark. When enterocytes that were irradiated in the presence of [3H] 7,7-azo-TC were fractionated and the resultant basolateral membrane fraction was subjected to SDS-PAGE, incorporation of radioactivity into the 54,000 and 59,000 mol. wt. polypeptides was seen. In contrast, when the brush-border membrane fraction was subjected to SDS-PAGE, greatest incorporation of radioactivity was seen in the previously described 99,000 mol. wt. polypeptide. These studies suggest that 7,7-azo-TC shared transporters with natural bile acid and identified polypeptides that may be involved in bile acid transport across the basolateral membrane and differ from that seen in the brush-border membrane of the ileal epithelial cell.

Affinity Labels↗
PubMed1988

[Conventional x-ray diagnosis of the knee joint following surgical repair of ligament injuries and after ligament replacement plasty].

Characteristic radiological findings following operative repair of the ligaments of the knee joint are described and related to the type of operative procedure. The operations depend on a number of basic principles which are discussed in detail. The majority of these surgical procedures are not apparent radiographically. It is only by having a knowledge of the individual procedures and of the resulting functional changes in the knee joint, that the informed radiologist can make a correct assessment of the post-operative radiographs. Defects resulting from bone drilling can be recognised tomographically in the early postoperative phase, or if they persist for any reason. This could be confirmed by experiments on pigs' knees.

Adolescent↗
PubMed1988

Analysis of left-ventricular changes associated with chronic hemodialysis. A noninvasive follow-up study.

To assess the reasons for the frequent cardiovascular complications in patients with end-stage renal disease (ESRD), 61 out of 131 normotensive ESRD patients originally examined (mean ESRD duration: 71 +/- 41 months) were followed over 2.5 years by echo-, electro- and mechanocardiography. Clinical and biochemical parameters were comparable. The prevalence of pericardial effusion (3%), pericardial thickening (14%), aortic valve sclerosis (14%) and mitral valve anulus sclerosis (12%) was unchanged. The interventricular septum diameter (14.3 +/- 3.0 vs. 16.4 +/- 3.4 mm), index of left-ventricular (LV) wall asymmetry (1.25 +/- 0.30 vs. 1.52 +/- 0.36) and left atrial diameter (38.3 +/- 5.4 vs. 42.6 +/- 3 mm) increased (p less than 0.001). The LV end-systolic diameter decreased slightly (35.8 + 6.3 vs. 34.2 +/- 6.4 mm; p less than 0.05), with no significant changes for end-diastolic diameter (50.4 +/- 6.3 vs. 49.3 +/- 6.1 mm), muscle mass index (189 +/- 57 vs. 197 +/- 50 g/m2), stroke volume (86.1 +/- 26.2 vs. 85.7 +/- 26.7 7 ml/m2) and fractional shortening (29.1 +/- 7 vs. 30.8 +/- 8.6%). We conclude that the predominant finding in ESRD is an LV hypertrophy progressing towards an asymmetric septum hypertrophy, while the increase of the primarily enlarged left atrial diameter over 30 months reflects a further deterioration of the diastolic LV dysfunction.

Cardiomegaly↗
PubMed1988

[Cardiac and extracardiac parameters as principles in differential therapeutic considerations for decreasing pre- and afterload].

A differential therapy of congestive heart failure requires the identification of the different stages of myocardial damage consequent to the underlying heart disease and the individual adaptive response to it. Because of their different load dependence, it is important to recognize the independent contribution of systolic and diastolic dysfunction in congestive heart failure. The vertical displacement of the diastolic pressure-volume loop is associated with a reduced therapeutic range. Reducing preload may limit the degree of diastolic filling necessary to fill a noncompliant heart and limit active myocardial stretch. The final outcome of any disorder is either to comprise cardiac filling, emptying, or both. When the end-stage is reached, only modification of excessive neurohormonal responses to heart failure remains. Efforts are necessary to avoid substances able to support the maladaptation. The rational use of vasodilator therapy in congestive heart failure is based on the concept that peripheral circulatory mechanisms overshoot, and may thereby act to exacerbate the heart failure. As important as it is to modify late neurohormonal responses, it is even more important to intervene at an earlier stage to reduce the degree of irreversible myocardial damage. Further studies have to prove the potential benefits of ACE-inhibitors in patients with mild heart failure in order to prevent or delay the progress of the disease.

Blood Pressure↗
PubMed1987

Identification of identical binding polypeptides for cephalosporins and dipeptides in intestinal brush-border membrane vesicles by photoaffinity labeling.

The uptake of a photolabile derivative of the orally effective cephalosporin cephalexin, N-(4-azidobenzoyl)cephalexin, was investigated in brush-border membrane vesicles. The compound was taken up into the intravesicular space and inhibited the active uptake of cephalexin in a concentration-dependent manner. Therefore, this probe interacts with the transport system shared by alpha-aminocephalosporins and dipeptides. Photoaffinity labeling of brush-border membrane vesicles from rat small intestine with N-(4-azido[3,5-3H]benzoyl) derivatives of the cephalosporin cephalexin and the dipeptide glycyl-L-proline resulted in the covalent incorporation of radioactivity into membrane polypeptides with apparent molecular weights of 127,000, 100,000, 94,000 and 86,000, the polypeptide of molecular weight 127,000 being predominantly labeled. The specificity of labeling was demonstrated by a decrease in the labeling of the polypeptide of apparent molecular weight 127,000 in the presence of beta-lactam antibiotics and dipeptides, whereas glucose, taurocholate or amino acids had no effect on the labeling pattern. These data demonstrate an interaction of cephalosporins and dipeptides with a common membrane protein of molecular weight 127,000, which could be a component of the intestinal transport system(s) responsible for the uptake of orally effective cephalosporins and dipeptides.

Affinity Labels↗
PubMed1987

Photoaffinity labeling studies of the rat renal sodium bile salt cotransport system.

The uptake of a photolabile taurocholate derivative, (7,7-azo-3 alpha, 12 alpha-dihydroxy-5 beta-cholan-24-oyl)-2-aminoethanesulfonate, 7,7-azo-TC, into rat renal brush-border membrane vesicles was stimulated by Na+ and inhibited by taurocholate indicating an interaction with the Na+/bile salt cotransport system. Irradiation of membrane vesicles in the presence of 7,7-azo-TC inhibited Na+-dependent taurocholate uptake irreversibly. Photoaffinity labeling with [3H]7,7-azo-TC resulted in a predominant incorporation of radioactivity into a polypeptide with apparent molecular weight of 99,000. These results suggest that the proteins involved in Na+/bile salt cotransport are similar in renal and ileal brush-border membranes, but differ from those in hepatocytes.

Affinity Labels↗
PubMed1987

[Results of the treatment of pathologic fractures of long tubular bones].

The treatment of pathological bone fractures will be analysed by 53 patients of the Surgical Clinic of the University of Tübingen in view of fractures reason, operative providing and prognosis. The first aim consist in correction or preservation of live quality. Fractures nearly joints can be treated by endoprothetics. With the help of compound osteosynthesis most fractures can be stabilized. Conservative treatment only in final phase is legitimated.

Adult↗
PubMed1987

[Determination of active components of amrinone by analysis of pressure-volume results; use of the conductance (volume) catheter technic and rapid load change by balloon occlusion of the inferior vena cava].

Endsystolic pressure-volume relationships (ESPVR) were determined using the conductance (volume) catheter-technique and the required rapid load changes by applying vasoactive drugs (nitroprusside = NP; phenylephrine = PE) or brief preload reduction by balloon occlusion of the vena cava inferior (BOVCI). With this load-independent index of contractility, we analyzed the hemodynamically active components of amrinone (AM) over a range of different LV-loading conditions. In 19 patients (study I) with still normal LV-function (LVF) in group A (dP/dtmax 1585 +/- 386 mmHg/s; n = 10) and impaired LVF in group B (dP/dtmax 1044 +/- 164 mmHg/s; n = 9) an infusion of AM, 1.5 mg/kg over 2 min, failed to induce changes in ESPVR (p greater than 0.05), but rather caused load changes, resembling like those seen with NP. During continuously paced heart rates (90/min) brief infusion phases with NP and PE provided the necessary load changes. In 11 patients (study II) with impaired LVF (dP/dtmax = 1177 +/- 163 mmHg/s) 2.5 mg/kg AM i.v. induced an increase in contractility, which was more pronounced after the additional application of dobutamine (DOB) at 10 micrograms/kg/min.; (1) Slope k rose from 0.52 to 0.80 mmHg/ml; (2) dP/dtmax increased by an average of 39% and 57% (p less than 0.01), respectively. The BOVCI provided the necessary load changes. Thus AM demonstrates its contractility-increasing effects in a dose-related fashion, probably (at least in part) via an increase in heart rate; inotropic effects further increased with the combination of AM and DOB. The use of the conductance technique for the assessment of ESPVR during acute decrease of preload by temporary BOVCI is innocuous to the patient, reproducible and can be carried out under the conditions of a routine cardiac catheter procedure. This technique seems to be useful in the assessment of relative inotropic effects of the newer cardiotonic drugs.

Amrinone↗