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Biomedical subjects

A Ludwig

Publications and source records attributed to A Ludwig.

At least 109 records · Page 6Linked to original sources
PubMed1995

The block of the expressed L-type calcium channel is modulated by the beta 3 subunit.

The alpha 1C subunit of the L-type calcium channel was stable, expressed alone or in combination with the beta 3 subunit in Chinese hamster ovary cells. The beta 3 subunit enhanced significantly the inactivation of barium currents indicating that both subunits interacted with each other. The beta 3 subunit decreased significantly the half-maximal inhibitory concentration of the calcium channel blockers (-)-gallopamil and verapamil, but did not affect significantly the block caused by isradipine and mibefradil at the holding potentials of -80 mV and -40 mV. These results suggest that the beta 3 subunit affects distinctly the interaction of the expressed alpha 1C subunit with different classes of organic calcium channel blockers.

Animals↗
PubMed1995

[Change in renin activity and blood pressure in the dog autologous kidney transplant model with modified HTK solution].

Investigations of changes in activity of renin and blood pressure after reperfusion of the kidney transplant using HTK solution were carried out by means of an autologous, heterotopic model of kidney transplantation applied to dogs. Duration of cold ischemia was 48 h. According to variations in the composition of the HTK perfusion solution three test groups were set up. During the first 20 min after recirculation in each test group the renal venous and arterial renin activities were measured. Parallel to renin activity, the arterial blood pressure was recorded. During the first few minutes following recirculation of the kidney transplant the renin levels in the venous blood of the kidney were higher in test group 1 (HTK solution, perfusion height 120 cm) than in either of the other two, showing a median maximal increase of 195 ng/ml.h. In test group 2 the maximal venous renin concentration fell to 145 ng/ml.h, while graphs take a more uniform course. Test group 3 (HTK/tryptophan) differed from the others in having further improved renin values. After the 7.5 min of observation normal venous renin concentrations were measured following earlier values for maximal increase between 23.1 ng/ml.h and 120 ng/ml.h (median 61.5 ng/ml.h). The best reperfusion of the kidney was observed in the tryptophan group, albeit without any recognizable positive effects on the other renal functions. Initially low renin values do not necessarily correlate with a smooth postoperative renal function and vice versa. Initial renin values cannot provide a secure basis for predicting instant as well as long-term postoperative functions.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Effects of various HTK solution regimens on proteinuria after renal transplantation in dogs.

Investigations were carried out by means of an autologous, heterotopic model for kidney transplantation applied to dogs. Duration of cold ischemia was 48 h. Four experimental groups were arranged. During the first 20 min following revitalization of the transplanted kidney, group 1 (HTK solution/80 cm perfusion height) showed a significant glomerular and tubular malfunction. In group 2 (HTK solution/120 cm perfusion height), only four urinary proteins with molecular weights of 25 kDa, 67 kDa, 100 kDa and > 100 kDa were found. The excretion of higher molecular proteins receded over the 20-min period of observation. In both group 3 (HTK/aspartate solution) and group 4 (HTK/tryptophan solution) the quantity of excreted glomerular and tubular protein was well above that of group 2. As opposed to the "Tryptophan" group, a complete restoration of renal function was observed in the "Aspartate" group after 4 weeks. In general, the "standard" HTK protective solution delivered with 120 cm perfusion pressure gave the most favorable results, with the lowest levels of proteinuria and a satisfactory recovery of renal function after revitalization.

Albuminuria↗
PubMed1995

Expression of the L-type calcium channel with two different beta subunits and its modulation by Ro 40-5967.

The smooth muscle alpha 1Cb subunit of the L-type calcium channel was expressed alone (CHO alpha 1 cell) or together with the skeletal beta 1 (CHO alpha 1 beta 1 cell) subunit or smooth muscle beta 3 (CHO alpha 1 beta 3 cell) subunit in Chinese hamster ovary (CHO) cells. The interaction of the expressed calcium channel with the non-dihydropyridine calcium channel blocker Ro 40-5967 was studied. Ro 40-5967 decreased isradipine binding by an apparent allosteric interaction and blocked the barium inward currents (IBa) in a voltage- and use-dependent manner in all cells. The steady-state inactivation curves were shifted to hyperpolarizing potentials in the presence of Ro 40-5967. The rate of channel inactivation was increased in CHO alpha 1 and CHO alpha 1 beta 3 cells. The shift in the steady-state inactivation curve and the increase in channel inactivation were less pronounced in CHO alpha 1 beta 1 cells than in the other cell lines. Low concentrations of Ro 40-5967 increased IBa by up to 198% in 33% of the CHO alpha 1 beta 1 cells. In addition, higher concentrations of Ro 40-5967 were required to inhibit IBa in 60% of the CHO alpha 1 beta 3 cells. These results suggest that the beta subunits modify the interaction of the non-dihydropyridine Ro 40-5967 with the expressed calcium channel alpha 1 subunit.

Animals↗
PubMed1995

Evaluation of polysaccharides intended for ophthalmic use in ocular dosage forms.

Different water-soluble polysaccharides were evaluated for their intended use in ocular dosage forms. The physicochemical characteristics and the viscosity of the solutions prepared with several iso-osmotic vehicles were measured. The influence of ions present in lacrimal fluid, mucin and cyclodextrin on the rheological behaviour was examined. Scleroglucan and xanthan gum exhibit viscoelastic and mucoadhesive properties useful for ocular dosage form formulations.

Cyclodextrins↗
PubMed1994

A cytolysin encoded by Salmonella is required for survival within macrophages.

A Salmonella gene encoding a cytolysin has been identified by screening for hemolysis on blood agar. DNA sequence analyses together with genetic mapping in Salmonella suggest that it is unrelated to other toxins or hemolysins. The gene (slyA) is present in every strain of Salmonella examined, in Shigella, and in enteroinvasive Escherichia coli but not in other Enterobacteriaceae. SlyA (salmolysin) purified from a derivative of the original clone has hemolytic and cytolytic activity and has a molecular weight predicted by the DNA sequence. The median lethal dose and infection kinetics in mice suggest that the toxin is required for virulence and facilitates Salmonella survival within mouse peritoneal macrophages.

Amino Acid Sequence↗
PubMed1994

Low temperature cultivation--a step towards process optimisation.

Adherent recombinant BHK cells were cultivated at temperatures between 30 and 37 degrees C. Batch and repeated-batch-cultivations in a 2-litre bioreactor showed a significant influence on metabolism and cell growth. The low-temperature-cultivations showed a lower growth rate and a lower glucose consumption rate and, therefore, less lactate production. On the other hand, the maximum cell density and productivity seemed not to be affected by the temperature reduction.

Animals↗
PubMed1994

Cell electrophoretic discrimination of Staphylococcus saprophyticus strains of different origin.

Staphylococcus saprophyticus isolates (n = 10) from urinary tract infections (UTIs) could be reproducibly distinguished from S. saprophyticus strains from respiratory tract infections (n = 5) by distinct electrophoretic mobility. Thus, the method of "bacteriopheresis" may offer a new approach to discriminate pathogenic microbial strains of different origin with respect to defined surface structures (hemagglutinins, lectins), generating distinct surface charges.

Electrophoresis↗
PubMed1994

Pore formation by the Escherichia coli alpha-hemolysin: role for mediator release from human inflammatory cells.

The Escherichia coli alpha-hemolysin represents a potent stimulus for inflammatory mediator release (O2-, beta-glucuronidase release, and leukotriene generation) from human polymorphonuclear granulocytes, for histamine release from a suspension of human lymphocyte/monocyte basophil cells (LMB), and for serotonin release and 12-hydroxyeicosatetraenoic acid generation from human platelets. In contrast, the E. coli alpha-hemolysin leads to a downregulation of cytokine release (interleukin-1 beta [IL-1 beta], IL-6, and tumor necrosis factor alpha) from human LMB. Recently, it became apparent that the E. coli alpha-hemolysin is composed of several functional structures. We analyzed the role of pore formation, pore stability, and calcium-dependent membrane binding for inflammatory mediator release by using washed bacteria as well as culture supernatants of isogenic recombinant E. coli strains expressing no hemolysin (Hly-), the wild-type hemolysin (Hly+), or hemolysin molecules deficient or modulated in defined functions (pore formation, calcium-dependent membrane binding, or pore stability). In human granulocytes and platelets, mutant hemolysin with enhanced pore stability did not lead to a further increase in induction; mutant hemolysin deficient in pore-forming activity or calcium-dependent membrane binding no longer induced leukotriene B4 generation or beta-glucuronidase release compared with the wild-type hemolysin. Similar results were obtained with regard to histamine release from human LMB. The induction of cytokine release from human LMB differed depending on the type of mutant E. coli alpha-hemolysin. The wild-type hemolysin, the mutant hemolysin with enhanced pore-forming activity, and, to a lesser degree, the mutant hemolysin deficient in pore-forming activity decreased cytokine release (IL-1 beta, IL-6, IL-8, and tumor necrosis factor) compared with untreated cells. In contrast, the mutant hemolysin deficient in calcium-dependent membrane binding led to an increase of up to 50% in cytokine release compared with that by unstimulated cells. Our results indicate that simultaneous expression of the pore-forming and calcium-dependent membrane-binding activities of the hemolysin molecule was necessary to obtain the full cellular inflammatory response pattern observed with the wild-type hemolysin.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1994

Synthesis and secretion of bacterial antigens by attenuated Salmonella via the Escherichia coli hemolysin secretion system.

We describe a plasmid system which allows the secretion of foreign antigens in attenuated Salmonella aroA strains by the secretion apparatus of E. coli hemolysin. The gene (or gene fragment) encoding the antigen is inserted in frame into a residual position of the hlyA gene, encoding the HlyA secretion signal (HlyAs). Generally, the fused gene is efficiently expressed and the synthesized antigen is in part secreted into the culture supernatant and in part exposed on the surface of the producing Salmonella strain. The successful use of this approach is demonstrated with two antigens of Salmonella typhimurium, PagC and SlyA, both of which are potent virulence factors but produced only in small amounts under in vitro culture conditions and two virulence proteins of Listeria monocytogenes, p60 and listeriolysin. Interestingly the listeriolysin fusion protein proved to be cytolytically active and allowed, when expressed in Salmonella, the escape of these bacteria into the cytoplasm of infected macrophages.

Antigens, Bacterial↗
PubMed1993

Oligomerization of Escherichia coli haemolysin (HlyA) is involved in pore formation.

Coexpression of pairs of nonhaemolytic HlyA mutants in the recombination-deficient (recA) strain Escherichia coli HB101 resulted in a partial reconstitution of haemolytic activity, indicating that the mutation in one HlyA molecule can be complemented by the corresponding wild-type sequence in the other mutant HlyA molecule and vice versa. This suggests that two or more HlyA molecules aggregate prior to pore formation. Partial reconstitution of the haemolytic activity was obtained by the combined expression of a nonhaemolytic HlyA derivative containing a deletion of five repeat units in the repeat domain and several nonhaemolytic HlyA mutants affected in the pore-forming hydrophobic region. The simultaneous expression of two inactive mutant HlyA proteins affected in the region at which HlyA is covalently modified by HlyC and the repeat domain, respectively, resulted in a haemolytic phenotype on blood agar plates comparable to that of wild-type haemolysin. However, complementation was not possible between pairs of HlyA molecules containing site-directed mutations in the hydrophobic region and the modification region, respectively. In addition, no complementation was observed between HlyA mutants with specific mutations at different sites of the same functional domain, i.e. within the hydrophobic region, the modification region or the repeat domain. The aggregation of the HlyA molecules appears to take place after secretion, since no extracellular haemolytic activity was detected when a truncated but active HlyA lacking the C-terminal secretion sequence was expressed together with a nonhaemolytic but transport-competent HlyA mutant containing a deletion in the repeat domain.

Bacterial Proteins↗
PubMed1993

Effect of cholesterol addition on growth kinetics and shear stress sensitivity of adherent mammalian cells.

The growth kinetics of adherent baby hamster kidney cells cultivated with additional cholesterol as well as the effect of cholesterol addition on shear stress sensitivity were investigated. The influence of various cholesterol preparations was tested, whereby dimethylsulfoxide and ethanol show negative effects at higher concentrations. With addition of cholesterol in the range of 90 micrograms ml-1, a positive effect on the shear stress resistance was achieved.

Animals↗
PubMed1993

Shear stress induced variation of cell condition and productivity.

Anchorage-dependent recombinant BHK-21 cells, which produce beta-D-galactosidase, were subjected to shear stress levels in the range between 0.1 and 1.55 N m-2 over a period of three days. To investigate cell constitution and productivity, the cell number and viability, as well as the lactate dehydrogenase and beta-D-galactosidase activity, were determined. Low (0.25-0.75 N m-2) and intermediate (0.75-1.25 N m-2) shear stress rates stimulated; higher shear stress rates (more than 1.25 N m-2) diminished productivity. Cell number and general condition were influenced negatively by all but the lowest levels of shear stress. An optimal shear stress level between 0.4 and 0.6 N m-2 can be deduced.

Animals↗
PubMed1993

Age dependency of red blood cell deformability and density: studies in transient erythroblastopenia of childhood.

Several investigators have demonstrated that red blood cell (RBC) deformability decreases progressively with increasing cell density and proposed that reduction in deformability plays a role in the senescence process of normal RBCs. Transient erythroblastopenia of childhood (TEC) results from temporary cessation of erythropoiesis. Since no new RBCs are produced for some time, the circulating RBCs are relatively old. RBS density (phthalate-oil method) and RBC deformability (RBC elongation in a counter-rotating rheoscope) were studied in seven children with TEC and in 10 control children. The mean values of MCHC, RBC density and RBC deformation were not significantly different between TEC and control children. Compared to controls, the frequency distribution of RBC density in TEC was slightly shifted to higher values. The percentage of RBCs with extremely low densities (< 1.090 g/ml) was 0.9 +/- 1.2% in the patients and 5.6 +/- 2.3% in the controls (P < 0.001). The percentage of RBCs with high density (> 1.106 g/ml) was 6.4 +/- 2.1% in the patients and 4.9 +/- 1.8% in the controls (P > 0.10). The reduction of RBCs with low density in TEC suggests that RBCs with low density are relatively young. Since the percentage of RBCs with high density increased only slightly in TEC, we conclude that only a fraction of dense RBCs is old. In TEC, the frequency distribution of RBC elongation was slightly shifted to lower values. 5% of the RBCs studied in the control children had RBC elongation values above 0.39 (TEC 1.2%) and 5% had elongation values below 0.16 (TEC 6.7%). Thus, only a small fraction of highly deformable RBCs was diminished in TEC. These data suggest that a decrease in deformability is not a significant part of the normal ageing process of human RBCs.

Child, Preschool↗
PubMed1993

Stable co-expression of calcium channel alpha 1, beta and alpha 2/delta subunits in a somatic cell line.

1. The high-voltage-activated L-type calcium channel is a multi-protein complex of alpha 1, alpha 2/delta, beta and gamma subunits. The alpha 1 subunit contains the voltage-dependent calcium-conducting pore. Chinese hamster ovary (CHO) cells were stably transfected with the complementary DNA of the alpha 1, beta and alpha 2/delta subunits. These subunits were not detected in wild-type CHO cells. 2. The alpha 1 (CaCh2b) subunit itself directed the expression of functional calcium channels which bound calcium channel blockers and showed voltage-dependent activation and inactivation. 3. The co-expression of the alpha 1 subunit with the beta subunit (CaB1 gene) enhanced the density of the dihydropyridine binding sites 2- to 3-fold and increased dihydropyridine-sensitive barium inward currents (IBa) up to 3.5-fold from -13.3 microA/cm2 (alpha 1 subunit) to -46.7 microA/cm2 (alpha 1 and beta subunits). 4. Co-expression of the beta subunit did not change the sensitivity of IBa towards dihydropyridines, but accelerated current activation and inactivation and shifted the half-maximal steady-state activation and inactivation to slightly more hyperpolarizing potentials. 5. The co-expression of the alpha 2/delta subunit together with alpha 1 and beta subunits accelerated the inactivation kinetics of the channel without a major effect on the other parameters. 6. These results indicate that the beta and alpha 2/delta subunit interact with the alpha 1 subunit and modulate thereby the properties of the alpha 1 subunit-dependent inward current.

Animals↗
PubMed1992

Influence of the temperature on the shear stress sensitivity of adherent BHK 21 cells.

The influence of temperature on the shear sensitivity of anchorage-dependent baby hamster kidney (BHK) cells was investigated. The temperature effect in general was compared for stressed and unstressed cells. Both the growth rate as well as the shear sensitivity are temperature-dependent. Decreasing the temperature lowered the growth rate and increased the ability of the BHK cells to withstand shear stress.

Animals↗