[Micromorphometry and stereography of rough depth relief coated implant surfaces by scanning electron microscopy and confocal laser scanning microscopy].
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Coated Materials, Biocompatible↗
Biomedical subjects
H Götz
Publications and source records attributed to H Götz.
[Different effects of various embedding techniques on confocal laser scanning microscopy image and mechanical properties of cortical bone].
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Animals↗
Fluoride-induced precipitates on enamel surface and subsurface areas visualised by electron microscopy and confocal laser scanning microscopy.
The present study examined the enamel surface after in vitro topical treatments with a neutral 2% NaF solution. For minimising the risk of artefacts, samples were inspected without pre-treatment as fresh, naturally wet specimens by complementary techniques: variable pressure electron microscopy (VP-SEM) and confocal laser scanning microscopy (CLSM). VP-SEM provided information on the surface morphology, whereas CLSM allowed non-destructive visualisation of subsurface areas. Neutral NaF solutions induced globular precipitates on the enamel surfaces. If the globules formed may be described as "calcium fluoride like material", the additional information of this experiment is that, after interaction with neutral solutions, they also contain considerable amounts of NaF. When the NaF solutions were soaked on pellets and then were brought into contact with the enamel surfaces, NaF crystallites of cubic shape are formed. Confocal optical tomographies of subsurface enamel after treatments with neutral NaF solutions revealed partly coroded enamel structures, whereas VP-SEM showed intact surfaces. In between the coroded areas, a fine granulate precipitate could be observed. This is evidence that fluoride induces the formation of sub-surface precipitates only when applied during demineralisation. The precipitate could be readily removed by 24-h contact with a KOH solution.
Adolescent↗
Growth reduction of Listeria spp. caused by undefined industrial red smear cheese cultures and bacteriocin-producing Brevibacterium lines as evaluated in situ on soft cheese.
The undefined microbial floras derived from the surface of ripe cheese which are used for the ripening of commercial red smear cheeses have a strong impact on the growth of Listeria spp. In some cases, these microbial consortia inhibit Listeria almost completely. From such undefined industrial cheese-ripening floras, linocin M18-producing (lin+) (N. Valdés-Stauber and S. Scherer, Appl. Environ. Microbiol. 60:3809-3814, 1994) and -nonproducing Brevibacterium linens strains were isolated and used as single-strain starter cultures on model red smear cheeses to evaluate their potential inhibitory effects on Listeria strains in situ. On cheeses ripened with lin+ strains, a growth reduction of L. ivanovii and L. monocytogenes of 1 to 2 log units was observed compared to cheeses ripened with lin strains. Linocin M18 activity was detected in cheeses ripened with lin+ strains but was not found in those ripened with lin strains. We suggest that production of linocin M18 contributes to the growth reduction of Listeria observed on model red smear cheeses but is unsufficient to explain the almost complete inhibition of Listeria caused by some undefined microbial floras derived from the surface of ripe cheeses.
Bacteriocins↗
Monosomy 6 in human cultured fibroblast-like cells after long-term stimulation with acidic fibroblast growth factor (FGF1).
Long-term exposure to fibroblast growth factor type 1 (FGF1) of fibroblast-like cells derived from neurofibromas of patients with neurofibromatosis type 1, from angiofibromas of patients with tuberous sclerosis, and from foreskin of unaffected donors resulted in the outgrowth of monosomy 6 in 7 out of 14 cell lines examined. After their initial detection by cytogenetic analysis, the proportion of cells which had lost one chromosome 6 was monitored by FISH using a-satellite probes specific for chromosome 6 and 7, and by PCR analysis of polymorphic microsatellite markers. Monosomy 6 exceeding baseline levels developed only in cultures exposed to FGF 1, and the emergence of monosomic cells could not be correlated with a given donor's genotype. During serial culture, the proportion of monosomic cells increased to over 90% in 5 of the 7 affected strains. A conspicuous change of cellular morphology from spindle-shaped to more epithelial-type cells was noted in monosomic cultures, even though none of them converted to a permanent cell line during the observation period. We conclude that long-term exposure of human fibroblast-like cell strains to FGF1 results in the emergence of monosomy 6 in 50% of the cultures so treated. A selective advantage for such monosomic cells is the most likely explanation for their steady increase during serial culture.
Adult↗
Studies on human porin: XIII. The type-1 VDAC 'porin 31HL' biotinylated at the plasmalemma of trypan blue excluding human B lymphocytes.
In 1989, we demonstrated for the first time the expression of the VDAC 'Porin 31HL' in the plasmalemma of human B lymphocytes, then giving first evidence of a multi-topological expression of VDAC. Meanwhile, data from this and other laboratories support our proposal of a multi-compartment distribution of the channel in mammalian tissues. Here, by biotinylation of plasmalemma-integrated proteins of proven living B lymphocytes, followed by two-dimensional electrophoresis, immuno- and streptavidin affinity blotting, we show that part of the channel molecules can be labelled at the outer membrane of the cells. Thus, by a relevant approach our results invalidate objections concerning putative cross-reactivity of anti-human Type-1 porin antibodies with non-VDAC proteins at the outer cell membrane.
B-Lymphocytes↗
Purification and characterization of two putative HLA class II associated proteins: PHAPI and PHAPII.
In addition to their well defined role in presentation of processed antigen on the cell surface, class II molecules are able to transduce signals into the cell after binding of ligands. The cytoplasmic regions of class II molecules might function as docking sites for as yet unidentified proteins that are components of this signalling pathway. Here we report on two putative HLA class II associated proteins (PHAPI and PHAPII) which have been purified from the cytosolic fraction of the human lymphoblastoid B-cell line H2LCL using an affinity matrix composed of the synthetic biotinylated cytoplasmic region of the DR2 alpha chain immobilized on avidin agarose. The sequence obtained for PHAPI revealed a novel primary structure with a leucine/isoleucine rich N-terminal region. Protein data and the cDNA sequence obtained for PHAPII agree with the cDNA sequence of SET that has been described recently. Both PHAPI and PHAPII have an extended highly acidic C-terminal region. Based on their primary structure we speculate that PHAPI and PHAPII are involved in the generation of intracellular signalling events that lead to regulation of transcriptional activity after binding of a ligand to HLA class II molecules.
Amino Acid Sequence↗
Human protein NEFA, a novel DNA binding/EF-hand/leucine zipper protein. Molecular cloning and sequence analysis of the cDNA, isolation and characterization of the protein.
The cDNA libraries constructed from the human acute lymphoblastic leukemia cell line KM3 in the expression vector lambda gt11, were screened with the anti-CALLA (common acute lymphoblastic leukemia antigen) mAb (monoclonal antibody) J5. The selected J5-positive clone I containing a partial cDNA insert was isolated and sequenced. For completing the cDNA sequence the cDNA libraries were further screened by hybridization with the DIG (digoxigenin)-labelled DNA probe derived from clone I, the 5'-end region was analysed by 5'-RACE (rapid amplification of cDNA ends) using a sequence specific primer. In total a 1639 bp cDNA sequence was determined. The cDNA sequence contains a 1260 bp open reading frame and the untranslated 3'- and 5'-end sides. The 420 residue amino acid sequence, deduced from the cDNA sequence, unexpectedly differs fundamentally from CALLA (CD10) although clones I and II were J5-positive in immuno screening. The mature protein corresponding to the cDNA was isolated and characterized from the KM3 cells using polyclonal antisera raised against the in vitro expressed polypeptide from clone I. The protein is expressed on plasma membrane, in cytosol and is secreted into culture medium, its relative molecular mass was determined to be 55 kDa on SDS-PAGE. The deduced amino acid sequence from cDNA was confirmed by peptide sequences. The new protein contains a basic amino acid rich putative DNA binding domain (b) with a potential nuclear targeting signal, two helix-loop-helix (HLH) motif regions, concurrently EF-hand motifs, an acidic amino acid rich region (a) between the EF-hands, and a leucine zipper (Z) motif. This DNA binding protein therefore is characterized by a linked motif "b/HLH/a/HLH/Z". The protein was designated NEFA: DNA binding/EF-hand/acidic amino acid rich region.
Amino Acid Sequence↗
Studies on human porin. VII. The channel properties of the human B-lymphocyte membrane-derived "Porin 31HL" are similar to those of mitochondrial porins.
Porin 31HL was isolated and purified from total membrane preparations of a human B-lymphocyte cell line. The protein showed a single band of apparent molecular mass 31 kDa on SDS-PAGE. Reconstitution of the protein into artificial lipid bilayer membranes defines its function as a channel-forming protein. The distribution of single-channel conductances had two maxima of 2.4 and 4.3 nS in 1M KCl. The channel formed by Porin 31HL of human B-lymphocytes was found to be voltage-dependent and switched to ion-permeable substates at membrane voltage larger than 20mV. In the open state the pore exhibited the characteristics of a general diffusion pore because the mobility sequence of the ions inside the pore was similar to that in the bulk aqueous phase. The effective diameter was estimated to be about 1.7 nm. The properties of the low conductance state of the channel were studied in detail. In this state the pore favored the passage of cations, in contrast to the open state which favored anions slightly. Monoclonal antibodies against the N-terminal end of Porin 31HL blocked its reconstitution but had otherwise no influence on the channel properties. This result suggested that the amphipathic alpha-helical structure at the N-terminal end is probably not involved in channel gating. The channel-forming properties of Porin 31HL were compared to those of porins isolated from mitochondrial outer membranes and to those of the "maxi chloride channel" observed in the cytoplasmic membrane of several eukaryotic cells.
Animals↗
Studies on human porin. VIII. Expression of "Porin 31HL" channels in the plasmalemma of the acute-lymphoblastic-leukemia cell line KM3 as revealed by light- and electron-microscopy.
First electron microscopy data on the expression of the VDAC "Porin 31HL" in the plasmalemma of a eucaryotic cell are presented. In a light and electron microscopic study we demonstrate the expression of the porin channel in the outer cell membrane of the pre-B lymphocyte type acute-lymphoblastic-leukemia cell line KM3. Monoclonal mouse anti-"Porin 31HL" antibodies were applied in indirect immunofluorescence or immunogold labelling experiments. The results confirm our early topological data on the expression of porin channels in the cytoplasmic membrane of different human cell types as revealed on the light microscopy level. Interestingly, only a pre-embedding immunoreaction approach was successful for gold particle labelling of the plasmalemma. This again is in agreement with our recent data on the accessibility of the acetylated N-terminal part of "Porin 31HL" molecules on the outer cell surface.
Antibodies, Monoclonal↗
Antagonistic effect of coryneform bacteria from red smear cheese against Listeria species.
A total of 187 coryneform bacteria were isolated from red smear and screened for inhibitory effects against 16 strains of Listeria species. Culture filtrates from Brevibacterium linens (16 strains), Arthrobacter nicotianae (4 strains) and Arthrobacter nucleogenes (3 strains) showed clear zones of inhibition. The antagonistic effect was seen against 26 to 87% of 91 Listeria strains tested. A. nicotianae and A. nucleogenes were more effective against Listeria innocua and Listeria ivanovii than against Listeria monocytogenes. No species specifically was observed for B. linens, but there was a difference regarding the inhibitory activity of individual culture filtrates. When culture filtrates of the test strains were added to Listeria broth cultures, the maximum growth level was not attained. Inhibition in broth cultures was dependent on the concentration of culture filtrates. Culture filtrates from the late stationary phase had a stronger inhibitory effect on the growth of L. monocytogenes. The nature of the inhibitory effects remained unclear. Attempts to characterize the nature of the antagonism showed that the culture filtrates lost their inhibitory activity upon heating, and the molecular size of the inhibitory substances were greater than 12-14 kDa.
Actinomycetales↗
Quantitation of soluble HLA class II molecules by an enzyme-linked immunosorbent assay.
In order to quantify soluble HLA-DR,DQ,DP molecules (sHLA-RQP) an enzyme-linked immunosorbent assay was developed utilizing two monoclonal antibodies specific for HLA-DR,DP (Tü35) and HLA-DQ (Tü22) gene products, respectively. Highly purified HLA class II molecules isolated from a lymphoblastoid cell line were used for calculation of exact sHLA-RQP protein values. Circadian variations of sHLA-RQP plasma levels were studied in 7 healthy probands showing no significant deviations; measurements in 4 probands at intervals between 4 and 6 weeks revealed that sHLA-RQP levels remain relatively stable. The population analysis of 209 unrelated, HLA-typed healthy donors resulted in an average protein concentration of 1.53 +/- 2.44 micrograms/ml plasma for sHLA-RQP. Four out of 209 probands (= 1.9%) had no detectable sHLA-RQP. Significant associations of high or low sHLA-RQP levels to particular HLA-DR or -DQ specificities were not observed. However, plasma derived from HLA-DR9 positive had the highest and from HLA-DR8 positive donors the lowest mean sHLA-RQP values. By comparing HLA identical with two-haplotype-different siblings we found no evidence that sHLA-RQP plasma levels are under genetic control of the HLA complex or closely linked genes. Furthermore, soluble HLA class I plasma concentrations in 100 probands analyzed showed no correlation to those of sHLA-RQP.
Antibodies, Monoclonal↗
Studies on human porin. VI. Production and characterization of eight monoclonal mouse antibodies against the human VDAC "Porin 31HL" and their application for histotopological studies in human skeletal muscle.
We report on the production and characterization of eight monoclonal mouse antibodies against the complete human VDAC "Porin 31HL". The antigen used was purified from a total membrane preparation of the transformed human B-lymphocyte cell line H2LCL. In Western blots all eight mAbs react with a single 31-kDa band in solubilized H2LCL membrane preparations thus demonstrating their specificity for the human VDAC "Porin 31HL". Concerning the epitope specificity we show that all eight mAbs equally react with the N-terminal part of human porin. Moreover, we demonstrate the expression of VDAC in the sarcolemma by indirect immunoenzyme labelling of cryosections of human skeletal muscle applying four of our mAbs. These data support our recent observations on the expression of porin channels in the plasmalemma of different normal and transformed human cell lines. VDAC in the plasmalemma is discussed as the molecular basis of the Blatz and Magleby channel.
Animals↗
[Studies on human porin. V. The expression of "porin31HL" in the plasmalemma is not by cell transformation].
In recent papers we proved "Porin 31HL" to be located on the surface of human, EBV-transformed B lymphocytes. Here we present proof of "Porin 31HL" in the plasmalemma of normal human blood lymphocytes. For this purpose B and T lymphocytes were isolated from human heparinized blood and examined by indirect immunofluorescence techniques using different monoclonal antibodies against purified "Porin 31HL" and some B and T cell markers, respectively. For comparison a number of established cell lines of different origin were employed. Hence it followed that normal B and T cells as well as transformed and leukemic cells express "Porin 31HL" in their membrane. No significant quantitative differences could be seen. Consequently, the location of "Porin 31HL" in the plasmalemma is not a product of transformation.
Animals↗
[Regional or general anesthesia in patients with pulmonary risks?].
Such partial functions of the respiratory system as ventilation, gas exchange, bronchomotor tone, respiratory regulation, secretion within the airways, mucociliary clearance, etc., are less impaired by such procedures as peridural or spinal anaesthesia than by general anaesthesia. As a result, it is often concluded that regional anaesthesia should always be used preferentially in the case of patients with a pulmonary risk. However, comparative studies have shown that pulmonary morbidity and mortality are little influenced by the anesthetic procedure. For, in the first instance, the incidence of post-operative complications affecting respiration are determined by previous lung pathologies, the age of the patient, and the location and duration of the surgical procedure. These are factors that cannot readily be influenced by anaesthesiological measures. For this reason, the pre-operative diagnostic evaluation and preparation, post-operative prophylaxis and treatment, including the components pain elimination, physiotherapy, respiratory therapy, broncholysis and secretolysis, are more important than the anaesthetic procedure itself.
Anesthesia, Conduction↗
[The mechanism of the reduction of protein catabolism following trauma and during sepsis using xylitol].
Total parenteral nutrition (TPN) after trauma and sepsis has two major goals. One is the reduction of enhanced protein catabolism; the second is the avoidance of enhancement of whole-body glucose turnover. Glucose and xylitol differ in their quantitative utilization rate after trauma and sepsis. Maximal glucose utilization is reduced during such states, while the utilization of xylitol is more than doubled. In order to investigate whether these differences are associated with beneficial effects with regard to whole-body glucose turnover rate of gluconeogenesis and protein sparing, we conducted two studies using animal models and two clinical studies. METHODS. For the determination of glucose and protein turnover, radioactive and stable isotope techniques were applied. In an animal model a primed constant infusion of 3-H-6-glucose, 14-C-1-alanine and 13-C-3-alanine and 14-C-U-acetate was used to determine total glucose appearance, gluconeogenesis from 3-C-precursors and alanine flux. In the human studies hepatic glucose production was determined by using a primed constant infusion of 6.6-D-2-glucose and urea synthesis rate was determined by a primed constant infusion of 2-N-15-urea. RESULTS. In the first rat model we were able to show that hypocaloric xylitol compared to glucose significantly reduced whole-body glucose turnover from 1741 +/- 232 mumol/h during glucose infusion to 449 +/- 49 mumol/h during xylitol infusion and gluconeogenesis from C-3 carbons form 382 +/- 24 mumol/h during glucose infusion to 155 +/- 39 mumol/h during xylitol infusion after a burn trauma. In a second septic rat model the exchange of glucose calories by xylitol in a proportion of 1:1 was associated with a significantly ameliorated N-balance from +144 +/- 90 mgN/kg body weight per day during glucose infusion to +699 +/- 80 mgN/kg body weight per day during glucose-xylitol infusion and a reduced 3-methyl-histidine excretion from 7.14 +/- 0.61 mumol/kg body wt. per day during glucose infusion to 4.10 +/- 0.56 mumol/kg per day during glucose-xylitol infusion, respectively. In two studies with surgical intensive care patients we were able to confirm the nitrogen-sparing properties of xylitol infusion, together with amino acids during hypocaloric feeding or during TPN with a glucose/xylitol mixture in a proportion of 1:1. From a basal urea production rate of 9.2 +/- 1.6 mumol/kg min. xylitol led to a significant reduction with 6.4 +/- 1.5 mumol/kg per min. Hepatic glucose production was significantly reduced during xylitol infusion from basal 4.8 +/- 0.6 mg/kg per min to 3.1 +/- 0.7 mg/kg per min, respectively. Equicaloric glucose in a dosage of 3 g/kg per day had no effect. During TPN glucose/xylitol, in a proportion of 1:1 at a total dosage of 0.24 g/kg per h, significantly reduced whole-body glucose turnover, endogenous glucose production and lactate concentrations compared to an isocaloric glucose infusion. DISCUSSION. In animal as well as in human studies hypocaloric xylitol as well as a glucose-xylitol mixture were more efficient in preserving body protein than glucose alone. Whole-body glucose turnover was significantly reduced during hypocaloric xylitol or glucose-xylitol infusion compared to isocaloric glucose infusion. During the acute phase after trauma we therefore recommend a carbohydrate supplementation of 3 g/kg body wt. per day using xylitol. During long-term TPN, a glucose-xylitol mixture in a proportion of 1:1 in a dosage of 3 g/kg body wt. per day each is recommended as energy source, together with amino acids and, if necessary, lipids.
Animals↗
[The effect of a thigh tourniquet on the pharmacokinetics of midazolam].
Although the general pharmacokinetics of midazolam (M) are well documented, little is known about the possible effects of a thigh tourniquet on the distribution and elimination of this drug. METHOD. Institutional approval for the study and individual informed consent were obtained. We studied 30 patients (ASA-I) without premedication who electively underwent a surgical procedure of the lower limb. Patients were divided into three groups of 10. The procedure was done in groups I and II with and in group III without tourniquet use. Anesthesia was induced in groups I and II with 0.1 mg/kg M, fentanyl 5 micrograms/kg, alcuronium-dichloride 0.15 mg/kg and etomidate 0.1-0.2 mg/kg i.v. and maintained with enflurane 0.3-1.0 vol.-%. About 20 min after midazolam injection and after exsanguination the tourniquet was applied on the proximal thigh in group I. In group II anesthesia was induced with etomidate 0.2 mg/kg and alcuronium-dichloride 0.15 mg/kg i.v., and maintained about 20 min with enflurane 1.0-1.5 vol.-% until exsanguination and tourniquet application. After this, these patients also received 0.1 mg/kg M and 5 micrograms/kg fentanyl i.v. Through an indwelling arterial line, blood samples were obtained prior to and 2, 15, 30, 45, 60, 75, 90, 105, 120, 135, 150, 165 and 180 min after M injection. Plasma M levels were measured by high-performance liquid chromatography with UV detection. These concentrations were fitted to a two-compartment open model. Comparison between groups was performed using the Kruskal-Wallis test and p less than 0.05 was considered to indicate significance. RESULTS. The groups were all comparable in age and weight, and groups I and II also in duration of thigh ischemia. Midazolam elimination half-time (t beta 1/2) was significantly shorter in group II than in groups III and I (52 min vs 126 min and 139 min; p less than 0.05). Of the calculated distribution volumes (volume of the central compartment, volume in the steady state and volume in the elimination phase), only the volume in the steady state was significantly smaller in group II than in groups III and I (p less than 0.05). Groups III and I did not differ significantly in the computed parameters. The measured initial midazolam mean concentrations in group II were twice those in groups III and I (655 ng/ml vs 323 ng/ml and 332 ng/ml). Since clearance was not significantly different between any two groups, the shorter t beta 1/2 in group II was probably due to the reduced distribution volume. CONCLUSION. These data demonstrate that in the presence of a thigh tourniquet the timing of the injection - before or after application of the tourniquet is of decisive importance. Injection after the application of a tourniquet leads to an higher plasma level and shortens the elimination half-life.
Adult↗
[Dosage equipment, concentration monitoring and the anesthesia system].
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Anesthesiology↗