Biomedical subjects

U Ziegler

Publications and source records attributed to U Ziegler.

At least 19 recordsLinked to original sources

PubMed2006

Ifosfamide metabolites CAA, 4-OH-Ifo and Ifo-mustard reduce apical phosphate transport by changing NaPi-IIa in OK cells.

Renal Fanconi syndrome occurs in about 1-5% of all children treated with Ifosfamide (Ifo) and impairment of renal phosphate reabsorption in about 20-30% of them. Pathophysiological mechanisms of Ifo-induced nephropathy are ill defined. The aim has been to investigate whether Ifo metabolites affect the type IIa sodium-dependent phosphate transporter (NaPi-IIa) in viable opossum kidney cells. Ifo did not influence viability of cells or NaPi-IIa-mediated transport up to 1 mM/24 h. Incubation of confluent cells with chloroacetaldehyde (CAA) and 4-hydroperoxyIfosfamide (4-OH-Ifo) led to cell death by necrosis in a concentration-dependent manner. At low concentrations (50-100 microM/24 h), cell viability was normal but apical phosphate transport, NaPi-IIa protein, and -mRNA expression were significantly reduced. Coincubation with sodium-2-mercaptoethanesulfonate (MESNA) prevented the inhibitory action of CAA but not of 4-OH-Ifo; DiMESNA had no effect. Incubation with Ifosfamide-mustard (Ifo-mustard) did alter cell viability at concentrations above 500 microM/24 h. At lower concentrations (50-100 microM/24 h), it led to significant reduction in phosphate transport, NaPi-IIa protein, and mRNA expression. MESNA did not block these effects. The effect of Ifo-mustard was due to internalization of NaPi-IIa. Cyclophosphamide-mustard (CyP-mustard) did not have any influence on cell survival up to 1000 microM, but the inhibitory effect on phosphate transport and on NaPi-IIa protein was the same as found after Ifo-mustard. In conclusion, CAA, 4-OH-Ifo, and Ifo- and CyP-mustard are able to inhibit sodium-dependent phosphate cotransport in viable opossum kidney cells. The Ifo-mustard effect took place via internalization and reduction of de novo synthesis of NaPi-IIa. Therefore, it is possible that Ifo-mustard plays an important role in pathogenesis of Ifo-induced nephropathy.

Acetaldehyde↗

PubMed2006

Impact of carbon dioxide versus air pneumoperitoneum on peritoneal cell migration and cell fate.

BACKGROUND: Postoperative systemic immune function is suppressed after open abdominal surgery, as compared with that after minimally invasive abdominal surgery. As a first line of defense, peritoneal macrophages (PMo) and polymorphonuclear neutrophil granulocytes (PMNs) are of primary importance in protecting the body from microorganisms. Previous studies have shown changes in these cell populations over time after open versus laparoscopic surgery. This study aimed to investigate the dynamics of cell recruitment and clearance of peritoneal cells. METHODS: Female NMRI mice (33 +/- 2 g) were randomly assigned to carbon dioxide (CO2) or air insufflation. Intravasal cells with phagocytic capabilities were selectively stained by intravenous injection of the fluorescent dye PKH26 24 h before surgery. Gas was insufflated into the peritoneal cavity through a catheter, and the pneumoperitoneum was maintained for 30 min. Peritoneal lavage was performed 1, 3, 8, or 24 h after surgery. Apoptotic cells were assessed by flow cytometry using a general caspase substrate. RESULTS: The total peritoneal cell count did not differ between groups. The PKH26-positive PMo level was significantly increased after CO2, as compared with air, at 1 h and 24 h. The ratio of apoptotic PMo did not differ between the groups. In the peritoneal lavage, polymorphonuclear leukocytes (PMNs) were tripled in the air group, as compared with the CO2 group, whereas the ratio of apoptotic PMNs was significantly decreased. There was a higher fraction of PKH26-positive PMNs after air exposure, as compared with that after CO2. CONCLUSIONS: Air exposure triggered a higher transmigration rate of PMNs from the blood compartment into the peritoneal cavity and decreased PMN apoptosis, as compared with CO2. The lower proportion of PKH26-positive peritoneal macrophages in the air group might have been attributable to a higher inflammatory stimulation than in the CO2 group, leading to increased emigration of PMo to draining lymph nodes. All the findings underscore a complex cell-specific regulation of cell recruitment and clearance in the peritoneal compartment.

Air↗

PubMed2006

Expression of smoothelin in the normal and the overactive human bladder.

PURPOSE: We established the expression pattern of smoothelin, a marker protein for contractile smooth muscle cells, in the human detrusor and investigated its possible impact on bladder overactivity. MATERIALS AND METHODS: Detrusor samples of 13 overactive bladders (sensory urge and detrusor instability) were obtained before botulinum toxin injection and compared to those of 8 normally contractile, nonobstructed bladders obtained during radical cystectomy. Smoothelin mRNA expression patterns were investigated by Northern blot and variant specific reverse transcriptase-polymerase chain reaction as well as by quantitative reverse transcriptase-polymerase chain reaction on laser capture, microdissected smooth muscle. At the protein level smoothelin was investigated by standard and quantitative immunohistochemistry. RESULTS: The bladder muscularis expressed vascular and visceral smoothelin isoforms, and 2 of the known splice variants. In the smooth muscle of patients with detrusor instability and sensory urge a significant 2.4 and 2.2-fold increase, respectively, in smoothelin variant 1 mRNA was observed in comparison to that of normal controls. Analyses at the smoothelin protein level confirmed significant up-regulation in these bladder dysfunctions by a factor of 2.3 and 1.8, respectively. No significant difference in smoothelin expression was observed between detrusor instability and sensory urge. CONCLUSIONS: Increased expression of smoothelin in patients with detrusor instability and sensory urge implies that the etiology of these dysfunctions includes changes in myogenic parameters. In addition, our data support the new classification of the International Continence Society for overactive bladder proposing that sensory urge and detrusor instability represent a single clinical entity.

Cytoskeletal Proteins↗

PubMed2005

Strain typing of German transmissible spongiform encephalopathies field cases in small ruminants by biochemical methods.

Following the implementation of a large scale transmissible spongiform encephalopathies (TSE) surveillance programme of small ruminants, evidence for a natural transmission of bovine spongiform encephalopathy (BSE) to a French goat has been found. During the years 2002-2004, a massive TSE rapid testing programme on >250,000 small ruminants was carried out in Germany. In this national survey, 186 scrapie-affected sheep were found which originated from 78 flocks. The majority of these cases were of the classical TSE type (115 sheep belonging to 14 outbreaks). However, 71 cases coming from 64 flocks were of the novel atypical scrapie type. According to the regulation EU 999/2001, all TSE cases in small ruminants have to be examined by strain typing methods to explore any possibility of the existence of BSE cases in the field sheep population. Here we report on a biochemical typing strategy (termed FLI-test), which includes the determination of molecular masses, antibody binding affinities and glycosylation pattern of the TSE induced abnormal prion protein. Based on this typing approach none of the analysed German classical TSE outbreaks (total number of analysed sheep: 36) displayed biochemical features indicative for a BSE infection. However, in two cases distinct but BSE-unrelated PrP(Sc) types were found, which alludes to the existence of different scrapie strains in the German sheep population.

Animals↗

PubMed2004

Atypical scrapie cases in Germany and France are identified by discrepant reaction patterns in BSE rapid tests.

The intensified surveillance of scrapie in small ruminants in the European Union (EU) has resulted in a substantial increase of the number of diagnosed cases. Four rapid tests which have passed the EU evaluation for BSE testing of cattle are also recommended currently and used for the testing of small ruminants by the EU authorities. These tests include an indirect ELISA (cELISA), a colorimetric sandwich ELISA (sELISA I), a chemiluminescent sandwich ELISA (sELISA II), and a Western blot (WB). To this point, the majority of samples have been screened by using either sELISA I (predominantly in Germany) or WB (predominantly in France). In this study, it is shown that a number of the German and French scrapie cases show inconsistent results using rapid and confirmatory test methods. Forty-eight German sheep, 209 French sheep and 19 French goat transmissible spongiform encephalopathy (TSE) cases were tested. All cases were recognised by the sELISA I and either one of the confirmatory methods (scrapie-associated fibrils (SAF)-immunoblot or immunohistochemistry). Surprisingly, three rapid tests failed to detect a significant number of scrapie cases (29 in France and 24 in Germany). The possible reasons for these inconsistent reaction patterns of scrapie cases are discussed. Similar discrepancies have not been observed during rapid testing of cattle for BSE, the disease for which all diagnostic methods applied have been evaluated.

Animals↗

PubMed2004

Morphological features of cell death.

Cell death is discriminated into two main forms: apoptosis and necrosis. In contrast to necrosis, apoptosis is a regulated, energy-dependent form of cell death leading to phagocytosis of cellular remnants by neighboring cells. Characteristic morphological features of these two forms of cell death will be discussed and correlated to underlying molecular mechanisms.

Animals↗

PubMed2002

Decreased alveolar oxygen induces lung inflammation.

Molecular mechanisms of the inflammatory reaction in hypoxia-induced lung injury are not well defined. Therefore, effects of alveolar hypoxia were studied in rat lungs, exposing rats to 10% oxygen over periods of 1, 2, 4, 6, and 8 h. An increase in the number of macrophages in bronchoalveolar lavage fluid of hypoxic animals was shown between 1 and 8 h. Extravasation of albumin was enhanced after 1 h and remained increased throughout the study period. NF-kappaB-binding activity as well as mRNA for TNF-alpha, macrophage inflammatory protein (MIP)-1beta, and monocyte chemoattractant protein (MCP)-1 were increased within the first 2 h of exposure to hypoxia. Hypoxia-inducible factor (HIF)-1alpha and intercellular adhesion molecule (ICAM)-1 mRNA were upregulated between 1 and 6 h. Elimination of alveolar macrophages by intratracheal application of liposome-encapsulated clodronate led to a decreased expression of NF-kappaB binding activity, HIF-1alpha, TNF-alpha, ICAM-1, and MIP-1beta. In summary, alveolar hypoxia induced macrophage recruitment, an increase in albumin leakage, and enhanced expression of inflammatory mediators, which were mainly macrophage dependent. Alveolar macrophages appear to have a prominent role in the inflammatory response in hypoxia-induced lung injury and the related upregulation of inflammatory mediators.

Animals↗

PubMed2002

Role of alveolar epithelial ICAM-1 in lipopolysaccharide-induced lung inflammation.

Intercellular adhesion molecule-1 (ICAM-1) is known to play a central role in lung inflammation. Limited information, however, is available regarding the expression and biological function of ICAM-1 in the alveolar epithelial compartment. The current report analyses the expression pattern of ICAM-1 in primary cultures of rat alveolar epithelial cells (AECs) and in the rat lung following instillation of bacterial endotoxin (lipopolysaccharide (LPS)) in order to better define the role of alveolar epithelial ICAM-1. AECs stimulated in vitro with LPS were evaluated for ICAM-1 and ICAM-1 messenger ribonucleic acid content. Adherence assays with neutrophils and macrophages were performed. Endotoxin-induced ICAM-1 upregulation on AECs was demonstrated in vivo by immunofluorescence staining. In addition, the effect of intratracheally-instilled anti-ICAM-1 was assessed. Significant upregulation of ICAM-1 occurred in vitro and in vivo on AECs after LPS stimulation. Adherence assays showed a 114% increase in adhesion of neutrophils to AECs. Antibody directed against ICAM-1 reduced this adhesion by 40%. A significant reduction in the number of neutrophils in bronchoalveolar lavage fluid and whole lung was seen under airway ICAM-1 blockade. These data indicate that intercellular adhesion molecule-1 participates in the inflammatory response to lipopolysaccharide-induced lung injury in the distal airways by interacting mainly with neutrophils.

Animals↗

PubMed2001

Two splice variants of the Wilms' tumor 1 gene have distinct functions during sex determination and nephron formation.

Alternative splicing of Wt1 results in the insertion or omission of the three amino acids KTS between zinc fingers 3 and 4. In vitro experiments suggest distinct molecular functions for + and -KTS isoforms. We have generated mouse strains in which specific isoforms have been removed. Heterozygous mice with a reduction of +KTS levels develop glomerulosclerosis and represent a model for Frasier syndrome. Homozygous mutants of both strains die after birth due to kidney defects. Strikingly, mice lacking +KTS isoforms show a complete XY sex reversal due to a dramatic reduction of Sry expression levels. Our data demonstrate distinct functions for the two splice variants and place the +KTS variants as important regulators for Sry in the sex determination pathway.

Active Transport, Cell Nucleus↗

PubMed2001

Calsyntenin-1, a proteolytically processed postsynaptic membrane protein with a cytoplasmic calcium-binding domain.

In a screen for proteins released from synapse-forming spinal cord neurons, we found the proteolytically cleaved N-terminal fragment of a transmembrane protein localized in the postsynaptic membrane of both excitatory and inhibitory synapses. We termed this protein calsyntenin-1, because it binds synaptic Ca2+ with its cytoplasmic domain. By binding Ca2+, calsyntenin-1 may modulate Ca2+-mediated postsynaptic signals. Proteolytic cleavage of calsyntenin-1 in its extracellular moiety generates a transmembrane stump that is internalized and accumulated in the spine apparatus of spine synapses. Therefore, the synaptic Ca2+ modulation by calsyntenin-1 may be subject to regulation by extracellular proteolysis in the synaptic cleft. Thus, calsyntenin-1 may link extracellular proteolysis in the synaptic cleft and postsynaptic Ca2+ signaling.

Animals↗

PubMed2001

[Sweat gland carcinoma in the axillary area. A case report and review of the literature].

The sweat gland carcinoma is a rare tumor, representing in approximately 1% of primary skin lesions. In the literature 220 cases of sweat gland carcinomas have been presented over the last three decades. The medium age is 57 years, with an equal male-to-female distribution. Topographically, the tumor is located at the lower limbs (32.9%), the upper extremities (28%), and the head (26%). Involvement of the trunk is rare. Diagnosis can be complicated as the carcinoma is a rare entity, with no correlation of its histologic classification and biologic presentation. This can only implicate the difficulties in treatment.

Adenocarcinoma↗

PubMed2000

Effect of streptolysin O on the microelasticity of human platelets analyzed by atomic force microscopy.

Atomic force microscopy (AFM) has been shown to be a suitable tool to probe biophysical properties of cells and cell fragments. We analysed biophysical alterations of human platelets by AFM using streptolysin O (SLO) as a model for pore forming proteins. Permeabilization of platelet membrane by SLO was confirmed by transmission electron and confocal microscopy. Using force volume imaging combined with FIEL analysis we were able to show dynamically the increase in the elasticity of platelets during the pore formation by SLO and could correlate the viscoelasticity to the morphology of platelets. Stabilizing the actin cytoskeleton by phalloidin resulted in partial restoration of the elasticity indicating that loss of stability in platelets by SLO is mediated by alterations of both plasma membrane and cytoskeleton.

Bacterial Proteins↗

PubMed2000

Bacterial population of chronic crural ulcers: is there a difference between the diabetic, the venous, and the arterial ulcer?

BACKGROUND: At the Surgical Department of Surgery of the University Hospital Würzburg microbiological examinations were performed of the ulcer grounds from patients with diabetic-neuropathic, diabetic-ischemic, venous, and arterial leg ulcers. The aim of the examination was to evaluate possible differences in the healing process of these ulcers based on the knowledge of their bacterial populations. PATIENTS AND METHODS: In a period of four months, 63 patients were consecutively examined by taking a bacteriological swab of their ulcer area. The healing process of their wounds was followed and related to the impact of bacterial colonisation and clinical signs of infection. RESULTS: 95% of the venous and arterial leg ulcers had a positive smear, whereas only 70% of diabetic ulcers were positive for bacterial growth. Bacterial population of the three ulcer entities, however did not differ significantly. 100% of the clinically infected venous and arterial ulcers but only 80% of the diabetic wounds revealed a positive smear. On the other hand, only 22% of the venous ulcers with a positive smear developed a clinical infection in contrast to 70% of the arterial and diabetic. Venous ulcers showed only in a few patients prolonged healing, even in cases of marked bacterial contamination. Despite of clinical signs of infection however, diabetic wounds sometimes did not reveal a positive wound smear (20%). All infected venous, but only 20% of the infected ischemic ulcers healed satisfactorily. Arterial wounds with no bacterial growth healed significantly better than contaminated wounds. This difference was not significant in the other entities. Radical removal of the infection by minor amputation increased the healing rate in diabetic ulcers over 80%, whereas ischemic wounds did not profit from this therapy. CONCLUSIONS: A positive bacterial wound smear is not inevitably correlated with a protracted leg ulcer healing. Nevertheless a fulminant infection often developed in diabetic ulcers despite the initial inability to demonstrate bacterial growth. In order to start antibiotic treatment as early as possible, a wound smear should be obtained routinely from patients with diabetic ulcers. In chronic venous ulcers, a routine swab does not appear to be indicated as it bears no clinical consequences. The same applies to patients with surgically fully treated peripheral arterial occlusive disease. As ischemia presents the limiting factor, antibiotic therapy in case of infection will not prevent imminent amputation.

Adult↗

PubMed2000

Lipopolysaccharide induces functional ICAM-1 expression in rat alveolar epithelial cells in vitro.

Lipopolysaccharide (LPS)-induced lung inflammation is known to increase pulmonary intercellular adhesion molecule-1 (ICAM-1) expression. In the present study, L2 cells, a cell line of alveolar epithelial cells, were stimulated with LPS, and ICAM-1 expression was studied. ICAM-1 protein on L2 cells peaked at 6 (38% increase; P < 0.01) and 10 (48% increase; P < 0.001) h after stimulation with Escherichia coli and Pseudomonas aeruginosa LPS, respectively. ICAM-1 mRNA expression was markedly increased, with a peak at 2-4 (E. coli) and 4-6 (P. aeruginosa) h. Adherence assays of neutrophils to LPS-stimulated L2 cells showed a threefold increase in adherence (P < 0.001). Pretreatment of the neutrophils with anti-lymphocyte function-associated antigen-1 and anti-Mac-1 antibodies reduced adherence by 54% (P < 0.001). Analysis of immunofluorescence staining for ICAM-1 showed an exclusive apical expression of ICAM-1. These results indicate that LPS upregulates functional active ICAM-1 on the apical part of the membrane in rat pneumocytes.

Animals↗

PubMed2000

Differential subcellular localization of ENaC subunits in mouse kidney in response to high- and low-Na diets.

Previous electrophysiological experiments on renal cortical collecting ducts indicated that dietary sodium intake and variations in aldosterone plasma levels regulate the abundance of functional epithelial Na channels (ENaC) in the apical plasma membrane. In mouse kidney we investigated by immunohistochemistry whether feeding for 3 wk a diet with high (3% Na) and low (0.05% Na) Na content influences the distribution pattern of ENaC. In mice of all experimental groups, ENaC was apparent in cells from the late portion of the distal convoluted tubule (DCT2) down to the medullary collecting duct (CD). In mice on a high-Na diet (plasma aldosterone: 40.8 +/- 2.0 ng/dl), the alpha-subunit was undetectable, and the beta- and gamma-ENaC were detected in the cytoplasm, but not in the apical plasma membrane of the cells. In contrast, in mice on a low-Na diet (plasma aldosterone: 93.6 +/- 9.3 ng/dl) all three ENaC subunits were displayed in the subapical cytoplasm and in the apical membrane of DCT2, connecting tubule (CNT), and, although less prominent, in cortical CD cells. Apical plasma membrane immunostaining progressively decreased along the cortical CD, simultaneously with increasing cytoplasmic staining for beta- and gamma-ENaC. Thus our data on mice adapted to moderately low and high Na intake suggest that regulation of ENaC function in vivo involves shifts of beta- and gamma-subunits from the cytoplasm to the apical plasma membrane and vice versa, respectively. The insertion of these subunits into the apical plasma membrane coincides with upregulation of the alpha-subunit and its insertion into the apical plasma membrane.

Aldosterone↗

PubMed2000

Embryoid bodies: an in vitro model of mouse embryogenesis.

Embryonic stem (ES) cells are pluripotent cells isolated from the inner cell mass of blastocysts. ES cells are able to differentiate into the three primitive layers (endoderm, mesoderm and ectoderm) of the organism, including the germline. To study early stages of development, as well as to investigate the impact of a gene knock-out in vitro, ES cells are differentiated into three-dimensional structures called embryoid bodies, because of their ability to mimick post-implantation embryonic tissues. This review summarises the work on ES cell differentiation into haematopoietic and vascular cells, neuronal and glial cells, myocytes, and adipocytes, using this in vitro model of early embryogenesis. We also present the potential of this method to analyse the impact of genetic alterations in vitro.

Animals↗

PubMed1999

cAMP-dependent and -independent downregulation of type II Na-Pi cotransporters by PTH.

Parathyroid hormone (PTH) leads to the inhibition of Na-Pi cotransport activity and to the downregulation of the number of type II Na-Pi cotransporters in proximal tubules, as well as in opossum kidney (OK) cells. PTH is known also to lead to an activation of adenylate cyclase and phospholipase C in proximal tubular preparations, as well as in OK cells. In the present study, we investigated the involvement of these two regulatory pathways in OK cells in the PTH-dependent downregulation of the number of type II Na-Pi cotransporters. We have addressed this issue by using pharmacological activators of protein kinase A (PKA) and protein kinase C (PKC), i.e., 8-bromo-cAMP (8-BrcAMP) and beta-12-O-tetradecanoylphorbol 13-acetate (beta-TPA), respectively, as well as by the use of synthetic peptide fragments of PTH that activate adenylate cyclase and/or phospholipase C, i.e., PTH-(1-34) and PTH-(3-34), respectively. Our results show that PTH signal transduction via cAMP-dependent, as well as cAMP-independent, pathways leads to a membrane retrieval and degradation of type II Na-Pi cotransporters and, thereby, to the inhibition of Na-Pi cotransport activity. Thereby, the cAMP-independent regulatory pathway leads only to partial effects (approximately 50%).

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed1999

Induction and nuclear translocation of hypoxia-inducible factor-1 (HIF-1): heterodimerization with ARNT is not necessary for nuclear accumulation of HIF-1alpha.

Hypoxia-inducible factor-1 (HIF-1) is a master regulator of mammalian oxygen homeostasis. HIF-1 consists of two subunits, HIF-1alpha and the aryl hydrocarbon receptor nuclear translocator (ARNT). Whereas hypoxia prevents proteasomal degradation of HIF-1alpha, ARNT expression is thought to be oxygen-independent. We and others previously showed that ARNT is indispensable for HIF-1 DNA-binding and transactivation function. Here, we have used ARNT-mutant mouse hepatoma and embryonic stem cells to examine the requirement of ARNT for accumulation and nuclear translocation of HIF-1alpha in hypoxia. As shown by immunofluorescence, HIF-1alpha accumulation in the nucleus of hypoxic cells was independent of the presence of ARNT, suggesting that nuclear translocation is intrinsic to HIF-1alpha. Co-immunoprecipitation of HIF-1alpha together with ARNT could be performed in nuclear extracts but not in cytosolic fractions, implying that formation of the HIF-1 complex occurs in the nucleus. A proteasome inhibitor and a thiol-reducing agent could mimic hypoxia by inducing HIF-1alpha in the nucleus, indicating that escape from proteolytic degradation is sufficient for accumulation and nuclear translocation of HIF-1alpha. During biochemical separation, both HIF-1alpha and ARNT tend to leak from the nuclei in the absence of either subunit, suggesting that heterodimerization is required for stable association within the nuclear compartment. Nuclear stabilization of the heterodimer might also explain the hypoxically increased total cellular ARNT levels observed in some of the cell lines examined.

Animals↗