Biomedical subjects

S Steiner

Publications and source records attributed to S Steiner.

At least 19 recordsLinked to original sources

PubMed1998

Cyclosporine A-induced decrease in calbindin-D 28 kDa in rat kidney but not in cerebral cortex and cerebellum.

Recently, we reported that in rat, cyclosporine A (CsA) markedly decreases the levels of calbindin-D (CABP-D) 28 kDa in kidney. CABP-D 28 kDa is a calcium-binding protein which is highly expressed in calcium-transporting tissues such as kidney or brain. In this study, we investigated whether, in addition to the kidney, CsA also has an effect on CABP-D 28 kDa in rat brain. Three groups of male Wistar rats received 15 mg/kg/day or 50 mg/kg/day of CsA orally for 12 days, whereas controls received vehicle solution for the same period. CABP-D 28-kDa protein and CsA were quantified in homogenates of kidney, cerebral cortex and cerebellum, and the localization of CABP-D 28 kDa was assessed in the different tissue sections by immunohistochemistry. In kidney, CABP-D 28 kDa was strongly and dose dependently decreased, and was located in tubular epithelial cells. In brain, CABP-D 28 kDa was not changed and was mainly located in pyramidal cells of the cortex and in cerebellum exclusively in Purkinje cells. High CsA concentrations were measured in kidney, more than 17-fold greater than those found in cortex. In cerebellum, CsA was below the limit of detection. These data suggest that at clinically relevant doses, CsA may not affect CABP-D 28-kDa levels in brain.

Animals↗

PubMed1998

The cyclosporine A-induced decrease in rat renal calbindin-D28kDa protein as a consequence of a decrease in its mRNA.

Cyclosporine A (CsA) is a potent immunosuppressant with the drawback of renal side-effects. We recently reported that relatively high doses of CsA markedly decreased the calcium-binding protein calbindin-D28kDa in kidneys of male Wistar rats, and showed that this decrease could be associated with some of the drug-induced adverse renal effects. To investigate the events leading to this decrease, the calbindin-D28kDa mRNA level in kidneys of rats treated with 15 or 50 mg/kg/day CsA for 12 days was analysed by reverse transcription followed by polymerase chain reaction. At both doses, a marked dose-dependent decrease in the calbindin-D28kDa mRNA level was found, one very similar to the decrease measured in the calbindin-D28kDa protein abundance. Thus, the CsA-mediated down-regulation of the renal calbindin-D28kDa protein is most likely the result of a decrease in the calbindin-D28kDa mRNA level.

Animals↗

PubMed1998

New insights into cyclosporine A nephrotoxicity by proteome analysis.

Using two-dimensional gel electrophoresis (2-DE), we recently discovered an association between decreased calcium-binding protein, calbindin-D 28 kDa, urinary calcium wasting and intratubular corticomedullary calcifications in rat kidney. This observation prompted us to investigate kidney tissues of other species, including man. In this paper we show that in dogs and monkeys, which are generally devoid of cyclosporine A (CsA)-mediated nephrotoxicity, renal calbindin levels were not affected by the CsA treatment whereas in CsA-treated human kidney-transplant recipients with renal vascular or tubular toxicity, a marked decrease in renal calbindin-D 28 kDa protein level was found in most of the kidney biopsy sections. The present results strongly suggest that calbindin is a marker for CsA-nephrotoxicity. The discovery of calbindin-D 28 kDa being involved in CsA toxicity has evolved from the application of 2-DE and has not been reported previously, proving that proteomics can provide essential information in mechanistic toxicology. Considering the current improvements in proteome methods it is expected that high throughput proteomics will become an indispensable tool in preclinical safety testing.

Animals↗

PubMed1998

Efficient identification of point mutations by automated DNA sequencing of artificial heterozygote samples.

DNA sequencing templates of individual point mutants of the lacI target gene were amplified by polymerase chain reaction (PCR). By mixing the PCR fragments from two individual mutants in a defined ratio, samples of artificial heterozygous composition were prepared. These samples were then submitted to automated DNA sequencing. The simultaneous, visual comparison of the mixed mutant traces using a graphics program efficiently revealed all heterozygous positions. Based on the individual intensities of the heterozygous base signals the identified point mutations could be assigned to the corresponding mutants. This efficient approach doubles the sample throughput for both the sequencing reactions and the gel electrophoresis using an automated DNA sequencing system.

Animals↗

PubMed1998

Changes in the liver protein pattern of female Wistar rats treated with the hypoglycemic agent SDZ PGU 693.

SDZ PGU 693 acts as a hypoglycemic agent by stimulating glucose utilisation in insulin-sensitive peripheral tissues, such as skeletal muscle and fat. In a 28 day toxicity study the compound was found to induce hepatocellular hypertrophy in Wistar rats treated with 300 mg/kg/day. To gain insights into the pathomechanism of these alterations, aliquots of liver samples from control and treated female Wistar rats were separated by two-dimensional protein gel electrophoresis and the digitized images of the protein patterns were searched for protein abundance changes. Significant treatment-related quantitative changes (P < 0.001) were found in 29 liver proteins. Major increases were observed in several microsomal proteins, including NADPH cytochrome P-450 reductase, cytochrome b5 and serine protease inhibitor. The changes in the cytochrome related enzymes, both known co-factors of the P-450 enzyme system, strongly suggest that SDZ PGU 693 induces microsomal proliferation and induction of the P-450 enzyme system. Decreases were observed in a series of mitochondrial proteins, such as F1ATPase-delta subunit and ornithine aminotransferase precursor as well as in several cytosolic proteins such as the liver fatty acid binding protein, arylsulfotransferase and the senescence marker protein-30. The changes in F1ATPase-delta subunit and liver fatty acid binding protein together suggest a down-regulation of the mitochondrial liver fatty acid metabolism, likely reflecting the pharmacological action of the compound. These results show that SDZ PGU 693 produces a complex pattern of gene expression changes which give insights into the molecular mechanisms of both its pharmacological action and a toxic response.

Animals↗

PubMed1998

[Results and complications of fiber bronchoscopy in HIV positive patients].

Fibreoptic bronchoscopy is an established diagnostic procedure for HIV-associated pulmonary infections. We retrospectively evaluated the diagnostic effectivity and safety of fibreoptic bronchoscopy with bronchoalveolar lavage (BAL) and transbronchial biopsy (TBB) in 153 patients with late-stage HIV infection and clinical signs of pulmonary infection or abnormal chest radiograph. Bronchoscopy leads to diagnosis in 82.4% and changed therapy in 54%. 45 patients (30%) were found to have pneumocystis carinii pneumonia (PCP), the most common bronchoscopic finding, followed by bacterial lung disease (29.3%). BAL had a sensitivity of 78% for PCP. Diagnostic yield of BAL for PCP was higher in patients without previous treatment (positive results in 82%) with regard to PCP independend of the prior treatment. Serious complication occurred in 22 cases (pneumothorax: 6 (3.9%), bleeding: 12 (7.8%), hypoxaemia: 4 (2.6%)). High serum levels of lactate dehydrogenase (LDH) correlated with pulmonary complications like pneumothorax. Age, sex and kind of pulmonary infection did not influence complication rates. 6 (3.9%) episodes of spontaneous pneumothorax occurred in the further course, 3 of them concurrently with PCP or prior history of PCP. We conclude that fibreoptic bronchoscopy is of great value for diagnosing pulmonary infection in HIV-seropositive patients. TBB provides incremental diagnostic information not available from BAL, especially in patients pretreated with cotrimoxazol or pentamidin. For that reason we believe that TBB should be performed in these patients.

AIDS-Related Opportunistic Infections↗

PubMed1997

Decrease in kidney calbindin-D 28kDa as a possible mechanism mediating cyclosporine A- and FK-506-induced calciuria and tubular mineralization.

The use of the immunosuppressant cyclosporine A (CsA) is limited by its adverse renal effects. Most recently, we reported that the drug markedly decreases the levels of the calcium-binding protein calbindin-D 28kDa in kidneys of male Wistar rats. In the present study, the potential relationship between drug-induced nephrotoxicity and the decrease in kidney calbindin-D 28kDa was investigated. Four groups of male Wistar rats were treated for 10 or 31 days with either the immunosuppressant CsA (50 mg/kg/day), FK-506 (5 mg/kg/day), rapamycin (5 mg/kg/day) or with the nonimmunosuppressive cyclosporine derivative 3'keto-[Bmt1]-[Val2]-CsA (SDZ PSC-833) (50 mg/kg/day), and the effects on calcium homeostasis, kidney histology and renal calbindin-D 28kDa were examined. Similar effects were found with CsA and FK-506; both drugs strongly reduced kidney calbindin-D 28kDa protein levels, increased urine calcium excretion, caused intratubular calcification, and induced basophilic tubules. In contrast, rapamycin and SDZ PSC-833 caused no decrease in renal calbindin-D 28kDa levels, no noticeable alterations in calcium metabolism, and no renal calcification. The results provide evidence for a link between decreased renal calbindin, increased calcium urine excretion, and intratubular kidney calcification. The present data show no correlation between the decrease in renal calbindin and the induction of basophilic tubules; however, it needs to be investigated if these apparently independent kidney effects may have a common origin upstream of calbindin expression.

Animals↗

PubMed1997

[Prognostically relevant parameters in patients with coronary heart disease, arterial hypertension and sleep apnea disorders].

Patients with untreated sleep apnea syndrome have a higher cardiovascular mortality. It is not known which mechanisms lead to this increase in mortality and whether it is independent from the often associated coronary heart disease and systemic hypertension. In 48 consecutive patients with coronary heart disease confirmed by angiography, exercise-ECG, Holter-ECG, echocardiography, spirometric tests, analysis of ventricular late potentials, heart rate variability and a test for sleep-disordered breathing with a screening device were performed. Seventeen patients showed disordered breathing during sleep (obstructive sleep apnea) with a desaturation index of > or = 10 (mean desaturation index 17.3 +/- 9.3 vs. 2.6 +/- 3.1 in the patients without sleep-disordered breathing). There are no significant differences in age (58.9 +/- 6.1 vs. 59.7 +/- 7.6 years), body-mass-index (28.6 +/- 3.7 vs. 27.7 +/- 3.3 kg/m2), left ventricular ejection fraction (57.2 +/- 13.6 vs. 64.0 +/- 14.6%), forced expiratory volume in 1 second/vital capacity 95.4 +/- 13.9 vs. 92.9 +/- 11.2% predicted, heart rate variability (standard deviation of the RR-intervals 39.4 +/- 29.4 vs. 37.2 +/- 17.0 ms), the frequency of premature ventricular beats over 24 h and at night, the frequency of multivessel disease (71 vs. 68%), additional hypertension 53 vs. 48%), status postmyocardial infarction (47 vs. 48%) and positive late potential analysis (24 vs. 13%). There were no ST segment depressions during the night. Patients with coronary heart disease and mild sleep-disordered breathing show no significant differences in the investigated parameters compared with patients without obstructive sleep apnea or sleep-disordered breathing.

Adult↗

PubMed1996

Cyclosporine A decreases the protein level of the calcium-binding protein calbindin-D 28kDa in rat kidney.

Despite the widespread use of cyclosporine A (CsA), its mechanism of action and side effects are not yet completely understood. There exists a large body of evidence suggesting that disturbance of calcium homeostasis is a critical step in the cascade of cellular and molecular events induced by the drug. As recently shown in our laboratory by two-dimensional protein gel electrophoresis (2-DE) analysis of kidney homogenates, CsA induced numerous changes in several kidney proteins. One kidney protein in particular was shown to be strongly down-regulated by the drug. In this work we report the identification of the strongly decreased kidney protein as calbindin-D 28kDa, a vitamin D-dependent calcium-binding protein associated with calcium handling by cells. The assignment of the down-regulated protein spot is based on its internal amino acid sequence analysis and its specific reaction with a monoclonal antibody raised against calbindin-D 28kDa. In kidney homogenates of male Wistar rats treated with 50 mg/kg/d CsA for up to 28 days, calbindin levels were measured by ELISA and were shown to be continuously decreased with prolonged CsA treatment. To our knowledge, this is the first report describing the effect of CsA on kidney calbindin-D 28kDa protein levels. Further studies are needed to elucidate whether the CsA-mediated down-regulation of the calcium-binding protein calbindin-D 28kDa may be a critical factor for the renal adverse effects induced by this drug.

Amino Acid Sequence↗

PubMed1996

Induction of the adipose differentiation-related protein in liver of etomoxir-treated rats.

The effects of etomoxir, an irreversible carnitine palmitoyltransferase I inhibitor, on the liver protein pattern and on liver morphology were examined by two-dimensional gel electrophoresis in female Sprague-Dawley rats treated with 125 mg/kg/day etomoxir for 28 days. In livers of treated animals a protein spot was found which was not present in controls. The spot was identified by internal amino acid sequence analysis as the adipose differentiation-related protein (ADRP). The expression of ADRP in liver is a novel finding as the protein has been described previously as adipocyte-specific. Additionally we found histopathologic evidence of lipid accumulation in the livers of etomoxir rats. The data show that for each treated rat there was a good correlation between ADRP levels and degree of lipid droplet formation. This observation may suggest a potential relationship between drug-induced expression of ADRP in liver and lipid accumulation.

Amino Acid Sequence↗

PubMed1996

[Skull injury in childhood: comparison of ultrasonography with conventional X-rays and computerized tomography].

PURPOSE: The purpose of our study was to compare the value of ultrasound, conventional x-ray diagnosis and CT in detecting skull fractures and intracranial haemorrhage in children suffering from a head injury. MATERIAL AND METHODS: We examined 210 children who had a head injury. In all cases the calvarium was investigated by ultrasound using a 7.0 MHz linear transducer. In children with an open fontanel (n = 190) the cerebrum was screened additionally by ultrasound following a standard protocol. The sonographic findings were correlated to the x-ray examination (n = 21) and CT (n = 13). RESULTS: Ultrasound enabled diagnosis of linear calvarial fractures (n = 29), depressed fractures (n = 6) and intracranial haemorrhage (n = 8). X-Ray and CT examination confirmed the diagnosis of linear calvarial fractures in 16 cases, of depressed fractures in 6 cases. CT confirmed the sonographic diagnosis of intracranial haemorrhage in 8 cases. CONCLUSION: Ultrasound as a primary method can replace the conventional x-ray in detecting calvarial fracture and posttraumatic sequelae. Additional CT examination depends on the sonographic and neurological status.

Age Factors↗

PubMed1995

Effects of cyclosporine A on the rat liver and kidney protein pattern, and the influence of vitamin E and C coadministration.

The effects of cyclosporine A (CsA), a potent immunosuppressive drug, were examined in rat liver and kidney samples using two-dimensional electrophoretic protein analysis. Of a total of 370 liver and 336 kidney spots analyzed, 8% (29 spots) and 6% (19 spots), respectively, showed a significant drug-induced change (p < 0.01), which was predominantly reflected in increased protein abundance (62% and 74% of the changes, respectively). Of the 48 proteins changed in either organ, 14 were most probably common to both tissues and one of these was significantly increased in both the liver and the kidney. Most of the other 13 showed similar trends (either increases or decreases) in both organs. However, the most striking drug effect seen in this study concerned an unidentified protein present only in the kidney, which completely disappeared upon CsA treatment. It was also investigated whether the drug-induced changes could be prevented by the coadministration of the radical scavengers vitamin E and C with CsA. Spots changed by the administration of the drug were classified according to three different categories, based on their response profiles in rats treated with CsA in combination with the vitamins: (i) spots which were changed by CsA as well as by CsA in combination with the vitamins (12 liver and 4 kidney spots), (ii) spots which were changed by CsA and showed an additional increase of this change by CsA plus the vitamins (no liver and 4 kidney spots), and (iii) spots which were changed by CsA but not by CsA in combination with the vitamins (8 liver and 6 kidney spots). These results showed that in both organs the vitamins were able to prevent around 30% of the effects caused by CsA, and that two-dimensional gel electrophoresis is an excellent tool to demonstrate such drug interactions at the molecular level.

Animals↗

PubMed1995

Protein variability in male and female Wistar rat liver proteins.

In the emerging field of mechanistic toxicology, growing attention is being paid to the interpretation of sex-related toxic responses. The Wistar rat outbred stock is a frequently used rodent for toxicity testing. Outbred strains, which display a relatively high degree of genetic variability between the individual animals are often used in risk assessment, as they are considered to best approximate the variability present in a human population. In this study two-dimensional (2-D) protein gel electrophoresis was applied to investigate the liver protein patterns of male and female Wistar rats and to search for (i) sex-related and (ii) interindividual qualitative and quantitative differences in protein expression. Among the sexes, six proteins were detected that were shown to be exclusively present in male rats and one that was present only in females. A male-specific protein was tentatively assigned to alpha 2u globulin. Seven protein spots showed statistically significant abundance changes (p < 0.001) between males and females, one was tentatively assigned to heme oxygenase 1 and another to the 23 kDa morphin-binding protein. Four sets of protein spots were detected that showed positional shifts in the individual patterns and are likely to represent polymorphic proteins inherent in the Wistar rat. These results form a valuable basis for future investigations of drug-induced changes in the male and female Wistar rat liver pattern.

Alpha-Globulins↗

PubMed1995

An updated two-dimensional gel database of rat liver proteins useful in gene regulation and drug effect studies.

We have improved upon the reference two-dimensional (2-D) electrophoretic map of rat liver proteins originally published in 1991 (N. L. Anderson et al., Electrophoresis 1991, 12, 907-930). A total of 53 proteins (102 spots) are now identified, many by microsequencing. In most cases, spots cut from wet, Coomassie Blue stained 2-D gels were submitted to internal tryptic digestion [2], and individual peptides, separated by high-performance liquid chromatography (HPLC), were sequenced using a Perkin-Elmer 477A sequenator. Additional spots were identified using specific antibodies.

Animals↗

PubMed1995

[MRI of focal liver lesions using a 1.5 turbo-spin-echo technique compared with spin-echo technique].

AIM: The aim of our study was to evaluate a T2-weighted turbo-spinecho sequence in comparison to a T2-weighted spinecho sequence in imaging focal liver lesions. METHODS: In our study 35 patients with suspected focal liver lesions were examined. Standardised imaging protocol included a conventional T2-weighted SE sequence (TR/TE = 2000/90/45, acquisition time = 10.20) as well as a T2-weighted TSE sequence (TR/TE = 4700/90, acquisition time = 6.33). Calculation of S/N and C/N ratio as a basis of quantitative evaluation was done using standard methods. A diagnostic score was implemented to enable qualitative assessment. RESULTS: In 7% (n = 2) the TSE sequence enabled detection of further liver lesions showing a size of less than 1 cm in diameter. Comparing anatomical details the TSE sequence was superior. S/N and C/N ratio of anatomic and pathologic structures of the TSE sequence were higher compared to results of the SE sequence. CONCLUSION: Our results indicate that the T2-weighted turbo-spinecho sequence is well appropriate for imaging focal liver lesions, and leads to reduction of imaging time.

Carcinoma, Hepatocellular↗

PubMed1995

Homologous recombination as the main mechanism for DNA integration and cause of rearrangements in the filamentous ascomycete Ashbya gossypii.

A slow and a fast growth phenotype were observed after transformation of the phytopathogenic fungus Ashbya gossypii using a plasmid carrying homologous DNA and as selectable marker the Tn903 aminoglycoside resistance gene expressed from a strong A. gossypii promoter. Transformations with circular plasmids yielded slowly and irregularly growing geneticin-resistant mycelia in which 1% of nuclei contained plasmid sequences. Occasionally, fast growing sectors appeared which were shown to be initiated by homologous integration of the transforming DNA. Transformants obtained with plasmids linearized within the homology region immediately exhibited fast radial growth. In all 28 transformants analyzed plasmid DNA was integrated homologously. Such apparent lack of nonhomologous recombination has so far not been observed in filamentous ascomycetes. In 14 transformants two to four tandemly integrated plasmid copies were found. They underwent several types of genetic changes, mainly in the older mycelium: excision of whole plasmid copies and rearrangements within the integrated DNA (inversions and deletions). These internal rearrangements involved 360-bp inverted repeats, remnants of IS-elements flanking the resistance gene, and 156-bp direct repeats, originating from the strong A. gossypii promoter. Improved vectors lacking sequence repetitions were constructed and used for stable one-step gene replacement in A. gossypii.

Aminoglycosides↗

PubMed1995

[Diagnosis of cerebral metastasis with standard dose gadobutrol vs. a high dose protocol. Intraindividual evaluation of a phase II high dose study].

PURPOSE: To assess the effectiveness and safety of normal and high doses of Gadobutrol versus a standard dose of Gadolinium DTPA in the MR evaluation of patients with brain metastases. MATERIAL AND METHODS: In a clinical phase-II study 20 patients who had been diagnosed as having brain metastases with CT or MRT were studied prospectively with Gadobutrol, a new nonionic, low osmolality contrast agent. Each patient received an initial injection of 0.1 mmol/kg body weight and an additional dose of 0.2 mmol/kg Gadobutrol 10 min later. Spin-echo images were obtained before and after the two applications of Gadobutrol. Dynamic scanning (Turbo-FLASH) was performed for 3 min after each injection of the contrast agent. Both quantitative and qualitative data were intraindividually evaluated. The primary tumor was a bronchial carcinoma in 11 cases; in 9 other cases there were different primary tumors. RESULTS: Forty-eight hours after the use of Gadobutrol there were no adverse signs in the clinical examination, vital signs or blood and urine chemistry. Statistical analysis (Friedman test and Wilcoxon test) of the C/N ratios between tumor and white matter, percentage enhancement, and visual assessment rating revealed statistically significant superiority of high-dose Gadobutrol injection in comparison to the standard dose. The percentage enhancement increased on average from 104% after 0.1 mmol/kg to 162% after 0.3 mmol/kg Gadobutrol. Qualitative delineation and contrast of the lesions increased significantly. The use of high-dose Gadobutrol improved the detection of 36 additional lesions in 6 patients. CONCLUSION: The first in vivo results prove the excellent contrast capacity of the nonionic contrast agent Gadobutrol for the diagnosis of intracerebral metastases.

Adult↗

PubMed1994

Cloning and sequencing of a gene coding for an actin binding protein of Saccharomyces exiguus.

The actin binding protein Abp1p of the yeast Saccharomyces cervisiae is thought to be involved in the spatial organisation of cell surface growth. It contains a potential actin binding domain and an SH-3 region, a common motif of many signal transduction proteins [1]. We have cloned and sequenced an ABP1 homologous gene of Saccharomyces exiguus, a yeast which is only distantly related to S. cerevisiae. The protein encoded by this gene is slightly larger than the respective S. cerevisiae protein (617 versus 592 amino acids). The two genes are 67.4% identical and the deduced amino acid sequences share an overall identity of 59.8%. The most conserved regions are the 148 N-terminal amino acids containing the potential actin binding site and the 58 C-terminal amino acids including the SH3 domain. In addition, both proteins contain a repeated motif of unknown function which is rich in glutamic acids with the sequence EEEEEEEAPAPSLPSR in the S. exiguus Abp1p.

Actins↗