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Barbara Kofler

Publications and source records attributed to Barbara Kofler.

11 recordsLinked to original sources
PubMed2005

Pharmacological and functional characterization of galanin-like peptide fragments as potent galanin receptor agonists.

The hypothalamic galanin-like peptide (GALP) was isolated by its ability to activate galanin receptors. The mature porcine GALP is a 60-amino acid neuropeptide proteolytically processed from a 120-amino acid precursor protein. It contains a region identical to the N-terminal 13-amino acids of the neuropeptide galanin. Within the sequence of human GALP (1-60) a potential proteolytic cleavage site between two basic amino acids is present at position 33, which might lead to a shorter C-terminally amidated peptide. In addition, the first two amino acids could be potentially removed via the action of dipeptidase IV. Ligand binding assays using the human neuroblastoma cell line SH-SY5Y transfected with the respective galanin receptors revealed that human GALP (1-60) displayed the highest affinity for the galanin receptor subtype GalR3 (IC50 = 10 nM) followed by GalR2 (IC50 = 28 nM) and GalR1 (IC50 = 77 nM). Ligand binding assays and functional studies showed that the human GALP (3-32) fragment was at least as potent as full length GALP (1-60). Other studies have shown that shorter fragments like human GALP (1-21) and GALP (22-60) were not effective on feeding responses in mice as compared to the full length peptide. Taken together these data suggest that the putative fragment GALP (3-32) might represent the strongest mediator of biological GALP activity. Furthermore it might be a useful tool to study the affinity of GALP to galanin receptors and to search for specific GALP receptors.

Amino Acid Sequence↗
PubMed2005

Galanin and galanin receptors in human cancers.

The increasing interest in peptides and peptide receptors in cancer is based on the possibility of receptor targeting, because peptide receptors are often expressed in different human tumors. The neuropeptide galanin has also been suggested to be involved in the development of neuroendocrine tumors based on the development of estrogen-induced tumors in estrogen-sensitive rats. This study summarizes our current knowledge on the expression of galanin peptide and galanin receptors in different human neuroendocrine tumors. The expression of both, peptide and corresponding receptor, seems to be a common feature of human gliomas, pheochromocytomas, pituitary and neuroblastic tumors. The co-expression of galanin and its receptors supports a role for galanin in tumor cell pathology via autocrine/paracrine mechanisms.

Galanin↗
PubMed2005

Rapid screening of the entire mitochondrial DNA for low-level heteroplasmic mutations.

Alterations of the mitochondrial DNA (mtDNA) are implicated in various pathological conditions. In this study, we used denaturing high performance liquid chromatography (DHPLC) as a method to rapidly screen the entire mtDNA for mutations. Overlapping DNA fragments, amplified by one single cycling protocol from frozen pre-formulated PCR mixes, were subjected to DHPLC analysis. Single DHPLC injections of fragments yielded straightforward interpretation of results with a detection limit down to 1% mtDNA heteroplasmy. Furthermore, collection and re-amplification of low degree heteroduplex peak-fractions allowed sequence analysis of mtDNA mutations down to the detection limit of the DHPLC method. In order to demonstrate that the method has diagnostic value, we analyzed and confirmed known mtDNA mutations in patient samples.

Chromatography, High Pressure Liquid↗
PubMed2004

Decrease of mitochondrial DNA content and energy metabolism in renal cell carcinoma.

To elucidate the relationship between tumor genesis and the mitochondrial energy metabolism in renal neoplasms, we studied three individual enzyme activities of the oxidative phosphorylation, two components of the Krebs cycle and the mitochondrial DNA content of renal carcinomas including 29 conventional, five papillary, two unclassified carcinomas with sarcomatoid features and one collecting duct carcinoma. A significant reduction of all mitochondrial enzyme activities including complex V, as well as of the mitochondrial DNA content was detected in 34 of 37 renal carcinoma tissues as compared with control kidney. Mitochondrial enzyme activities and mitochondrial DNA levels were not statistically different between the conventional, papillary and unclassified sarcomatoid type of renal carcinoma and did not correlate with tumour grade, metastasis, ploidy and proliferative activity as determined by Ki-67 staining. Taken together, our data indicate that a co-ordinated down-regulation of all components necessary for mitochondrial energy metabolism occurs in most renal carcinomas as an early event in carcinoma formation, which does not change with progression of the disease.

Adult↗
PubMed2003

Galanin receptor subtype GalR2 mediates apoptosis in SH-SY5Y neuroblastoma cells.

Recently we have shown that galanin binding significantly correlates with survival in neuroblastoma patients, indicating a possible modulatory role of galanin receptors in neuroblastic tumor biology. However, the molecular mechanisms beyond this correlation have not been elucidated. Here, the cellular effects on activation of specific galanin receptor subtypes in human SH-SY5Y neuroblastoma cells were analyzed using a tetracycline-controlled expression system. Pharmacological studies confirmed the inducible expression of high affinity binding sites for galanin in SH-SY5Y cells transfected with the galanin receptors GalR1 (SY5Y/GalR1) and GalR2 (SY5Y/GalR2). Microphysiometry revealed that both receptor subtypes were able to mediate an intracellular signal upon galanin application. Interestingly, induction of receptor expression and treatment with 100 nm galanin resulted in a dramatic decrease in cell viability in SY5Y/GalR2 cells (93 +/- 3%) compared with a less pronounced effect in SY5Y/GalR1 cells (19 +/- 10%). The antiproliferative potency of galanin was 100-fold higher in SY5Y/GalR2 (50% effective concentration, 1.1 nm) than in SY5Y/GalR1 cells (50% effective concentration, 190 nm). Furthermore, activation of receptor expression and exposure to galanin resulted in apparent morphological changes indicative of apoptosis in SY5Y/GalR2 cells only. Induction of cell death by the apoptotic process was confirmed by poly-(ADP-ribose)-polymerase cleavage, caspase-3 activation, and the typical laddering of DNA. This study indicates that a high level of GalR2 expression is able to inhibit cell proliferation and induce apoptosis in neuroblastoma cells and therefore identifies GalR2 as a possible target for pharmacological intervention in neuroblastoma.

Apoptosis↗
PubMed2003

Galanin and galanin receptors in human gliomas.

Galanin-like immunoreactivity (GAL-LI) and specific GAL binding sites have been shown to be widely distributed in the central nervous system (CNS) and in CNS tumors. GAL and its receptors have also been shown to be present in glial cells, but to date it is still unknown whether human gliomas produce GAL and express GAL receptors. In this study 20 brain tumors consisting of 15 glioblastomas, 4 meningiomas and 1 gliosarcoma were investigated for the presence of GAL-LI and GAL receptors. Immunofluorescence analysis revealed a dense network of GAL-LI positive cellular processes and cell bodies in 18 of the 20 tumors. In contrast, in vitro (125)I-labeled GAL receptor autoradiography showed substantial GAL binding in only 6 glioblastoma tissues. Reverse transcription-PCR analysis detected mRNA of all three known galanin receptors in the tumor tissues, with most tumors expressing multiple receptor subtypes. Pharmacological analysis of tumor membrane homogenates with GAL and the specific GAL receptor GalR2 agonist, AR-M1896, revealed that the GAL receptor GalR1 is most likely the receptor responsible for the observed GAL binding in the glioblastomas. No correlation could be found between GAL-LI, the level of GAL binding and proliferative activity as determined by immunostaining with the cell proliferation marker Ki-67.

Adult↗
PubMed2002

Severe depletion of mitochondrial DNA in spinal muscular atrophy.

Spinal muscular atrophy (SMA) is a neuromuscular disorder in childhood leading to a dramatic loss of muscle strength. Functional investigations with high-resolution polarography and enzyme measurements of the respiratory chain revealed lowered activities in muscle tissue of SMA patients. To gain a better understanding of this low energy supply we analyzed the amount of mitochondrial DNA (mtDNA) in skeletal muscle of 20 unrelated children with genetically proven SMA and 31 controls. Quantitative Southern blot analysis revealed a severe and homogeneous decrease in the content of muscle mtDNA in relation to nuclear DNA in SMA patients (90.3+/-7.8%), whereas by immunofluorescence no decrease in the number of mitochondria was detected. In addition, a two- to threefold reduction of the nuclear-encoded complex II (succinate dehydrogenase) activity was detected in SMA muscle tissue. Western blot analysis showed a significant reduction of both mitochondrial- and nuclear-encoded cytochrome c oxidase subunits. Our results indicate that mtDNA depletion in SMA is a consequence of severe atrophy, and has to be differentiated by measurement of complex II from an isolated reduction of mtDNA as found in patients with mitochondriocytopathies and the so-called mtDNA depletion syndrome.

Adolescent↗
PubMed2002

125I-labeled galanin binding sites in congenital innervation defects of the distal colon.

Neuropeptides have turned out to be promising new parameters, in addition to the routinely performed histochemical diagnosis, of Hirschsprung's disease (HD). Studies of the peptidergic innervation of the affected intestinal segment of patients with HD have demonstrated a marked reduction in the density of several neuropeptide-containing nerve fibers. The frequency of nerve fibers storing the neuropeptide galanin (GAL) was found to be unchanged or slightly reduced in HD, but nothing is known about the occurrence of GAL receptors. In this study, in vitro receptor autoradiography using (125)I-labeled GAL and GAL immunofluorescence have been performed on frozen tissue sections from colon biopsies of 10 patients diagnosed with HD, 8 patients with intestinal neuronal dysplasia (IND B) and 20 patients with chronic obstruction but normal innervation. Binding sites were mainly detected in the mucosal and muscular layer, in acetylcholinesterase-positive nerve fiber bundles and ganglia within the submucosal layer and in close association to blood vessels. An increased population of GAL receptor positive, parasympathetic nerve fibers was seen in the aganglionic segment of HD as compared to controls and IND B. In contrast, GAL immunostaining which was unchanged in HD revealed a significant lack of GAL-positive structures in IND B colon biopsies. Colocalization of GAL and GAL binding sites was only observed in thick nerve fibers in the submucosa. The presence of GAL binding sites in different cellular structures suggests an involvement of GAL in various physiological functions of the gastrointestinal tract.

Adolescent↗
PubMed2002

Elevated expression of galanin receptors in childhood neuroblastic tumors.

The neuropeptide galanin (GAL) has been shown to be present in certain brain tumors. In order to learn more about GAL and its receptors in human tumors of the peripheral nervous system, we investigated the expression of the GAL peptide and the GAL receptors in tumor tissue from childhood neuroblastic tumors. GAL peptide concentrations up to 674 +/- 166 fmol/mg of tissue were detected by radioimmunoassay, but no significant correlation with standard tumor markers or the prognosis of the 14 patients investigated was observed. Ligand binding experiments showed different levels of GAL binding in all 28 primary neuroblastomas and 7 ganglioneuromas investigated. All three human GAL receptor subtypes cloned to date could be detected, with the GALR1 receptor subtype being expressed most prominently. GAL binding did not significantly correlate with genetic markers such as unfavorable DNA ploidy, amplification of the oncogene MYCN and allelic loss of chromosome 1p. However, low galanin binding was significantly correlated with survival (p = 0.021) in this limited analysis of neuroblastic tumor samples. These results raise the possibility that the expression of GAL binding sites may play a role in neuroblastic tumor biology and behavior.

Brain Neoplasms↗
PubMed2002

Preliminary observations on the need for control angiography after peripheral endovascular brachytherapy using a centering balloon.

PURPOSE: To examine the need for completion angiography after endovascular femoropopliteal brachytherapy using a centering catheter. METHODS: Nine consecutive patients (7 men; mean age 68 years, range 53-79) were enrolled in a double-blinded multicenter randomized trial of endovascular brachytherapy after femoropopliteal balloon angioplasty. All patients underwent postdilation angiography, after which the centering catheter was placed in the dilated segment. The patients were randomized and transferred to the radiotherapy unit; the centering catheter was inflated only in patients randomized to receive brachytherapy (18 Gy of gamma radiation delivered to the target site 2 mm from an iridium-192 source axis). Because the local ethics committee required angiography after any interventional procedure (not stipulated in the study protocol), our patients were taken back to the angiosuite for angiography. RESULTS: There were no complications of angioplasty or centering catheter introduction. In the 5 patients randomized to brachytherapy, the median inflation time of the centering catheter was 12.1 minutes (range 9.1-13.3). The completion angiogram following brachytherapy showed peripheral embolization in 2 patients and lesion recoil that required redilation in another; all the complications were confined to the treatment group. CONCLUSIONS: Three of 5 patients who received brachytherapy via a centering catheter demonstrated sequelae on postradiation angiography. Therefore, as in any other peripheral vascular intervention, completion angiography after brachytherapy seems to be essential in preventing delayed diagnosis of procedural complications.

Aged↗