Heidegard Hilbig

Publications of Heidegard Hilbig

• Proliferation, Apoptosis and Expression of Non-Collagenous Proteins: Differences between the Upper and the Lower Jaw Bone in vitro.

  Authors: Hans-Ludwig Graf, Martin Brueckner, Ursula Troeger, Heidegard Hilbig

  Cells, tissues, organs. 04/2011; 195(3):244-51.

  One of the effects observed during several screening studies for osteocompatibility in vitro was that cells derived from the upper and lower jaw exhibited distinct differences regardingOne of the effects observed during several screening studies for osteocompatibility in vitro was that cells derived from the upper and lower jaw exhibited distinct differences regarding proliferation. Therefore, the aim of this study was to examine systematically whether a single osteoblast possesses abilities which are specific to the upper or lower jaw. Both human maxillary and mandibular bone samples without any clinical or radiographic evidence of pathology were obtained from 4 male donors aged between 40 and 45 years. Cells were cultured for up to 25 days to investigate in vitro development. Total and apoptotic cell numbers were estimated by image analysis. Cells were identified as bone-like cells using immunocytochemical determination of bone sialoprotein (BSP) and osteocalcin expression. The number of healthy cells was significantly higher for cells of the lower jaw compared to those of the upper jaw. The number of apoptotic cells showed an inverse pattern. The expression pattern of osseo-inductive BSP correlated with the proliferation rate of the cells. The pattern of osteocalcin expression was related to the number of apoptotic cells. Our findings are new but were anticipated regarding the well-known differences in the healing process around implants in the lower jaw versus the upper jaw. Additionally, a relationship between our results and some diseases of the lower/upper jaw seems obvious. Future work on cell responses to biomaterials should define the origin of the cells more precisely.

• Long-term benefit of human fetal neuronal progenitor cell transplantation in a clinically adapted model after traumatic brain injury.

  Authors: Marco Skardelly, Khaled Gaber, Swen Burdack, Franziska Scheidt, Heidegard Hilbig, Johannes Boltze, Annette Förschler, Sigrid Schwarz, Johannes Schwarz, Jürgen Meixensberger, Martin U Schuhmann

  Journal of neurotrauma. 11/2010; 28(3):401-14.

  Experimental human fetal neural progenitor cell (hfNPC) transplantation has proven to be a promising therapeutic approach after traumatic brain injury (TBI). However, the long-term efficacy andExperimental human fetal neural progenitor cell (hfNPC) transplantation has proven to be a promising therapeutic approach after traumatic brain injury (TBI). However, the long-term efficacy and safety, which are both highly important for clinical translation of this approach, have thus far not been investigated. This study investigated the effect of local (L, 1 × 10(5) cells) and systemic (S, 5 × 10(5) cells) administration of PKH-26-labeled pre-differentiated hfNPCs over a period of 12 weeks, beginning 24 h after severe controlled cortical impact TBI in Sprague-Dawley rats. Accelerating rotarod testing revealed a trend toward functional improvement beginning 1 week after transplantation, and persisting until the end of the experiment. The traumatic lesion volume as quantified by magnetic resonance imaging was smaller in both treatment groups compared to control (C) animals (C = 54.50 mm(3), L = 32 mm(3), S = 37.50 mm(3)). Correspondingly, neuronal (NeuN) staining showed increased neuronal survival at the border of the lesion in both transplanted groups (S = 92.4%; L = 87.2%; 72.5%). Histological analysis of the brain compartments revealed transiently increased angiogenesis and reduced astroglial reaction during the first 4 weeks post-transplantation. PKH-26-positive cells were detected exclusively after local transplantation without any evidence of tumor formation. However, graft differentiation was seen only in very rare cases. In conclusion, transplantation of hfNPCs improved the long-term functional outcome after TBI, diminished trauma lesion size, and increased neuronal survival in the border zone of the lesion. This therapeutic effect was not likely due to cell replacement, but was associated with transiently increased angiogenesis and reduced astrogliosis.

• Expression of Zonulin, c-kit, and Glial Fibrillary Acidic Protein in Human Gliomas.

  Authors: Marco Skardelly, Franz Paul Armbruster, Jürgen Meixensberger, Heidegard Hilbig

  Translational oncology. 09/2009; 2(3):117-20.

  The hallmarks of human malignant gliomas are their marked invasiveness and vascularity. Because angiogenesis and tumor invasion have been associated with extracellular matrix degradation andThe hallmarks of human malignant gliomas are their marked invasiveness and vascularity. Because angiogenesis and tumor invasion have been associated with extracellular matrix degradation and intercellular tight junctions, the involvement of zonulin in glioma biology is in the focus. We selected for histological examination five cases of glioblastoma WHO IV (nomenclature of the World Health Organization) and one case each from astrocytoma WHO III, meningioma WHO III, and meningioma WHO I as control samples. The meningioma WHO I is regarded as benign, whereas the meningioma WHO III is recognized as the transition form of malignant tumors in humans. The visualization of a newly designed antibody against human zonulin was studied in triple-labeling studies using fluorescence immunocytochemistry and compared with the expression of c-kit and glial fibrillary acidic protein in differently developed human gliomas. We found that increasing the expression of c-kit is accompanied by an increase of zonulin expression. Both are correlated to the degree of malignancy of human brain tumors. The expression of zonulin is correlated to the degradation of the blood-brain barrier as revealed by Griffonia simplicifolia lectin. In differently graded tumors, we found differently graded involvement of blood vessels in the tumor development, explaining patients' survival.

• Relaxin as a protective substance in the preserving solution for liver transplantation: spectrophotometric in vivo imaging of local oxygen supply in an isolated perfused rat liver model.

  Authors: Markus U Boehnert, Franz Paul Armbruster, Heidegard Hilbig

  Annals of the New York Academy of Sciences. 05/2009; 1160:320-1.

  Ischemia reperfusion injury (IRI) is a problem in organ transplantation. Relaxin is known to have a protective effect against liver injury caused by IRI. Using a model of isolated perfused rat liver,Ischemia reperfusion injury (IRI) is a problem in organ transplantation. Relaxin is known to have a protective effect against liver injury caused by IRI. Using a model of isolated perfused rat liver, the local oxygen supply in liver tissue was investigated by spectrophotometric in vivo imaging and compared to the protective effect of relaxin shown by immunohistochemical measurement of myeloperoxidase and malonyldialdehyde activities as determinants of oxidative stress. In relaxin-treated liver tissue, spectrophotometry showed a better oxygen supply and decreased myeloperoxidase and malonyldialdehyde activities. Our data suggest that relaxin can influence the oxygen distribution in liver tissue and reduce cell damage caused by IRI.

• Superior olivary complex organization and cytoarchitecture may be correlated with function and catarrhine primate phylogeny.

  Authors: Heidegard Hilbig, Boris Beil, Henrik Hilbig, Josep Call, Hans-Jürgen Bidmon

  Brain structure & function. 02/2009;

  In the mammalian auditory system, the medial nucleus of the trapezoid body and the lateral superior olive (MNTB-LSO system) contribute to binaural intensity processing and lateralization.In the mammalian auditory system, the medial nucleus of the trapezoid body and the lateral superior olive (MNTB-LSO system) contribute to binaural intensity processing and lateralization. Localization precision varies with the sound frequencies. As recency of common ancestry with human beings increases, primates have improved low-frequency sensitivity and reduced sensitivity to higher frequencies. The medial part of the MNTB is devoted to higher frequency processing. Thus, its high-frequency-dependent function is nearly lost in humans and its role in binaural processing as part of the contralateral pathway to the LSO remains questionable. Here, Nissl-stained sections of the superior olivary complex of man (Homo sapiens), bonobo (Pan paniscus), chimpanzee (Pan troglodytes), gorilla (Gorilla gorilla), orangutan (Pongo pygmaeus), gibbon (Hylobates lar), and macaque (Macaca fascicularis) were compared to reveal differences and coincidences. From chimpanzees to humans, the size of the LSO decreased, and the MNTB as a compact nucleus nearly disappears. From chimpanzees to humans, the LSO/MNTB ratio increases dramatically too, whereas the LSO/MSO ratio remains 1.1; a finding that probably corresponds to the phylogenetic proximity between the species.

• Imaging anti-angiogenic treatment response with DCE-VCT, DCE-MRI and DWI in an animal model of breast cancer bone metastasis.

  Authors: Tobias Bäuerle, Sönke Bartling, Martin Berger, Annette Schmitt-Gräff, Heidegard Hilbig, Hans-Ulrich Kauczor, Stefan Delorme, Fabian Kiessling

  European journal of radiology. 12/2008;

  As current classification systems for the assessment of treatment response in bone metastasis do not meet the needs of oncologists, new imaging biomarkers are desirable. Therefore, the diagnosticAs current classification systems for the assessment of treatment response in bone metastasis do not meet the needs of oncologists, new imaging biomarkers are desirable. Therefore, the diagnostic impact of dynamic contrast enhanced (DCE)-volumetric computed tomography (VCT) (descriptive analysis), DCE-MRI (two-compartment model) and diffusion weighted imaging (DWI) for monitoring anti-angiogenic therapy effects of the VEGF antibody bevacizumab in breast cancer bone metastases in rats was studied. Nude rats (n=8 animals treated with bevacizumab and n=9 untreated control rats) with site-specific osteolytic bone metastasis of the hind leg were imaged with a 1.5T clinical MRI-scanner in an animal coil as well as in a volumetric CT-scanner at days 30, 40, 50 and 60 after inoculation of MDA-MB-231 human breast cancer cells. From these data, osteolytic lesion size (OLS), peak enhancement (PE), area under the curve (AUC), amplitude (A), exchange rate constant (k(ep)) and apparent diffusion coefficient (ADC) were determined in bone metastases. Prior to changes in OLS (p</=0.05 at days 50 and 60) there was already a significant decrease in PE, AUC and A (p</=0.05 at days 40-60) in treated animals compared to controls. However, for k(ep) and ADC there were no significant differences between the groups at any time point (p>0.05 at days 40-60). In conclusion, anti-angiogenic treatment response in osteolytic breast cancer bone metastases can be assessed early with surrogate markers of vascularization, while DWI appears to be insensitive.

• Bevacizumab inhibits breast cancer-induced osteolysis, surrounding soft tissue metastasis, and angiogenesis in rats as visualized by VCT and MRI.

  Authors: Tobias Bäuerle, Heidegard Hilbig, Sönke Bartling, Fabian Kiessling, Astrid Kersten, Annette Schmitt-Gräff, Hans-Ulrich Kauczor, Stefan Delorme, Martin R Berger

  Neoplasia (New York, N.Y.). 06/2008; 10(5):511-20.

  The aim of this study was to evaluate the effect of an antiangiogenic treatment with the vascular endothelial growth factor antibody bevacizumab in an experimental model of breast cancer boneThe aim of this study was to evaluate the effect of an antiangiogenic treatment with the vascular endothelial growth factor antibody bevacizumab in an experimental model of breast cancer bone metastasis and to monitor osteolysis, soft tissue tumor, and angiogenesis in bone metastasis noninvasively by volumetric computed tomography (VCT) and magnetic resonance imaging (MRI). After inoculation of MDA-MB-231 human breast cancer cells into nude rats, bone metastasis was monitored with contrast-enhanced VCT and MRI from day 30 to day 70 after tumor cell inoculation, respectively. Thereby, animals of the treatment group (10 mg/kg bevacizumab IV weekly, n = 15) were compared with sham-treated animals (n = 17). Treatment with bevacizumab resulted in a significant difference versus control in osteolytic as well as soft tissue lesion sizes (days 50 to 70 and 40 to 70 after tumor cell inoculation, respectively; P < .05). This observation was paralleled with significantly reduced vascularization in the treatment group as shown by reduced increase in relative signal intensity in dynamic contrast-enhanced MRI from days 40 to 70 (P < .05). Contrast-enhanced VCT and histology confirmed decreased angiogenesis as well as new bone formation after application of bevacizumab. In conclusion, bevacizumab significantly inhibited osteolysis, surrounding soft tissue tumor growth, and angiogenesis in an experimental model of breast cancer bone metastasis as visualized by VCT and MRI.

• Characterization of neuronal subsets surrounded by perineuronal nets in the rhesus auditory brainstem.

  Authors: Heidegard Hilbig, Sandra Nowack, Katrin Boeckler, Hans-Jürgen Bidmon, Karl Zilles

  Journal of anatomy. 06/2007; 210(5):507-17.

  The distribution of perineuronal nets and the potassium channel subunit Kv3.1b was studied in the subdivisions of the cochlear nucleus, the medial nucleus of the trapezoid body, the medial andThe distribution of perineuronal nets and the potassium channel subunit Kv3.1b was studied in the subdivisions of the cochlear nucleus, the medial nucleus of the trapezoid body, the medial and lateral superior olivary nuclei, the lateral lemniscal nucleus and the inferior colliculus of the rhesus monkey. Additional sections were used for receptor autoradiography to visualize the patterns of GABAA and GABAB receptor distribution. The Kv3.1b protein and perineuronal nets [visualized as Wisteria floribunda agglutinin (WFA) binding] were revealed, showing corresponding region-specific patterns of distribution. There was a gradient of labelled perineuronal nets which corresponded to that seen for the intensity of Kv3.1b expression. In the cochlear nucleus intensely and faintly stained perineuronal nets were intermingled, whereas in the medial nucleus of the trapezoid body the pattern changed to intensely stained perineuronal nets in the medial part and weakly labelled nets in its lateral part. In the inferior colliculus, intensely labelled perineuronal nets were arranged in clusters and faintly labelled nets were arranged in sheets. Using receptor autoradiography, GABAB receptor expression in the anterior ventral cochlear nucleus was revealed. The medial part of the medial nucleus of the trapezoid body showed a high number of GABAA binding sites whereas the lateral part exhibited more binding sites for GABAB. In the inferior colliculus, we found moderate GABAB receptor expression. In conclusion, intensely WFA-labelled structures are those known to be functionally involved in high-frequency processing.

• Treatment of bone metastasis induced by MDA-MB-231 breast cancer cells with an antibody against bone sialoprotein.

  Authors: Tobias Bäuerle, Jenny Peterschmitt, Heidegard Hilbig, Fabian Kiessling, Franz P Armbruster, Martin R Berger

  International journal of oncology. 03/2006; 28(3):573-83.

  The extracellular bone matrix protein bone sialoprotein (BSP) is considered to play an important role in the pathogenesis of lytic skeletal lesions which are associated with severe morbidity inThe extracellular bone matrix protein bone sialoprotein (BSP) is considered to play an important role in the pathogenesis of lytic skeletal lesions which are associated with severe morbidity in breast, prostate or lung cancer patients. In addition to in vitro studies, nude rats were implanted with 10(5) MDA-MB-231 cells transfected with GFP into a small branch of the femoral artery. Osteolytic lesions of the respective hind leg were detected by X-ray and CT analysis as well as by immunohistochemistry. Exposure of MDA-MB-231GFP cells in vitro to an antibody against BSP (0-400 microg/ml) decreased proliferation, colony formation and migration of these cells by up to 95, 83 and 89 T/C%, respectively. In nude rats, pre-incubation of MDA-MB-231GFP cells prior to inoculation (25-100 microg/ml) reduced the mean osteolytic lesion size to 22 T/C% after 90 days of observation (p<0.05). Treatment of overt lytic metastasis with the anti-BSP antibody (10 mg/kg) resulted in a significantly smaller mean lesion size of 57 T/C% at the end of the observation period (p<0.05) as well as in new bone formation. Immunohistochemical analysis revealed the presence of BSP in MDA-MB-231GFP cells and in vessel endothelium cells during processes such as migration and invasion. In conclusion, an anti-BSP antibody decreased proliferation, colony formation and migration of MDA-MB-231GFP cells in vitro and reduced osteolysis besides inducing bone formation in a nude rat model.

• Characterization of the rhesus monkey superior olivary complex by calcium binding proteins and synaptophysin.

  Authors: Ivonne Bazwinsky, Hans-Jürgen Bidmon, Karl Zilles, Heidegard Hilbig

  Journal of anatomy. 01/2006; 207(6):745-61.

  This study was performed in order to characterize the main nuclei of the rhesus monkey superior olivary complex by means of antibodies against the calcium binding proteins parvalbumin, calbindin andThis study was performed in order to characterize the main nuclei of the rhesus monkey superior olivary complex by means of antibodies against the calcium binding proteins parvalbumin, calbindin and calretinin and the synaptic vesicle protein synaptophysin. These markers revealed the neuronal morphology and organization of nuclei located within the rhesus monkey superior olivary complex. The architectural details included the distribution of axonal terminals on neurons. The medial superior olivary nucleus was present as a column of neurons. No clear segregation of calretinin-positive terminals was noticed on the medial and lateral dendritic fields of these neurons. The lateral superior olivary nucleus was characterized by a distinct nuclear shape. Calretinin-, parvalbumin- or calbindin-positive terminals contacted somata and dendrites. The medial nucleus of trapezoid body could be clearly differentiated as a distinct region in the rhesus monkey superior olivary complex. Somata of that nucleus showed calbindin- and parvalbumin-labelling whereas somatic calyces of Held were reavealed by calretinin and synaptophysin labelling. The results are discussed with respect to the processing of acoustic information in primate species and their ability to hear high and low frequencies, which is reflected by anatomical correlates.

• Characterization of a rat model with site-specific bone metastasis induced by MDA-MB-231 breast cancer cells and its application to the effects of an antibody against bone sialoprotein.

  Authors: Tobias Bäuerle, Hassan Adwan, Fabian Kiessling, Heidegard Hilbig, Franz P Armbruster, Martin R Berger

  International journal of cancer. Journal international du cancer. 07/2005; 115(2):177-86.

  Metastasis into the skeleton is a serious complication of certain neoplastic diseases such as breast, prostate and lung cancer, but the reasons for this osteotropism are poorly understood. Our aimMetastasis into the skeleton is a serious complication of certain neoplastic diseases such as breast, prostate and lung cancer, but the reasons for this osteotropism are poorly understood. Our aim was to establish a physiologically relevant animal model that is characterized by osteolytic lesions confined to the hind leg of nude rats. For this purpose, we injected 1x10(5) MDA-MB-231 human breast cancer cells transfected with GFP into the superficial epigastric artery, which is an anastomosing vessel between the femoral and iliac arteries. As assessed with the aid of X-rays, computed tomography and immunohistochemisty, osteolytic lesions occurred exclusively in the femur, tibia and fibula of the animals. The tumor take rate was 93% in a series of 96 rats and the increase in lesion size was observed up to 110 days after tumor cell inoculation. When applying this animal model to the effects of an antibody against bone sialoprotein (BSP), a significantly reduced osteolytic lesion size was observed after preincubation of cells (2 hr, 600 microg/ml anti-BSP) prior to intra-arterial tumor cell injection resulting in 19 T/C% at day 60 after tumor implantation (p < 0.05). In addition, the osteolytic lesion size was also significantly reduced after s.c. treatment of the animals with the antibody (20 mg/kg anti-BSPx3 within 5 days after tumor implantation), resulting in 30 T/C% at day 60 after tumor cell implantation (p < 0.05). In conclusion, the novel rat model for site-specific osteolytic lesions provides in vivo evidence that preincubation of MDA-MB-231GFP cells and treatment of rats after tumor implantation with an antibody against BSP significantly reduces the size of lytic lesions in bone.

• Relaxin as an additional protective substance in preserving and reperfusion solution for liver transplantation, shown in a model of isolated perfused rat liver.

  Authors: Markus U Boehnert, Heidegard Hilbig, Franz P Armbruster

  Annals of the New York Academy of Sciences. 06/2005; 1041:434-40.

  Reperfusion injury is a problem in organ transplantation. Relaxin causes vessel dilation and inhibition of platelet and mast cell activation. The study investigates the protective effect of relaxinReperfusion injury is a problem in organ transplantation. Relaxin causes vessel dilation and inhibition of platelet and mast cell activation. The study investigates the protective effect of relaxin on liver tissue against cell damage during organ preservation and reperfusion. Liver transplantation was simulated in a model of isolated perfused rat liver. Relaxin was applicated during reperfusion and/or preservation. To quantify cell damage, we examined the perfusate for malonyldialdehyde (MDA) and myeloperoxidase activity (MPO), and liver tissue underwent immunohistochemical study. Relaxin as an additional substance in preserving/reperfusion solution decreases MPO and MDA levels in the perfusate and immunohistochemical study. Relaxin seems to have a protective effect against cell damage in ischemia and reperfusion injury.

• Effects of dental implant surfaces on the expression of bone sialoprotein in cells derived from human mandibular bone.

  Authors: Heidegard Hilbig, Thomas Wiener, Franz Paul Armbruster, Assegedech Bekele, Mathias Kirsten, Hans-Ludwig Graf

  Medical science monitor : international medical journal of experimental and clinical research. 05/2005; 11(4):BR111-5.

  BACKGROUND: Protein adsorption is believed to be the first event that takes place after contact of natural tissue with an artificial surface. Bone sialoprotein (BSP) is one of the majorBACKGROUND: Protein adsorption is believed to be the first event that takes place after contact of natural tissue with an artificial surface. Bone sialoprotein (BSP) is one of the major noncollagenous proteins in the extracellular matrix of bone. The expression of BSP coincides with initial bone mineralization and is believed to be a center of crystallization for hydroxyapatite formation. MATERIAL/METHODS: We used a variety of four differently designed dental implant surfaces (SLA, CPT, ANOX, TICER) to investigate the effects on the development of adult human mandibular bone at days 5, 10, 15, 20, and 25 in vitro. The time course of the expression of BSP and labeling of fibroblasts was visualized immunohistochemically. The distribution patterns of cells were determined semiquantitatively on both the surface and the tissue-implant borderline. RESULTS: BSP immunoresponse increased from day 5 before decreasing after day 15 in vitro. The distribution of BSP-expressing cells changed during that time. Cell counts revealed that the time course of the settlement of cells depended on the design of the surface of the implant. The design of the border of the implant affected both the cell distribution patterns and the survivals of cells to a higher degree than did the design of the implant surface. CONCLUSIONS: Investigation of novel biomaterials for bone engineering represents an essential area for the design of tissue-engineering strategies. The hydroxyapatite-based implant material TICER could be a good scaffold to guide and promote the regeneration of bone tissue.

• Angiopoietin-1 promotes neurite outgrowth from dorsal root ganglion cells positive for Tie-2 receptor.

  Authors: Joanna Kosacka, Maciej Figiel, Jürgen Engele, Heidegard Hilbig, Mariusz Majewski, Katharina Spanel-Borowski

  Cell and tissue research. 05/2005; 320(1):11-9.

  The effects of vascular factors on the nervous system are still poorly investigated. Angiopoietin-1 (Ang-1), an endothelial cell growth factor with influences on blood vessel stabilization, has beenThe effects of vascular factors on the nervous system are still poorly investigated. Angiopoietin-1 (Ang-1), an endothelial cell growth factor with influences on blood vessel stabilization, has been recently reported to prevent apoptosis in a neuroblastoma cell line via a pathway dependent on Tie-2 receptor. The present study focuses on the effect of Ang-1 on cultured dorsal root ganglion (DRG) cells isolated from 1-day-old rats. Three-day-old DRG cultures were exposed to Ang-1 treatment under serum-free condition for another 5 days and stained with antibodies against neurofilament (NF) 200 protein. Neurite length and density increased compared with those of controls. Double-immunofluorescence staining demonstrated the co-localization of the Tie-2 receptor in some NF-200-positive perikarya. The reverse transcription/polymerase chain reaction technique identified Tie-2 receptor mRNA in intact DRG and in Ang-1-stimulated DRG cell cultures, but not in a Schwann cell line or in primary astrocyte cultures. Western blotting confirmed that the expression of NF 68 protein in cultures treated with Ang-1 or nerve growth factor was higher than that in cultures treated with medium alone. When the Tie-2 receptor was blocked with anti-Tie-2 receptor antibody, neurite outgrowth was severely impeded. Induction of trkA-receptor protein expression was observed to be dependent on the presence of Tie-2 receptors. We conclude that Ang-1 promotes neurite outgrowth from DRG cells positive for Tie-2 receptor. The signalling pathway appears to involve transactivation of the trkA receptor.

• Influence of post-mortem delay and storage temperature on the immunohistochemical detection of antigens in the CNS of mice.

  Authors: Heidegard Hilbig, Hans-Jürgen Bidmon, Oliver Till Oppermann, Torsten Remmerbach

  Experimental and toxicologic pathology : official journal of the Gesellschaft für Toxikologische Pathologie. 01/2005; 56(3):159-71.

  The aim of this work was to compare the results of histochemical and immunohistochemical methods using mouse brains which were fixed with various post-mortem delays and storage temperatures (at aThe aim of this work was to compare the results of histochemical and immunohistochemical methods using mouse brains which were fixed with various post-mortem delays and storage temperatures (at a constant 4 degrees C or 22 degrees C, or at gradually decreasing post-mortem temperatures, mimicking conditions of human corpse). We studied the effects of post-mortem delay on glial fibrillary acidic protein, extracellular matrix components to which Wisteria floribunda agglutinin binds, non-phosphorylated neurofilament H, synaptophysin, calbindin and nitric oxide synthase isoenzymes. At the light microscopic level first signs of post-mortem changes were detectable after 6 h. Glial fibrillary acidic protein was most affected by post-mortem delay since its immunoreactivity increased dramatically with increasing post-mortem delay. N-acetylgalactosamines-beta1 labeled lectin binding sites, calbindin and intraneuronal non-phosphorylated neurofilament H seemed to be stable up to 12 h post-mortem. Storage temperature influenced the NADPH-d activity and the content of synaptophysin immunoreactivity to higher degree than all of the other parameters. We found only marginal differences of alterations comparing neocortex, hippocampus and corpus callosum. Our results indicate that different antigens are affected differently by the ongoing catabolic processes during post-mortem delay.

• Characterization of the human superior olivary complex by calcium binding proteins and neurofilament H (SMI-32).

  Authors: Ivonne Bazwinsky, Heidegard Hilbig, Hans-Jürgen Bidmon, Rudolf Rübsamen

  The Journal of comparative neurology. 03/2003; 456(3):292-303.

  This study provides a morphologic characterization of the human superior olivary complex as revealed by immunohistochemistry by using antibodies against the calcium binding proteins parvalbumin,This study provides a morphologic characterization of the human superior olivary complex as revealed by immunohistochemistry by using antibodies against the calcium binding proteins parvalbumin, calbindin, calretinin, and the nonphosphorylated neurofilament H SMI-32. By combining these markers, it was possible to establish the neuronal architecture and details of the morphologic organization (including axonal terminals) of the different nuclei. The medial superior olivary nucleus is formed by a sheet of parallel-oriented cells. A clear segregation of axon terminals was noticed on the medially and laterally oriented dendrites of the mostly bipolar neurons. The lateral superior olivary nucleus lacked a distinct nuclear shape but was formed by several patches of rather irregularly arranged neurons. Calretinin or parvalbumin immunoreactive afferent terminals were observed which contacted somata or dendrites of these neurons. The immunolabeling also revealed the boundaries of the dorsal periolivary nucleus and morphologic detail of its neurons. A coherent nuclear structure that could be addressed as the medial nucleus of the trapezoid body was not identified by any single one or by combinations of the markers used. The data were also used to establish a three-dimensional-reconstruction of the three major subnuclei of the superior olivary complex. The results are discussed with respect to the possible role of the superior olivary complex in the processing of spatial acoustic information in the azimuthal plane.

• Enriched environmental conditions reverse age-dependent gliosis and losses of neurofilaments and extracellular matrix components but do not alter lipofuscin accumulation in the hindlimb area of the aging rat brain.

  Authors: Heidegard Hilbig, Hans-Jürgen Bidmon, Susanne Steingrüber, Heinrich Reinke, Hubert R Dinse

  Journal of chemical neuroanatomy. 04/2002; 23(3):199-209.

  We provide a description of a correlation of lipofuscin accumulation and expression of glial fibrillary acidic protein in the cerebral cortex of aged rats. Glial fibrillary acidic protein showed aWe provide a description of a correlation of lipofuscin accumulation and expression of glial fibrillary acidic protein in the cerebral cortex of aged rats. Glial fibrillary acidic protein showed a complementary distribution pattern to perineuronal nets, visualized with Wisteria floribunda agglutinin. With progressing age (12-36 months), a strong increase of lipofuscin and gliosis occurred in functionally characterized cortical areas, whereas a concomitant, area-specific loss of perineuronal nets was found in the cortical somatosensory representation of the hindlimbs. In contrast to lipofuscin accumulation and increased gliosis, the loss of perineuronal nets and the reduction of non-phosphorylated neurofilament H were in part reduced or prevented by housing the animals under enriched environmental conditions between 33 and 36 months of age. Especially the reduction of astrocytosis by 20% which coincided with a reduction in the loss of extracellular matrix components involved in forming the glia-neuron-interface demonstrates, that the aging cortex retains its potential for functional plasticity.

• In contrast to neuronal NOS-I, the inducible NOS-II expression in aging brains is modifled by enriched environmental conditions.

  Authors: Heidegard Hilbig, Josefin Holler, Hubert R Dinse, Hans-Jïrgen Bidmon

  Experimental and toxicologic pathology : official journal of the Gesellschaft für Toxikologische Pathologie. 03/2002; 53(6):427-31.

  Changes in the expression of the nitric oxide synthase isoforms (NOS) have been implicated in age-related neurodegeneration, although mechanisms responsible for specific differences such as reducedChanges in the expression of the nitric oxide synthase isoforms (NOS) have been implicated in age-related neurodegeneration, although mechanisms responsible for specific differences such as reduced cognitive and motor functions are not well understood. Immunohistochemical examination of the age-dependent NOS-I and NOS-II expression in aging rats at 3, 12, 24, 36 months of age as well as in 36 months old rats who spent their last 3 months in an enriched environment, revealed a significant decrease in NOS-I expression with aging. In contrast, NOS-II increased at 36 months. Old rats kept under enriched environmental conditions, however, showed strongly reduced NOS-II expression. These results indicate that both NOS-I and II may contribute to age-related degenerative processes, but in contrast to NOS-I the age-dependent changes of NOS-II are reduced or even reversed by stimulating environmental conditions.

• Nitric oxide synthase isoenzymes during in vitro development of rat neuronal and human non-neuronal cells

  Authors: Heidegard Hilbig, Heike Franke, Hans-Jürgen Bidmon, Peter Illes

  Neuroscience Letters.

  We studied the expression of neuronal (n), endothelial (e) and inducible (i) nitric oxide synthase (NOS) in cell cultures of rat mesencephalic neurons (embryonic day 14), human keratinocytes fromWe studied the expression of neuronal (n), endothelial (e) and inducible (i) nitric oxide synthase (NOS) in cell cultures of rat mesencephalic neurons (embryonic day 14), human keratinocytes from juvenile epidermis, human endothelial cells from juvenile coronary arteries, and human osteoblasts. All cell types were cultured for 5, 10 or 15 days. During proliferation (round cells without processes), the intracellular distribution and the intracellular amount of the calcium-dependent NOS isoforms (n- and e-NOS) did not change whereas the calcium-independent i-NOS changed from a cytosolic distribution pattern to compartmentalized distribution. A striking decrease of i-NOS immunoreactivity was measured by means of image analysis. Our results support the opinion that i-NOS acts as a switch between proliferation and differentiation of cells.

• Influence of post-mortem delay and storage temperature on the immunohistochemical detection of antigens in the CNS of mice

  Authors: Heidegard Hilbig, Hans-Jürgen Bidmon, Oliver Till Oppermann, Torsten Remmerbach

  Experimental and Toxicologic Pathology.

  The aim of this work was to compare the results of histochemical and immunohistochemical methods using mouse brains which were fixed with various post-mortem delays and storage temperatures (at aThe aim of this work was to compare the results of histochemical and immunohistochemical methods using mouse brains which were fixed with various post-mortem delays and storage temperatures (at a constant 4 °C or 22 °C, or at gradually decreasing post-mortem temperatures, mimicking conditions of human corpse). We studied the effects of post-mortem delay on glial fibrillary acidic protein, extracellular matrix components to which Wisteria floribunda agglutinin binds, non-phosphorylated neurofilament H, synaptophysin, calbindin and nitric oxide synthase isoenzymes. At the light microscopic level first signs of post-mortem changes were detectable after 6 h. Glial fibrillary acidic protein was most affected by post-mortem delay since its immunoreactivity increased dramatically with increasing post-mortem delay. N-acetylgalactosamines-ß1 labeled lectin binding sites, calbindin and intraneuronal non-phosphorylated neurofilament H seemed to be stable up to 12 h post-mortem. Storage temperature influenced the NADPH-d activity and the content of synaptophysin immunoreactivity to higher degree than all of the other parameters. We found only marginal differences of alterations comparing neocortex, hippocampus and corpus callosum. Our results indicate that different antigens are affected differently by the ongoing catabolic processes during post-mortem delay.