Joachim Weis

Publications of Joachim Weis

• Efficacy of clinically relevant temozolomide dosing schemes in glioblastoma cancer stem cell lines.

  Authors: Dagmar Beier, Beate Schriefer, Konstantin Brawanski, Peter Hau, Joachim Weis, Jörg B Schulz, Christoph P Beier

  Journal of neuro-oncology. 04/2012;

  The effectiveness of temozolomide (TMZ) dosing schemes and the "rechallenge" of recurrent glioblastoma (GBM) with TMZ are controversial. We therefore compared the efficacy of different TMZ dosingThe effectiveness of temozolomide (TMZ) dosing schemes and the "rechallenge" of recurrent glioblastoma (GBM) with TMZ are controversial. We therefore compared the efficacy of different TMZ dosing schemes against GBM cancer stem cell (CSC) lines in vitro. In O(6)-methyl-guanidine-methyl-transferase (MGMT)-negative CSC lines, all schedules (1 day on/27 days off, 5 days on/23 days off, 7 days on/7 days off, 21 days on/7 days off, continuous low-dose TMZ) depleted clonogenic cells. In TMZ-resistant CSC lines, the 7 days on/7 days off scheme showed higher toxicity as compared with the other schemes. However, clinically feasible concentrations remained ineffective in highly resistant CSC lines. In addition, none of the schedules induced long-term depletion of clonogenic cells even at the highest concentrations (up to 250 μM). After sublethal TMZ treatment for 5 days, TMZ rechallenge of recovering CSC lines remained effective. Our data advocate CSC lines as in vitro model to address clinical questions. Using this model, our data suggest the effectiveness of TMZ in MGMT-negative CSC lines and support the concept of TMZ rechallenge. The 7 days on/7 days off scheme consistently showed the best activity of all schedules in TMZ-resistant CSC lines.

• Early muscle and brain ultrastructural changes in polymerase gamma 1-related encephalomyopathy.

  Authors: Kay W Nolte, Sonja Trepels-Kottek, Dagmar Honnef, Joachim Weis, Christian G Bien, Andreas van Baalen, Klaus Ritter, Birgit Czermin, Sabine Rudnik-Schöneborn, Norbert Wagner, Martin Häusler

  Neuropathology : official journal of the Japanese Society of Neuropathology. 04/2012;

  Mutations affecting the mitochondrial DNA-polymerase gamma 1 (POLG1) gene have been shown to cause Alpers-Huttenlocher disease. Ultrastructural data on brain and muscle tissue are rare. We report onMutations affecting the mitochondrial DNA-polymerase gamma 1 (POLG1) gene have been shown to cause Alpers-Huttenlocher disease. Ultrastructural data on brain and muscle tissue are rare. We report on ultrastructural changes in brain and muscle tissue of two sisters who were compound heterozygous for the c.2243G>C and c.1879C>T POLG1 mutations. Patient 1 (16 years) presented with epilepsia partialis continua that did not respond to antiepileptic treatment. Neuroimaging showed right occipital and bithalamic changes. Light microscopy from a brain biopsy performed after 3 weeks suggested chronic encephalitis showing astro- and microgliosis as well as perivascular CD8-positive T-cells. However, immunosuppressive therapy failed to improve her condition. When her 17-year-old sister (patient 2) also developed epilepsy, an intensified search for metabolic diseases led to the diagnosis. On electron microscopy mitochondrial abnormalities mainly affecting neurons were detected in the brain biopsy of patient 1, including an increase in number and size, structural changes and globoid inclusions. In patient 2, light and electron microscopy on a muscle biopsy confirmed a mitochondrial myopathy, also revealing an increase in mitochondrial size and number, as well as globoid inclusions. Neurons may be the primary target of mitochondrial dysfunction in brains of patients with Alpers disease related to POLG1 mutations. During early disease stages, brain histopathology may be misleading, showing reactive inflammatory changes.

• Dexmedetomidine is neuroprotective in an in vitro model for traumatic brain injury.

  Authors: Marc Schoeler, Philip D Loetscher, Rolf Rossaint, Astrid V Fahlenkamp, Georg Eberhardt, Steffen Rex, Joachim Weis, Mark Coburn

  BMC neurology. 04/2012; 12(1):20.

  ABSTRACT: BACKGROUND: The alpha2-adrenoreceptor agonist dexmedetomidine is known to provide neuroprotection under ischemic conditions. In this study we investigated whether dexmedetomidine has aABSTRACT: BACKGROUND: The alpha2-adrenoreceptor agonist dexmedetomidine is known to provide neuroprotection under ischemic conditions. In this study we investigated whether dexmedetomidine has a protective effect in an in vitro model for traumatic brain injury. METHODS: Organotypic hippocampal slice cultures were subjected to a focal mechanical trauma and then exposed to varying concentrations of dexmedetomidine. After 72 h cell injury was assessed using propidium iodide. In addition, the effects of delayed dexmedetomidine application, of hypothermia and canonical signalling pathway inhibitors were examined. RESULTS: Dexmedetomidine showed a protective effect on traumatically injured hippocampal cells with a maximum effect at a dosage of 1 muM. This effect was partially reversed by the simultaneous administration of the ERK inhibitor PD98059. CONCLUSION: In this TBI model dexmedetomidine had a significant neuroprotective effect. Our results indicate that activation of ERK might be involved in mediating this effect.

• Combining xenon and mild therapeutic hypothermia preserves neurological function after prolonged cardiac arrest in pigs*.

  Authors: Michael Fries, Anne Brücken, Ayşegül Cizen, Maren Westerkamp, Céline Löwer, Jan Deike-Glindemann, Nora K Schnorrenberger, Steffen Rex, Mark Coburn, Kay W Nolte, Joachim Weis, Rolf Rossaint, Matthias Derwall

  Critical care medicine. 04/2012; 40(4):1297-303.

  : Despite the introduction of mild therapeutic hypothermia into postcardiac arrest care, cerebral and myocardial injuries represent the limiting factors for survival after cardiac arrest.: Despite the introduction of mild therapeutic hypothermia into postcardiac arrest care, cerebral and myocardial injuries represent the limiting factors for survival after cardiac arrest. Administering xenon may confer an additional neuroprotective effect after successful cardiopulmonary resuscitation due to its ability to stabilize cellular calcium homeostasis via N-methyl-D-aspartate-receptor antagonism. : In a porcine model, we evaluated effects of xenon treatment in addition to therapeutic hypothermia on neuropathologic and functional outcomes after cardiopulmonary resuscitation. : Prospective, randomized, laboratory animal study. : Fifteen male pigs. : Following 10 mins of cardiac arrest and 6 mins of cardiopulmonary resuscitation, ten pigs were randomized to receive either mild therapeutic hypothermia (33°C for 16 hrs) or mild therapeutic hypothermia 1 xenon (70% for 1 hr). Five animals served as normothermic controls. : Gross hemodynamic variables were measured using right-heart catheterization. Neurocognitive performance was evaluated for 5 days after cardiopulmonary resuscitation using a neurologic deficit score before the brains were harvested for histopathological analysis. All animals survived the observation period in the mild therapeutic hypothermia 1 xenon group while one animal in each of the other two groups died. Mild therapeutic hypothermia 1 xenon preserved cardiac output during the induction of mild therapeutic hypothermia significantly better than did mild therapeutic hypothermia alone (4.6 6 0.6 L/min vs. 3.2 6 1.6 L/min, p # .05). Both treatment groups showed significantly fewer necrotic lesions in the cerebral cortex, caudate nucleus, putamen, and in hippocampal sectors CA1 and CA3/4. However, only the combination of mild therapeutic hypothermia and xenon resulted in reduced astrogliosis in the CA1 sector and diminished microgliosis and perivascular inflammation in the putamen. Clinically, only the mild therapeutic hypothermia 1 xenon-treated animals showed significantly improved neurologic deficit scores over time (day 1 = 59.0 6 27.0 vs. day 5 = 4.0 6 5.5, p ø .05) as well as in comparison to the untreated controls on days 3 through 5 after cardiopulmonary resuscitation. : These results demonstrate that even a short exposure to xenon during induction of mild therapeutic hypothermia results in significant improvements in functional recovery and ameliorated myocardial dysfunction. (Crit Care Med 2012; 40:-1303).

• The Modified Glasgow Outcome Score for the prediction of outcome in patients after cardiac arrest: a prospective clinical proof of concept study.

  Authors: Obaida R Rana, Jörg W Schröder, Julia S Kühnen, Esra Saygili, Christopher Gemein, Matthias D H Zink, Patrick Schauerte, Johannes Schiefer, Robert H G Schwinger, Joachim Weis, Nikolaus Marx, Malte Kelm, Christian Meyer, Erol Saygili

  Clinical research in cardiology : official journal of the German Cardiac Society. 02/2012;

  The Glasgow-Pittsburgh cerebral performance categories (GP-CPC) and the Glasgow Outcome Score (GOS) have been used to categorize patients according to their neurological outcome for prognosticThe Glasgow-Pittsburgh cerebral performance categories (GP-CPC) and the Glasgow Outcome Score (GOS) have been used to categorize patients according to their neurological outcome for prognostic predictors in patients after cardiac arrest (CA). We postulated that inclusion of deaths without knowing the cerebral status into the group of patients with poor outcome after CA using the GP-CPC and GOS will lead to dilution of the prognostic power of the investigated biochemical marker. The present study was conducted to verify this issue by employing a modified outcome score, which we termed as Modified Glasgow Outcome Score (MGOS). In the present study, 97 patients were enrolled in a prospective manner. Serum NSE and S100B levels were measured daily for 7 days after admission to the intensive care unit. Neurological outcome was assessed by employing the GOS and MGOS after 6 months. By employing the GOS, 46 patients were categorized into the group of patients with poor outcome and 51 patients survived with good neurological outcome. Patients who died without certified brain damage or with unknown cerebral status after CA (n = 20) were separated from patients with poor outcome in the MGOS. The magnitude of NSE (S100B) elevation in patients with poor outcome categorized by the MGOS was approximately 1.7-fold (1.5) higher as compared with patients divided by the GOS. The mean calculated sensitivities and area under the curve values of NSE and S100B predicting poor outcome classified by the MGOS were significantly higher as compared with the GOS. Conclusively, inclusion of deaths without certified brain damage or with unknown cerebral status into the group of patients with poor outcome will lead to underestimation of the prognostic power of investigated biochemical markers such as NSE and S100B. The MGOS will help to avoid this bias.

• SOX10 mutation with peripheral amyelination and developmental disturbance of axons.

  Authors: Kathleen Parthey, Malte Kornhuber, Christian Kunze, Dorothea Wand, Kay W Nolte, Stefan Nikolin, Joachim Weis, J Michael Schröder

  Muscle & nerve. 02/2012; 45(2):284-90.

  In this study we describe a case of a term infant with the neurological variant of Waardenburg syndrome type 4 (i.e., PCWH = peripheral demyelinating neuropathy, central dysmyelinatingIn this study we describe a case of a term infant with the neurological variant of Waardenburg syndrome type 4 (i.e., PCWH = peripheral demyelinating neuropathy, central dysmyelinating leukodystrophy, Waardenburg syndrome, and Hirschsprung disease, as defined in OMIM #609136) due to a novel heterozygous base exchange (c.671C>G) in exon 4 of SOX10. Magnetic resonance imaging suggested central myelin deficiency with cerebral and cerebellar hypoplasia. Hirschsprung disease was confirmed by rectal biopsy. Sural nerve biopsy revealed hypoplasia due to amyelination (with the exception of a single, small myelinated fiber) and severe reduction in the number of axons.

• Cetuximab induces mitochondrial translocalization of EGFRvIII, but not EGFR: involvement of mitochondria in tumor drug resistance?

  Authors: Agnieszka Dreier, Stefan Barth, Anand Goswami, Joachim Weis

  Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine. 02/2012; 33(1):85-94.

  Dysregulation of growth factor receptors such as the epidermal growth factor receptor (EGFR) and of its truncated form EGFRvIII is frequently found in human tumors. EGFRvIII is a promising target forDysregulation of growth factor receptors such as the epidermal growth factor receptor (EGFR) and of its truncated form EGFRvIII is frequently found in human tumors. EGFRvIII is a promising target for selective molecular tumor therapy because it is exclusively expressed by tumor cells. Cetuximab/Erbitux is a monoclonal antibody which targets EGFR and EGFRvIII. The effects of cetuximab on EGFRvIII but still the exact function and mechanism of cetuximab in relation to EGFR and EGFRvIII are incompletely understood. Therefore, we investigated the influence of cetuximab on EGFRvIII signaling and cellular survival. We found that cetuximab leads to increased internalization of EGFRvIII in NR6M cells but is unable to inhibit neither the activation of EGFRvIII nor its downstream signaling pathways. Incubation with cetuximab also did not alter the survival and proliferation of EGFRvIII-expressing cells. However, it caused increased mitochondrial activity and an increase in co-localization of EGFRvIII with mitochondria. These results demonstrate that interaction of EGFRvIII with mitochondria could play a role in survival of cetuximab-treated NR6M cells. Thus, a role of mitochondria in resistance to cetuximab has to be considered.

• Role for CD74 and CXCR4 in clathrin-dependent endocytosis of the cytokine MIF.

  Authors: Verena Schwartz, Alexander Krüttgen, Joachim Weis, Christian Weber, Tammo Ostendorf, Hongqi Lue, Jürgen Bernhagen

  European journal of cell biology. 10/2011;

  Macrophage migration inhibitory factor (MIF) is a pleiotropic cytokine that plays a role in innate and adaptive immunity. Depending on the cellular context and disease state, MIF signaling isMacrophage migration inhibitory factor (MIF) is a pleiotropic cytokine that plays a role in innate and adaptive immunity. Depending on the cellular context and disease state, MIF signaling is mediated by its receptors CXCR2, CXCR4 and/or CD74. Although it is known that MIF is endocytosed, the exact mechanism has remained unknown. In exploring the mechanism of MIF endocytosis with biologically active Alexa(546)MIF, pathway-specific inhibitors (monodansylcadaverine, MDC; chlorpromazine, CPZ; dynasore; dominant-negative dynamin, bafilomycin, nocodazole) and receptor overexpression and blockade approaches, we identified a clathrin/dynamin-dependent endocytosis pathway as the main track for MIF internalization. MIF endocytosis was rapid and colocalization with both early and late endosomal vesicles in a microtubule- and acidification-dependent manner was observed. LDL endocytosis (which is clathrin-mediated) served as a control and was similarly inhibited by MDC or dynasore. When MIF endocytosis was compared to that of transferrin, acetylated LDL, and choleratoxin B (the latter internalized by a clathrin-independent pathway) by colocalization studies, the MIF internalization pathway clearly resembled that of LDL but also shared early trafficking with transferrin, whereas no colocalization with choleratoxin was noted. To identify the receptors involved in MIF endocytosis, we focused on CD74 and CXCR4 which form a heteromeric complex. Ectopic overexpression of CD74 in HEK293 and HeLa cells, which do not endogenously express CD74, led to a marked acceleration of MIF endocytosis while pharmacological blockade of CXCR4, which is endogenously expressed on these cells, with AMD3100 led to a 20% reduction of MIF endocytosis in HEK293-CD74 transfectants, whereas in untransfected cells, a blockade of 40% was observed. Of note, both CD74 and CXCR4 strongly colocalize with Alexa(546)MIF both on the plasma membrane and in endosomal compartments. Moreover, MIF-stimulated AKT signaling, which was previously shown to involve both CD74 and CXCR4, was reduced by endocytosis inhibitors, indicating that MIF signaling is at least in part due to endosomal signaling mechanisms. Thus, MIF uptake follows a rapid LDL-like, clathrin- and dynamin-dependent endocytosis pathway, which is dependent on the receptors CD74 and CXCR4 and leads to the initiation of endosomal signaling responses.

• Rate and irregularity of electrical activation during atrial fibrillation affect myocardial NGF expression via different signalling routes.

  Authors: Erol Saygili, Obaida R Rana, Claudia Günzel, Gediminas Rackauskas, Esra Saygili, Fawad Noor-Ebad, Christopher Gemein, Matthias D Zink, Robert H G Schwinger, Karl Mischke, Joachim Weis, Nikolaus Marx, Patrick Schauerte

  Cellular signalling. 08/2011; 24(1):99-105.

  An irregular ventricular response during atrial fibrillation (AF) has been shown to mediate an increase in sympathetic nerve activity in human subjects. The molecular mechanisms remain unclear. ThisAn irregular ventricular response during atrial fibrillation (AF) has been shown to mediate an increase in sympathetic nerve activity in human subjects. The molecular mechanisms remain unclear. This study aimed to investigate the impact of rate and irregularity on nerve growth factor (NGF) expression in cardiomyocytes, since NGF is known to be the main contributor to cardiac sympathetic innervation density. Cell cultures of neonatal rat ventricular myocytes were electrically stimulated for 48 h with increasing rates (0, 5 and 50 Hz) and irregularity (standard deviation (SD)=5%, 25% and 50% of mean cycle length). Furthermore, we analyzed the calcineurin-NFAT and the endothelin-1 signalling pathways as possible contributors to NGF regulation during arrhythmic stimulation. We found that the increase of NGF expression reached its maximum at the irregularity of 25% SD by 5 Hz (NGF: 5 Hz 0% SD=1 vs. 5Hz 25% SD=1.57, P<0.05). Specific blockade of the ET-A receptor by BQ123 could abolish this NGF increase (NGF: 5 Hz 25% SD+BQ123=0.66, P<0.05). High frequency electrical field stimulation (HFES) with 50 Hz decreased the NGF expression in a significant manner (NGF: 50Hz=0.55, P<0.05). Inhibition of calcineurin-NFAT signalling with cyclosporine-A or 11R-VIVIT abolished the HFES induced NGF down-regulation (NGF: 50 Hz+CsA=1.14, P<0.05). In summary, this study reveals different signalling routes of NGF expression in cardiomyocytes exposed to increasing rates and irregularity. Whether this translates into different degrees of NGF expression and possibly neural sympathetic growth in various forms of ventricular rate control during AF remains to be elucidated in further studies.

• Mechanical stretch of sympathetic neurons induces VEGF expression via a NGF and CNTF signaling pathway.

  Authors: Erol Saygili, Maimouna Pekassa, Esra Saygili, Gediminas Rackauskas, Dorothee Hommes, Fawad Noor-Ebad, Christopher Gemein, Matthias D H Zink, Robert H G Schwinger, Joachim Weis, Nikolaus Marx, Patrick Schauerte, Obaida R Rana

  Biochemical and biophysical research communications. 06/2011; 410(1):62-7.

  Mechanical stretch has been shown to increase vascular endothelial growth factor (VEGF) expression in cultured myocytes. Sympathetic neurons (SN) also possess the ability to express and secrete VEGF,Mechanical stretch has been shown to increase vascular endothelial growth factor (VEGF) expression in cultured myocytes. Sympathetic neurons (SN) also possess the ability to express and secrete VEGF, which is mediated by the NGF/TrkA signaling pathway. Recently, we demonstrated that SN respond to stretch with an upregulation of nerve growth factor (NGF) and ciliary neurotrophic factor (CNTF). Whether stretch increases neuronal VEGF expression still remains to be clarified. Therefore, SN from the superior cervical ganglia of neonatal Sprangue Dawley rats were exposed to a gradual increase of stretch from 3% up to 13% within 3days (3%, 7% and 13%). Under these conditions, the expression and secretion of VEGF was analyzed. Mechanical stretch significantly increased VEGF mRNA and protein expression (mRNA: control=1 vs. stretch=3.1; n=3/protein: control=1 vs. stretch=2.7; n=3). ELISA experiments to asses VEGF content in the cell culture supernatant showed a time and dose dependency in VEGF increment due to stretch. NGF and CNTF neutralization decreased stretch-induced VEGF augmentation in a significant manner. This response was mediated in part by TrkA receptor activation. The stretch-induced VEGF upregulation was accompanied by an increase in HIF-1α expression. KDR levels remained unchanged under conditions of stretch, but showed a significant increase due to NGF neutralization. In summary, SN respond to stretch with an upregulation of VEGF, which is mediated by the NGF/CNTF and TrkA signaling pathway paralleled by HIF-1α expression. NGF signaling seems to play an important role in regulating neuronal KDR expression.

• Age-related regional differences in cardiac nerve growth factor expression.

  Authors: Erol Saygili, Rahel Kluttig, Obaida R Rana, Esra Saygili, Christopher Gemein, Matthias D Zink, Gediminas Rackauskas, Joachim Weis, Robert H G Schwinger, Nikolaus Marx, Patrick Schauerte

  Age (Dordrecht, Netherlands). 05/2011;

  Age has been identified as an independent risk factor for cardiovascular diseases. A shift of the cardiac autonomic nervous system towards an increase in sympathetic tone has been reported in theAge has been identified as an independent risk factor for cardiovascular diseases. A shift of the cardiac autonomic nervous system towards an increase in sympathetic tone has been reported in the elderly. Nerve growth factor (NGF) is the main neurotrophic factor that increases the sympathetic activity of the heart. If there is a shift of NGF expression in old compared to young cardiomyocytes and whether there are regional differences in the heart still remain unclear. Therefore, we chose a rat model of different-aged rats (3-4 days = neonatal, 6-8 weeks = young, 20-24 months = old), and isolated cardiomyocytes from the left and the right atrium (LA, RA), as well as from the left and the right ventricle (LV, RV), were used to determine NGF expression on mRNA and protein levels. In neonatal, young, and old rats, NGF amount in LA and RA was significantly lower as compared to LV and RV. In young and old rats, we found significant higher NGF protein levels in LA compared to RA. In addition, both atria showed an increase in NGF expression between age groups neonatal, young, and old. In both ventricles, we observed a significant decrease in NGF expression from neonatal to young rats and a significant increase from young to old rats. The highest NGF amount in LV and RV was observed in neonatal rats. Regarding tyrosine kinase A receptor (TrkA) expression, the main receptor for NGF signaling, both atria showed the largest expression in old rats; while in LV and RV, TrkA was expressed mainly in young rats. These results point to a contribution of nerve growth factors to the change of autonomic tone observed in elderly patients.

• Chronic electrical neuronal stimulation increases cardiac parasympathetic tone by eliciting neurotrophic effects.

  Authors: Obaida R Rana, Erol Saygili, Christopher Gemein, Matthias D H Zink, Alexandra Buhr, Esra Saygili, Karl Mischke, Kay W Nolte, Joachim Weis, Christian Weber, Nikolaus Marx, Patrick Schauerte

  Circulation research. 03/2011; 108(10):1209-19.

  Recently, we provided a technique of chronic high-frequency electric stimulation (HFES) of the right inferior ganglionated plexus for ventricular rate control during atrial fibrillation in dogs andRecently, we provided a technique of chronic high-frequency electric stimulation (HFES) of the right inferior ganglionated plexus for ventricular rate control during atrial fibrillation in dogs and humans. In these experiments, we observed a decrease of the intrinsic ventricular rate during the first 4 to 5 months when HFES was intermittently shut off. We thus hypothesized that HFES might elicit trophic effects on cardiac neurons, which in turn increase baseline parasympathetic tone of the atrioventricular node. In mongrel dogs atrial fibrillation was induced by rapid atrial pacing. Endocardial HFES of the right inferior ganglionated plexus, which contains abundant fibers to the atrioventricular node, was performed for 2 years. Sham-operated nonstimulated dogs served as control. In chronic neurostimulated dogs, we found an increased neuronal cell size accompanied by an increase of choline acetyltransferase and unchanged tyrosine hydroxylase protein expression as compared with unstimulated dogs. Moreover, β-nerve growth factor (NGF) and neurotrophin (NT)-3 were upregulated in chronically neurostimulated dogs. In vitro, HFES of cultured neurons of interatrial ganglionated plexus from adult rats increased neuronal growth accompanied by upregulation of NGF, NT-3, glial-derived neurotrophic factor (GDNF), ciliary neurotrophic factor (CNTF) and brain-derived neurotrophic factor (BDNF) expression. NGF was identified as the main growth-inducing factor, whereas NT-3 did not affect HFES-induced growth. However, NT-3 could be identified as an important acetylcholine-upregulating factor. HFES of cardiac neurons in vivo and in vitro causes neuronal cellular hypertrophy, which is mediated by NGF and boosters cellular function by NT-3-mediated acetylcholine upregulation. This knowledge may contribute to develop HFES techniques to augment cardiac parasympathetic tone.

• Sympathetic neurons express and secrete MMP-2 and MT1-MMP to control nerve sprouting via pro-NGF conversion.

  Authors: Erol Saygili, Patrick Schauerte, Maimouna Pekassa, Esra Saygili, Gediminas Rackauskas, Robert H G Schwinger, Joachim Weis, Christian Weber, Nikolaus Marx, Obaida R Rana

  Cellular and molecular neurobiology. 01/2011; 31(1):17-25.

  Recently, we have shown that high frequency electrical field stimulation (HFES) of sympathetic neurons (SN) induces nerve sprouting by up-regulation of nerve growth factor (NGF) which targets theRecently, we have shown that high frequency electrical field stimulation (HFES) of sympathetic neurons (SN) induces nerve sprouting by up-regulation of nerve growth factor (NGF) which targets the tyrosine kinase A receptor (TrkA) in an autocrine/paracrine manner. There is increasing evidence that matrix metalloproteinase-2 (MMP-2) is not only involved in extracellular matrix (ECM) turnover but may also exert beneficial effects during neuronal growth. Therefore, this study aimed to investigate the regulation and function of MMP-2 and its major activator membrane type 1-matrix metalloproteinase (MT1-MMP) as well its inhibitor TIMP-1 in SN under conditions of HFES. Moreover, we analyzed molecular mechanisms of the beneficial effect of losartan, an angiotensin II type I receptor (AT-1)blocker on HFES-induced nerve sprouting. Cell cultures of SN from the superior cervical ganglia (SCG) of neonatal rats were electrically stimulated for 48 h with a frequency of 5 or 50 Hz. HFES increased MMP-2 and MT1-MMP mRNA and protein expression, whereas TIMP-1 expression remained unchanged. Under conditions of HFES, we observed a shift from pro- to active-MMP-2 indicating an increase in MMP-2 enzyme activity. Specific pharmacological MMP-2 inhibition contributed to an increase in pro-NGF amount in the cell culture supernatant and significantly reduced HFES-induced neurite outgrowth. Losartan abolished HFES-induced nerve sprouting in a significant manner by preventing HFES-induced NGF, MMP-2, and MT1-MMP up-regulation. In summary, specific MMP-2 blockade prevents sympathetic nerve sprouting (SNS) by inhibition of pro-NGF conversion while losartan abolishes HFES-induced SNS by reducing total NGF, MMP-2 and MT1-MMP expression.

• Targeted high-throughput sequencing identifies mutations in atlastin-1 as a cause of hereditary sensory neuropathy type I.

  Authors: Christian Guelly, Peng-Peng Zhu, Lea Leonardis, Lea Papić, Janez Zidar, Maria Schabhüttl, Heimo Strohmaier, Joachim Weis, Tim M Strom, Jonathan Baets, Jan Willems, Peter De Jonghe, Mary M Reilly, Eleonore Fröhlich, Martina Hatz, Slave Trajanoski, Thomas R Pieber, Andreas R Janecke, Craig Blackstone, Michaela Auer-Grumbach

  American journal of human genetics. 01/2011; 88(1):99-105.

  Hereditary sensory neuropathy type I (HSN I) is an axonal form of autosomal-dominant hereditary motor and sensory neuropathy distinguished by prominent sensory loss that leads to painless injuries.Hereditary sensory neuropathy type I (HSN I) is an axonal form of autosomal-dominant hereditary motor and sensory neuropathy distinguished by prominent sensory loss that leads to painless injuries. Unrecognized, these can result in delayed wound healing and osteomyelitis, necessitating distal amputations. To elucidate the genetic basis of an HSN I subtype in a family in which mutations in the few known HSN I genes had been excluded, we employed massive parallel exon sequencing of the 14.3 Mb disease interval on chromosome 14q. We detected a missense mutation (c.1065C>A, p.Asn355Lys) in atlastin-1 (ATL1), a gene that is known to be mutated in early-onset hereditary spastic paraplegia SPG3A and that encodes the large dynamin-related GTPase atlastin-1. The mutant protein exhibited reduced GTPase activity and prominently disrupted ER network morphology when expressed in COS7 cells, strongly supporting pathogenicity. An expanded screen in 115 additional HSN I patients identified two further dominant ATL1 mutations (c.196G>C [p.Glu66Gln] and c.976 delG [p.Val326TrpfsX8]). This study highlights an unexpected major role for atlastin-1 in the function of sensory neurons and identifies HSN I and SPG3A as allelic disorders.

• Reducing the duration of 100% oxygen ventilation in the early reperfusion period after cardiopulmonary resuscitation decreases striatal brain damage.

  Authors: Anne Brücken, Aaref Bani Kaab, Kai Kottmann, Rolf Rossaint, Kay Wilhelm Nolte, Joachim Weis, Michael Fries

  Resuscitation. 12/2010; 81(12):1698-703.

  Previous data indicate that 100% O(2) ventilation during early reperfusion after cardiac arrest (CA) and cardiopulmonary resuscitation (CPR) increases neuronal death. However, current guidelinesPrevious data indicate that 100% O(2) ventilation during early reperfusion after cardiac arrest (CA) and cardiopulmonary resuscitation (CPR) increases neuronal death. However, current guidelines encourage the use of 100% O(2) during resuscitation and for an undefined period thereafter. We retrospectively analyzed data from a porcine CA model and hypothesized that prolonged hyperoxic reperfusion would be associated with increased neurohistopathological damage and impaired neurological recovery. Fifteen male pigs underwent 8 min of CA and 5 min of CPR. After resuscitation animals were ventilated with either 100% oxygen for 60 min (hyperoxia; n=8) or 10 min (normoxia; n=7). Physiological variables were obtained at baseline and 10, 60 and 240 min after resuscitation. Daily functional performance was assessed using an established neurocognitive test in parallel to a neurological deficit score (NDS). On day 5, brains of the re-anaesthetized pigs were harvested for neurohistopathological analyses. At baseline there were no differences in hemodynamics and neurological status between groups. Post-arrest only PaO(2), as a result of the different inspired oxygen fractions, was significantly higher in the hyperoxia group. There was a numerical trend towards improved clinical recovery in both the NDS and the neurocognitive testing for animals exposed to 10 min of 100% oxygen. However, hyperoxic animals showed a significantly greater degree of necrotic neurons and perivascular inflammation in the striatum in comparison to normoxic animals. In this retrospective analysis prolonged hyperoxia after CA aggravated necrotic brain damage and perivascular inflammation in the striatum of pigs.

• Myopathy with hexagonally cross-linked crystalloid inclusions: delineation of a clinico-pathological entity.

  Authors: Kristl G Claeys, Jean-François Pellissier, Federico Garcia-Bragado, Joachim Weis, Andoni Urtizberea, Juan-Jose Poza, Ana-Maria Cobo, Gisela Stoltenburg, Dominique Figarella-Branger, Patrick J Willems [......] Jean Pouget, Monique Piraud, Guy Brochier, Norma B Romero, Michel Fardeau, Hans H Goebel, Carsten G Bönnemann, Thomas Voit, Bruno Eymard, Pascal Laforêt

  Neuromuscular disorders : NMD. 11/2010; 20(11):701-8.

  A novel myopathy characterized by hexagonally cross-linked tubular arrays has been reported in five patients. We studied the clinical and histopathological features of five additional unrelatedA novel myopathy characterized by hexagonally cross-linked tubular arrays has been reported in five patients. We studied the clinical and histopathological features of five additional unrelated patients with this myopathy. Patients experienced exercise intolerance with exercise-induced myalgia and weakness, without rhabdomyolysis. One patient additionally presented mild permanent pelvic girdle muscle weakness. Age at onset varied between 13 and 56 years. The inclusions were eosinophilic on H and E, bright red with modified Gomori's trichrome stains, present in type 2 fibers, and revealed immunoreactivity selectively for a caveolin-3-antibody. Ultrastructurally, the inclusions showed a highly organized, hexagonally cross-linked crystalloid structure. Mutations in the caveolin-3 encoding gene were excluded. Biochemical assessment of glycogenolysis in muscle was normal. Inherited or sporadic myopathy with hexagonally cross-linked tubular arrays is associated with a homogeneous clinical and histopathological phenotype. This myopathy should be included in the differential diagnosis of patients with exercise intolerance and myalgia.

• Acetylcholine as an age-dependent non-neuronal source in the heart.

  Authors: Obaida R Rana, Patrick Schauerte, Rahel Kluttig, Jörg W Schröder, Rory R Koenen, Christian Weber, Kay W Nolte, Joachim Weis, Rainer Hoffmann, Nikolaus Marx, Erol Saygili

  Autonomic neuroscience : basic & clinical. 08/2010; 156(1-2):82-9.

  In the heart, acetylcholine (ACh) slows pacemaker activity, depresses contractility and slows conduction in the atrioventricular node. Beside these cardiovascular effects, ACh has also beenIn the heart, acetylcholine (ACh) slows pacemaker activity, depresses contractility and slows conduction in the atrioventricular node. Beside these cardiovascular effects, ACh has also been associated with an anti-inflammatory and anti-apoptotic pathway. There is no evidence for ACh synthesis and excretion in other cell types than neuronal cells in the heart. Therefore, this study investigates whether cardiomyocytes are able to synthesize, transport and excrete ACh in the heart. We chose a rat model of different aged rats (neonatal, 6-8 week = young, 20-24 month = old). By real-time PCR, Western blot and immunofluorescence experiments we could demonstrate that adult, but not neonatal cardiomyocytes, express the choline acetyltransferase (ChAT). The expression level of ChAT is down-regulated in old cardiomyocytes. Furthermore, we found that young and old cardiomyocytes express the ACh transport proteins choline transporter-1 (CHT-1) and the vesicular acetylcholine transporter (VAChT). The amount of ACh excretion detected by high performance liquid chromatography (HPLC) is significantly down-regulated in old cardiomyocytes. Bromo-acetylcholine (BrACh), a specific ChAT inhibitor, significantly decreased ACh concentrations in cardiomyocyte supernatants demonstrating that ChAT is the main ACh synthesizing enzyme in cardiomyocytes. In conclusion, we could demonstrate that adult, but not neonatal, cardiomyocytes are able to synthesize, transport and excrete ACh in the rat heart. The expression level of ChAT and the ACh excretion amount are significantly down-regulated in old cardiomyocytes. This finding may provide new physiological/pathological aspects in the communication between cardiomyocytes and other cell types in the myocardium, e.g. fibrocytes, neurocytes or endothelial cells.

• Hydrogen sulfide does not increase resuscitability in a porcine model of prolonged cardiac arrest.

  Authors: Matthias Derwall, Maren Westerkamp, Céline Löwer, Jan Deike-Glindemann, Nora Katharina Schnorrenberger, Mark Coburn, Kay Wilhelm Nolte, Nadine Gaisa, Joachim Weis, Katharina Siepmann, Martin Häusler, Rolf Rossaint, Michael Fries

  Shock (Augusta, Ga.). 08/2010; 34(2):190-5.

  Treatment options to improve resuscitability and neurological prognosis after cardiac arrest (CA) are limited. Hydrogen sulfide has demonstrated remarkable improvements in outcomes in small animalTreatment options to improve resuscitability and neurological prognosis after cardiac arrest (CA) are limited. Hydrogen sulfide has demonstrated remarkable improvements in outcomes in small animal models of severe hypoxia or hemorrhage. We investigated the influence of sodium sulfide (Na2S), a liquid hydrogen sulfide donor, on resuscitability, postresuscitation hemodynamics, and neurological performance in a porcine model of prolonged CA and cardiopulmonary resuscitation. Twenty-four male pigs were instrumented with arterial and pulmonary artery catheters before 10 min of CA was induced. During resuscitation, animals were randomized to receive either high-dose (1 mg/kg; n = 8) or low-dose (0.3 mg/kg; n = 8) Na2S (IK-1001; Ikaria, Clinton, NJ) or control (saline placebo; n = 8) i.v. injection and consecutive infusion. Cardiopulmonary resuscitation was performed for 6 min before defibrillation was attempted. Hemodynamic variables were taken at baseline and 10, 30, 60, 120, and 240 min after successful resuscitation. Neurological outcome was evaluated on 4 postoperative days before brains and hearts were harvested for histopathologic analysis. No differences in hemodynamic parameters were observed at baseline. Initial resuscitability was not improved by Na2S. Animals exposed to high- and low-dose Na2S showed significantly reduced cardiac output, heart rate, and pulmonary arterial pressure compared with control animals during the early postresuscitation period. Strikingly, two of the high-dose Na2S animals died during the postresuscitation period, whereas all other animals survived. High-dose Na2S significantly decreased microglial activation in striatal areas, although this did not translate into improved neurological outcome. Although animals receiving Na2S developed higher troponin T serum levels, these differences remained insignificant. In this investigation, Na2S did not improve resuscitability but significantly compromised postresuscitation hemodynamics.

• Differential effects of myopathy-associated caveolin-3 mutants on growth factor signaling.

  Authors: Eva Brauers, Agnes Dreier, Andreas Roos, Berthold Wormland, Joachim Weis, Alexander Krüttgen

  The American journal of pathology. 07/2010; 177(1):261-70.

  Caveolin-3 is an important scaffold protein of cholesterol-rich caveolae. Mutations of caveolin-3 cause hereditary myopathies that comprise remarkably different pathologies. Growth factor signalingCaveolin-3 is an important scaffold protein of cholesterol-rich caveolae. Mutations of caveolin-3 cause hereditary myopathies that comprise remarkably different pathologies. Growth factor signaling plays an important role in muscle physiology; it is influenced by caveolins and cholesterol-rich rafts and might thus be affected by caveolin-3 dysfunction. Prompted by the observation of a marked chronic peripheral neuropathy in a patient suffering from rippling muscle disease due to the R26Q caveolin-3 mutation and because TrkA is expressed by neuronal cells and skeletal muscle fibers, we performed a detailed comparative study on the effect of pathogenic caveolin-3 mutants on the signaling and trafficking of the TrkA nerve growth factor receptor and, for comparison, of the epidermal growth factor receptor. We found that the R26Q mutant slightly and the P28L strongly reduced nerve growth factor signaling in TrkA-transfected cells. Surface biotinylation experiments revealed that the R26Q caveolin-3 mutation markedly reduced the internalization of TrkA, whereas the P28L did not. Moreover, P28L expression led to increased, whereas R26Q expression decreased, epidermal growth factor signaling. Taken together, we found differential effects of the R26Q and P28L caveolin-3 mutants on growth factor signaling. Our findings are of clinical interest because they might help explain the remarkable differences in the degree of muscle lesions caused by caveolin-3 mutations and also the co-occurrence of peripheral neuropathy in the R26Q caveolinopathy case presented.

• Resistance to antiangiogenic therapy is directed by vascular phenotype, vessel stabilization, and maturation in malignant melanoma.

  Authors: Iris Helfrich, Inka Scheffrahn, Sönke Bartling, Joachim Weis, Verena von Felbert, Mark Middleton, Masahi Kato, Süleyman Ergün, Dirk Schadendorf

  The Journal of experimental medicine. 03/2010; 207(3):491-503.

  Angiogenesis is not only dependent on endothelial cell invasion and proliferation, it also requires pericyte coverage of vascular sprouts for stabilization of vascular walls. Clinical efficacy ofAngiogenesis is not only dependent on endothelial cell invasion and proliferation, it also requires pericyte coverage of vascular sprouts for stabilization of vascular walls. Clinical efficacy of angiogenesis inhibitors targeting the vascular endothelial growth factor (VEGF) signaling pathway is still limited to date. We hypothesized that the level of vessel maturation is critically involved in the response to antiangiogenic therapies. To test this hypothesis, we evaluated the vascular network in spontaneously developing melanomas of MT/ret transgenic mice after using PTK787/ZK222584 for anti-VEGF therapy but also analyzed human melanoma metastases taken at clinical relapse in patients undergoing adjuvant treatment using bevacizumab. Both experimental settings showed that tumor vessels, which are resistant to anti-VEGF therapy, are characterized by enhanced vessel diameter and normalization of the vascular bed by coverage of mature pericytes and immunoreactivity for desmin, NG-2, platelet-derived growth factor receptor beta, and the late-stage maturity marker alpha smooth muscle actin. Our findings emphasize that the level of mural cell differentiation and stabilization of the vascular wall significantly contribute to the response toward antiangiogenic therapy in melanoma. This study may be useful in paving the way toward a more rational development of second generation antiangiogenic combination therapies and in providing, for the first time, a murine model to study this.