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Hearing rehabilitation in SERAC1 related MEGD(H)EL syndrome – implications from a multi-center retrospective cohort study
Get PDFObjective
3-methylglutaconic aciduria (MEG), dystonia-deafness (D), (hepatopathy (H)), encephalopathy (E), and Leigh-like-syndrome (L) (MEGD(H)EL) syndrome is a rare, severely disabling progressive mitochondrial disease associated with biallelic pathogenic variants in SERAC1. Knowledge about hearing loss (HL) and hearing rehabilitation is scarce but highly sought after for best possible care in the absence of causative treatment.
Methods
Retrospective cross-sectional study.
Results
This study analyzed the audiometric data of 36 MEGD(H)EL patients (14 unpublished). Bilateral HL was diagnosed in 31 individuals (86 %). Detailed audiometric data, available for 23 of 31 patients, did not allow for general statements on site and degree of HL. HL was mostly congenital (n = 14/31), pre-lingual in six and post-lingual in nine cases (median age 2 years, n = 15/31; age unknown in n = 2).
In four of the five patients without HL, the severity of the other clinical-neurological symptoms was milder and less progressive, and their onset was significantly later than in the patients with HL. Five of 36 patients acquired spoken language, these were 4 of the 5 individuals without and one with HL. Twenty-two individuals received hearing rehabilitation with conventional hearing aids, followed by cochlear implant (CI) surgery in six. One of these six individuals acquired spoken language, which lessened in clarity as disease progressed.
Conclusions
Congenital HL represents a ubiquitous symptom in severe types of MEGD(H)EL syndrome, being absent in late onset milder forms. Regularly, severely affected MEGD(H)EL patients do not achieve spoken language, even with CI. Hence, hearing rehabilitation with CIs needs to be discussed very critically
Data archive of Synergistic olfactory nerve input and cholinergic neuromodulation can activate ERK in rat olfactory bulb vasopressin cells
No full textSocial discrimination in rats relies on vasopressin cells (VPCs) intrinsic to the olfactory bulb (OB). We had observed that VPCs responded to electrical stimulation of the olfactory nerve in acute OB slices with inhibitory postsynaptic potentials, and that the neuromodulator acetylcholine could revert these responses to excitation, resulting in action potentials (AP). Moreover, in behaving rats that were exposed to conspecifics, more VPCs were immunopositive to the neural activity marker pERK (pERK+ VPC) than in control rats. However, it is unclear whether these two observations, the increased ERK activation in vivo, and the generation of APs in the presence of ACh in vitro, can be actually mapped onto each other. Here we investigated ERK activation in acute OB slices from transgenic VP-eGFP rats upon either chemical stimulation or tetanic olfactory nerve stimulation. Both KCl and NMDA stimulation resulted in substantial pERK induction across bulbar layers and caused VPC spiking in whole-cell recordings, but only NMDA slightly increased pERK+ VPC percentage. Tetanic olfactory nerve stimulation yielded localized, column-like ERK activation of neurons across bulbar layers. The presence of ACh during tetanic stimulation substantially and specifically increased percentages of pERK+ VPCs within columns, indicating that columnar pERK+ VPCs were preferentially activated by coincident cholinergic neuromodulation and synaptic input.
Our results validate pERK induction as a tool to monitor synaptic VPC excitation in the OB, and imply that depolarization of VPCs alone is insufficient to activate ERK. We propose that synaptically evoked APs are a prerequisite for pERK induction in VPCs
Expression of Versican in the Retina and Its Implication in Retinal Disease
No full textChondroitin sulfate proteoglycans are characterized by their sulfated glycosaminoglycan chains covalently attached to the core protein. Versican, encoded by the CSPG2 gene, is a chondroitin sulfate proteoglycan highly expressed in the human and mouse retina. Due to alternative splicing, it exists in four different isoforms: V0, V1, V2 and V3. Interactions with a variety of proteins lead to its main functions, such as influencing cellular adhesion, migration, proliferation and regulating inflammatory processes. Versican expression is high during retinal development and decreases in the mature tissue, remaining mostly in the retinal pigment epithelium, Bruch’s membrane and choroid blood vessel walls. Mutations in the CSPG2 gene resulting in an imbalance of Versican splice variants cause erosive vitreoretinopathy and Wagner disease. Its regulatory role in a variety of functions, especially those influencing extracellular matrix composition and inflammation, suggests a crucial role in the pathomechanisms of retinal degenerative disorders and should be the purpose of future research
Novel water-soluble and highly efficient dual type I/II next generation inhibitors of FMS-like tyrosine kinase 3 (FLT3)
No full textMutations of the FMS-like tyrosine kinase (FLT3) occur in acute myeloid leukemia (AML) and are associated with very poor prognosis. Available FLT3 inhibitors are potent but either show a lack of selectivity regarding other tyrosine kinases or only transient efficacy due to emerging resistance under therapy. Water-soluble derivatives of the tyrosine kinase inhibitor Marbotinib are highly selective dual-type I/II inhibitors of FLT3.
The bisarylmethanone-based compound 29 and its carbamate derivative 42 show excellent results in various biological tests. They inhibit FLT3-ITD (internal tandem duplication) as well as therapy-associated FLT3-TKD point mutations. Additionally, good water solubility and consequently biological availability was achieved by attaching amine functions to appropriate scaffold positions, suggested by modeling of inhibitor binding at inactive and active FLT3 states. Subsequent formation of different salts led to very promising results in in vivo studies and improvements compared to midostaurin (1b), Quizartinib (7), Marbotinib 10 and its carbamate 11c
rTMS for the treatment of psychiatric disorders: a review about training courses and materials and the presentation of the training materials of the German Society for Brain Stimulation in Psychiatry
Get PDFThere is an increasing interest in the treatment ofmental disorders such as depression
using repetitive transcranial magnetic stimulation (rTMS). Treatment guidelines for the
application of rTMS are available, but they focus primarily on scientific evidence for
efficacy and tolerability. Other aspects that are relevant for the training of practitioners,
such as implementation in the national health system as well as organizational, formal,
and practical aspects, are not covered in detail in these guidelines.Here, we present an
overview of German rTMS courses and present training materials for the hands-on
workshop of the German Society for Brain Stimulation in Psychiatry [Deutsche
Gesellschaft für Hirnstimulation in der Psychiatrie (DGHP) e.V.] in German and
English as an example. Publishing national standard operating procedures is
important in order to harmonize rTMS practice all over the world
Design and Characterization of Vaccine Candidates for Therapeutic Vaccination Against Papillomaviruses
No full textDespite the availability of highly efficacious prophylactic vaccines against human papillomaviruses (HPV), the predominantly sexually transmitted viruses are still highly prevalent worldwide and represent the most significant part of virus-induced tumor diseases. Infection with HPV is the leading cause of cervical dysplasia and cancer, but moreover, it can also cause tumors in the other parts of the anogenital tract and in the head and neck area, especially in the oropharynx. Thus, the global burden of morbidity and mortality is immense. The prophylactic vaccines only protect from virus infection but cannot extirpate virus-infected cells, which renders them ineffective as a treatment for HPV-induced dysplasia and cancer. Hence, a novel therapy against HPV-induced malignancies is of the highest urgency.
Natural regression of HPV-induced dysplasia is connected to rigid T cell responses against the regulatory early proteins of the virus. This renders these proteins a possible candidate for therapeutic vaccination. Therapeutic vaccine candidates are mainly focused on the oncoproteins E6 and E7, thus varying in clinical efficacy. Hence, this thesis aims to expand the restricted antigen usage in order to generate novel type-specific therapeutic vaccine candidates.
First, with the lack of a small animal model for sexually-transmitted HPV, Macaca fascicularis papillomavirus type 3 (MfPV3) was chosen as a model virus due to its high homology and phylogenic relationship to HPV16 and its ability to induce HPV-like dysplasia in persistently infected Macaca fascicularis. A set of antigens of MfPV3 was designed based on the regulatory early proteins E1, E2, E6, and E7 and the N-terminal fusion of the MHC-II-associated invariant chain (Ii). Biochemical and molecular biological in vitro as well as immunological in vivo characterization of DNA vaccine vector- and adenoviral vector-delivered antigens proved that the artificially fused polyprotein Ii-E1E2E6E7 is not inferior compared to the combination of single antigens, thus renders this antigen the lead therapeutic vaccine candidate for pre-clinical efficacy trials in persistently-infected macaques.
Second, HPV16 represents the most critical clinical relevant HPV type. Hence, HPV16 antigens were designed based on the blueprints of the MfPV3 antigens. Biochemical, molecular, and cell biological characterization proved the absence of transformation capacity of the antigens. Moreover, in-depth immunological characterization of adenoviral-delivered antigens revealed potent anti-tumor efficacy in a translational HPV genome-positive tumor model in inbred mice, especially in synergy with cisplatin. Therefore, the HPV16 Ii-E1E2E6E7 therapeutic vaccine candidate might be a promising contender in future clinical investigations to combat HPV16-induced malignancies.
Finally, recombinant poxviral vectors based on the Modified Vaccinia virus Ankara (MVA) comprising MfPV3 Ii-E1E2E6E7 were generated in a duck-derived continuous cell line, which was modified to knock down transgene expression. Thorough characterization of the cell line side-by-side with the non-modified parental cell line exhibited better replication, resulting in higher virus titers, and enhanced genetic stability across serial passaging of both recombinant MVA (rMVA) in the absence of transgene expression. Furthermore, it was shown that the genetic stability depends on the integration site within the rMVA. particularly the mode of action how instability is manifested differs between the integration sites. Thus, the knock-down of transgene expression while rMVA generation and production might be a promising strategy for manufacturing recombinant viruses with demanding transgenes.
In summary, the generation of novel therapeutic vaccine constructs against HPV-associated disease with broader antigen capacity due to the inclusion of E1 and E2 and also of vectors suitable for antigen delivery is shown in this work. The data presented here argues for the potential beneficial use of the Ii-E1E2E6E7 antigen in the therapy against HPV-induced malignancies. A future pre-clinical efficacy trial with an adenovirus-prime and MVA-boost scheme in persistently MfPV3-infected macaques may guide the way for the clinical development of the promising HPV16 Ii-E1E2E6E7 therapeutic vaccine candidate
Identification of molecular characteristics of CNS-seeking cancer cells
No full textTumor cells that are detected in the cerebrospinal fluid or the membranes surrounding the brain and spinal cord (meninges), or both, are referred to as leptomeningeal metastases (LM). They are characterized as a serious complication of advanced cancer, can lead to drastic neurological symptoms and occur in 5-10% of patients with solid tumors. Breast cancer, lung cancer and melanoma are the most common associated diseases, but cases with an unknown primary tumor (cancer of unknown primary (CUP) syndrome) can also be affected. LM are rare but aggressive in nature and have limited treatment options and an unfavorable central nervous system (CNS) location, which is associated with an unfavorable prognosis and difficult therapeutic decision-making. The diagnosis of LM relies mainly on CSF cytology and magnetic resonance imaging (MRI) evaluation, both of which are the current gold standard in diagnosis, and, where available, information on the primary tumor. As the latter is not always accessible and CSF cytology and magnetic resonance therapy (MRI) have a low sensitivity, the results of which are of a purely quantitative nature, a molecular approach for CSF fluid biopsy was developed in this study, which should make it easier to predict treatment for affected patients in the future.
In a monocentric feasibility study, a qualitative comparison between CSF cytology, MRI data and molecular liquid biopsy results of adult patients with suspected LM and different primary tumors was performed. Disseminated cancer cells (DCCs), cell-free tumor DNA (ctDNA) and formalin-fixed paraffin-embedded (FFPE) tissue were molecularly characterized. The results were used to answer the following three questions:
(i) Does molecular fluid biopsy of CSF provide comparable results to the current clinical gold standard, currently CSF cytology and MRI, for patients with suspected LM?
ii) Is the isolation of single cells from CSF a valuable option to provide relevant molecular data and how reliable are these results compared to the analysis of circulating cell-free tumor DNA from CSF in terms of quality, significance and applicability?
iii) Does the heterogeneous landscape of tumor and metastatic tissue indicate the sedimentation of specific subclones colonizing the CNS?
In the first, technical part of this work, a molecular diagnostic workflow was established and optimized with the aim of detecting, enriching and isolating DCCs in CSF. Depending on the underlying primary tumor and its associated expression markers, it was evaluated whether cells could be detected either with the FDA (U.S. Food and Drug Administration) approved CellSearch® technology, based on the EpCAM (epithelial cell adhesion molecule) expression pattern and cytokeratin staining of carcinoma cells, or by density gradient centrifugation followed by specific fluorescence staining for melanoma cells. These results served to validate the CellSearch® method for CSF in this population in comparison with simultaneous cytological analyses and associated MRI data. The great advantage of this detection method is the ability to identify and isolate individual cells after cell enrichment based on their specific staining. This possibility was used to build up a single cell collective obtained from the CSF for subsequent molecular analysis.
Two different isolation methods were evaluated and the differences in DNA quality depending on the origin of the cell and the technical reprocessing were assessed: The manual method of fluorescence microscopy and automated isolation using DEPArrayTM technology. The categorization of DNA quality served as an additional tool to differentiate between low and high quality samples, with the latter being used for further molecular analyses. Whole genome sequencing was used to determine allele frequencies, which made it possible to determine the origin of the sample and to investigate subclonal diversity within different isolated CSF single cells. In addition to the tumor cells, circulating tumor DNA was also isolated from the CSF. Based on these samples, the workflow for the extraction of cell-free DNA was optimized, a new amplification strategy was tested and validated. The molecular data from these samples were then used to determine whether the enriched DNA was tumor DNA or not and whether the molecular data matched the data from the isolated DCCs.
The advantages and disadvantages of the two biomarkers in the CNS were compared in terms of quality, informative value, applicability and reliability. The analysis of the tumor tissue of the patients included in the study made it possible to investigate intratumoral heterogeneity. For this purpose, tumor and stromal cell populations were isolated from formalin-fixed paraffin-embedded (FFPE) tissue to validate the technical feasibility of the DEPArrayTM technology and to optimize the extraction protocol for different tumor types and materials. After the isolation process, the cell populations were analyzed at the molecular level and the data isolated from non-CNS or CNS tissue was compared with data from CSF isolated DNA.
This work has shown that the CSF single cell analysis approach provides comparable results to conventional CSF cytology and, together with MRI, can therefore represent a more sensitive alternative to the current clinical gold standard. A major advantage is the possibility of isolating high-quality cells and subsequently characterizing them at the molecular level. This means that CSF could also be a valuable source for the cultivation and functional analysis of vital cells, which can be used to detect clinically relevant, tumor-specific information that could contribute to the decision-making process for suitable therapy in the future. A fast and cost-effective method of analyzing tumor DNA is the isolation of ctDNA from cerebrospinal fluid. However, the reliability and informative value of this biomarker is significantly lower compared to single cell analysis within this collective. An analysis of both CSF biomarkers can be a practicable solution to quickly obtain initial information from ctDNA and, based on this, to follow up with a more in-depth molecular analysis of the individual cells.
The fact that the genetic profile of cells that sediment into the CNS is clearly different from cells isolated from the visceral area of the affected person highlights the importance of molecular analysis of CSF DNA. This enables the detection of possible CNS-specific markers for individual therapeutic approaches that would remain undetected without the analysis of CSF
The induced deletion of fibronectin disrupts the formation of renal lymphatic vessels
No full textFibronectin (FN) is a major constituent of the renal fibrillar extracellular matrix (ECM). To explore FN's specific functions in the interstitium of the kidney, mutant mice with induced FN deficiency were generated applying a tamoxifen-dependent Cre/LoxP system. Newborn CAGG-Cre-ERTM/Fnfl/fl mice were treated with tamoxifen eye drops. By the third postnatal day (P3) treatment resulted in a substantial (ca. 90 %) loss of FN and its mRNA in kidney and liver as seen by quantitative RT-PCR and Western blot analysis. The loss caused a dramatic reduction of immunoreactivity for fibrillar FN in the renal interstitium consistent with loss of both cellular (kidney-derived) and plasma (liver-derived) FN. FN loss resulted in the formation of cyst-like structures at the cortex-medulla transitional area. The localization correlated with the area of highest renal FN expression and deposition in normal mice as seen by in situ hybridization and immunohistochemistry. In mutant mice the structures rapidly expanded replacing after one week most of the normal kidney parenchyma. The cyst-like structures were not lined by tubular epithelium but by interstitial cells and were not connected with the lumen of tubuli. In addition, they contained non-fibrillar extracellular matrix components that were visualized by hyaluronan synthase and alcian-blue staining. Overall, the findings indicated that the cyst-like structures resulted from a rapid expanding interstitial edema. The edema did not result from changes in renal blood capillaries that showed an essentially normal network in both medulla and cortex. In addition, no changes were observed in the formation of capillary fenestrae. Instead, a significant reduction in the number of LYVE-1 positive migratory cells and putative lymphangioblasts was observed within the medulla of FN-deficient mice. The observation was consistent with failure of lymphovasculogenesis resulting in attenuated formation of lymphatic capillaries that normally occurs in the first postnatal week. In support of this assumption was the finding that induced deficiency of FN in one-week-old mice with an established network of lymphatic capillaries did not cause obvious structural changes in the kidney.
In summary the findings strongly indicate that the major function of renal FN is to provide an interstitial scaffolding that allows the migration of lymphangioblasts to form the normal network of renal lymphatic capillaries. The formation of renal lymphatic vessels occurs by FN-dependent lymphangiovasculogenesis
Essays on Round-Number Effects and Moral Balancing
No full textThis dissertation investigates behavioral phenomena in economic decision-making across three projects.
The first project examines the influence of round numbers in real estate price negotiations. Based on a large dataset of residential and commercial transactions in Germany, clustering at round-number prices in residential real estate is identified—mirroring findings from the U.S. market, but with notable cultural differences in what constitutes a "round" number. When extending the analysis to commercial real estate, professionals are found to rely less on round-number pricing than non-professionals. Hedonic regressions and machine learning models reveal that residential transactions at round prices occur at values exceeding the estimated hedonic prices of the respective properties.
The second project investigates the mechanisms underlying round-number clustering in negotiations. Two explanations are considered: cognitive bias and focal-point coordination. Analysis of observational eBay negotiation data shows that successful negotiations often conclude at round prices, which are also associated with faster agreements. To distinguish between the two mechanisms, an incentivized online experiment is conducted. The findings indicate that both bias and focal-point effects contribute to the clustering, highlighting the dual function of round numbers in negotiation outcomes.
The third project explores moral balancing in pro-environmental behavior. It is examined whether even minor environmentally beneficial actions are used to justify subsequent environmentally harmful behavior. In a two-stage economic experiment, participants who acquire a “moral license” are found to offset less carbon than those who do not. This effect is moderated by environmental concern: participants with stronger environmental values are less likely to engage in such compensatory behavior