Biology subjects

Voss, H. L.

Publications and source records attributed to Voss, H. L..

3 recordsLinked to original sources

bioRxiv2023

Assay for characterizing adsorption-properties of surfaces (APS) sample preparation prior to quantitative omics

Analytes during their journey from their natural sources to their identification and quantification are prone to adsorption to surfaces before they enter an analytical instrument, causing false quantities. This problem is especially severe in diverse omics. Here, thousands of analytes with a broad range of chemical properties and thus different affinities to surfaces are quantified within a single analytical run. For quantifying adsorption effects caused by surfaces of sample handling tools, an assay was developed, applying LC-MS/MS-based differential bottom-up proteomics and as probe a reference mixture of thousands of tryptic peptides, covering a broad range of chemical properties. The assay was tested by investigating the adsorption properties of several vials composed of polypropylene, including low-protein-binding polypropylene vials, borosilicate glass vials and low-retention glass vials. In total 3531 different peptides were identified and quantified across all samples and therefore used as probes. A significant number of hydrophobic peptides adsorbed on polypropylene vials. In contrast, only very few peptides adsorbed to low-protein-binding polypropylene vials. The highest number of peptides adsorbed to glass vials, driven by electrostatic as well as hydrophobic interactions. Calculation of the impact of the adsorption of peptides on differential quantitative proteomics showed significant false results. In summary, the new assay is suitable to characterize adsorption properties of surfaces getting into contact with analytes during sample preparation, thereby giving the opportunity to find parameters for minimizing false quantities. Insert Table of Contents artwork here O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=106 SRC="FIGDIR/small/551632v3_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@bafb12org.highwire.dtl.DTLVardef@1b98219org.highwire.dtl.DTLVardef@c3b45org.highwire.dtl.DTLVardef@1074180_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2023

Cystatin C loaded in brain-derived extracellular vesicles rescues synapses after ischemic insult in vitro and in vivo

Synaptic loss is an early event in the undersupplied but not yet irreversibly injured penumbra area after an ischemic stroke. Promoting synaptic preservation in this area would likely improve functional neurological recovery. In the present study, we aimed to detect proteins involved in endogenous protection mechanisms of synapses in the penumbra after stroke and to analyse the potential beneficial effect of these candidates for a prospective stroke treatment. For this, we performed Liquid Chromatography coupled to Mass Spectrometry (LC-MS)-based proteomics of synaptosomes isolated from the ipsilateral hemispheres of mice subjected to experimental stroke at different time points (24 h, 4 and 7 days) and compared them to sham-operated mice. Proteomic analyses indicated that among the differentially expressed proteins between the two groups, cystatin C (CysC) was significantly increased at 24 h and 4 days following stroke, before returning to steady-state levels at 7 days, thus indicating a potential transient and intrinsic rescue mechanism attempt of neurons. When CysC was applied to primary neuronal cultures subjected to an in vitro model of ischemic damage, this treatment significantly improved the preservation of synaptic structures. Notably, similar effects were observed when CysC was loaded into brain-derived extracellular vesicles (BDEVs). Finally, when CysC contained in BDEVs was administered intracerebroventricularly to stroked mice, it significantly increased the expression of synaptic markers such as SNAP25, Homer-1, and NCAM in the penumbra area compared to the group supplied with empty BDEVs. Thus, we show that CysC-loaded BDEVs promote synaptic protection after ischemic damage in vitro and in vivo, opening the possibility of a therapeutic use in stroke patients.

neuroscience↗

bioRxiv2023

Multiomic profiling of medulloblastoma reveals subtype-specific targetable alterations at the proteome and N-glycan level

Medulloblastomas (MBs) are malignant pediatric brain tumors that are molecularly and clinically very heterogenous. To unravel phenotypically relevant MB subtypes, we compiled a harmonized proteome dataset of 167 MBs and integrated findings with DNA methylation and N-glycome data. Six proteome MB subtypes emerged, that could be assigned to two main molecular programs: transcription/translation (pSHHt, pWNT and pGroup3-Myc), and synapses/immunological processes (pSHHs, pGroup3 and pGroup4). Multiomic analysis revealed different conservation levels of proteome features across MB subtypes at the DNA-methylation level. Aggressive pGroup3-Myc MBs and favorable pWNT MBs were most similar in cluster hierarchies concerning overall proteome patterns but showed different protein abundances of the vincristine resistance associated multiprotein complex TriC/CCT and of N-glycan turnover associated factors. The N-glycome reflected proteome subtypes and complex-bisecting N-glycans characterized pGroup3-Myc tumors. Our results shed light on new targetable alterations in MB and set a foundation for potential immunotherapies targeting glycan structures. SignificanceWhereas the application of omics technologies has significantly improved MB tumor classification and treatment stratification, it is still of debate, which features predict best clinical outcome. Moreover, treatment options - especially for high-risk groups - are still unsatisfactory. In contrast to nucleic acids, the proteome and their N-glycans may reflect the phenotype of a tumor in a more direct way and thus hold the potential to discover clinically relevant phenotypes and potentially targetable pathways. We show that these analyses are feasible on formalin fixed and paraffine embedded tissue. Compiling a comprehensive MB dataset, we detected new biomarkers and characteristics for high- and low-risk MB subtypes that were not reflected by other omic data modalities before. Specifically, we identified subtype specific abundance differences in proteins of the vincristine resistance associated multiprotein complex TriC/CCT and in proteins involved in N-glycan turnover. Changes in the N-glycans are considered as potential hallmarks of cancer and we show that N-glycan profiles can distinguish MB subtypes. These tumor-specific N-glycan structures hold a strong potential as new biomarkers, as well as immunotherapy targets. Highlights- Integration of in-house proteome data on formalin fixated paraffine embedded medulloblastoma (MB) and publicly available datasets enables large scale proteome analysis of MB - Six proteome MB subtypes can be assigned to two main molecular programs: replication/ translation versus synapse/immune system - Identification and validation of IHC compatible protein-biomarkers for high and low risk MB subtypes, such as TNC and PALMD. - Subtype specific correlation of the DNA methylome and the proteome reveals different conserved molecular characteristics across MB subtypes. - pGroup3-Myc subtype MBs are associated with high-risk features including high abundances of vincristine resistance associated TriC/CCT member proteins - Proteome MB subtypes show differential N-glycosylation patterns, revealing complex-bisecting glycans as potentially immunotargetable hallmarks of the high risk pGroup3-Myc subtype.

cancer biology↗