Biology subjects

Hammer, M.

Publications and source records attributed to Hammer, M..

15 recordsLinked to original sources

bioRxiv2026

Molecular context of pathogenic variants is associated with phenotype and treatment response in SCN8A-related disorders

Objectives: Genotype-phenotype studies in rare epilepsies typically relate a pathogenic DNA sequence change to clinical outcome, without considering the broader molecular context of a given variant. Here we ask whether clinical heterogeneity in SCN8A-related disorders (SCN8A-RD) is patterned along molecular dimensions that go beyond the specific genetic alteration, to include position in the linear channel topology, proximity to post-translational modification (PTM) and structural sites, and predicted effects on mRNA splicing. Using the International SCN8A Registry, we map these features against seizure, developmental, and treatment-response phenotypes to identify the regions and molecular feature classes with which clinical variability is associated. Methods: We identify "hot-spots" across the coding sequence for seizure types and severity of developmental disability (DD). We then test for enrichment of clinical features near sites related to PTM and protein structure, identify variants that may alter mRNA splicing, and evaluate the interaction between these features and clinical subgroups. Results: We expand on previous work identifying regions of coding DNA sequence that are pathogenic or benign "hot-spots". We show regional enrichment of numerous seizure types and severities of DD. Enrichment analysis of PTM and protein structural sites shows distinct enrichment profiles for developmental, seizure, and medication-response features. Finally, we identify 14 variants that are likely to alter RNA splicing. Significance: We provide a higher-resolution map of pathogenic variation across SCN8A and show that seizure types, developmental severity, and medication response are regionally organized along the coding sequence. Proximity to structural and glycosylation sites distinguishes phenotypic subgroups is associated with differential drug response, including a glycosylation-gabapentin relationship, and 14 missense variants are flagged as likely splice-altering, nominating targets for splice-directed therapy and providing candidates for experimental validation of dual pathogenic mechanisms.

molecular biology↗

bioRxiv2026

Segmentation and classification of retinal pigment granules in fluorescence lifetime imaging microscopy (FLIM) data

Alterations of fluorescence properties in retinal pigment epithelium (RPE) cells caused by diseases such as age-related macular degeneration (AMD) highlight the need for detailed analysis of the fluorescent RPE granules at the individual level. Precise segmentation and classification of these granules remain challenging due to their limited visual separability. In this study, we present Classi4RPE, a computational algorithm designed to accurately segment RPE granules and classify them into three categories -- lipofuscin (L), melanolipofuscin (ML), and melanin (M) -- based on fluorescence lifetime imaging data, which provide distinctive contrast. The method is implemented in a custom Python framework and employs seeded watershed segmentation to isolate individual granules. Lipofuscin granules are identified as hyperfluorescent structures with longer lifetimes, while granules with shorter lifetimes are further analyzed based on their spatial lifetime distribution from the center to edge, enabling discrimination of ML from other melanin-rich granules. Our approach achieves high performance, with mean sensitivities of 0.99 for L granules and 0.90 for ML granules, and corresponding specificities of 0.93 and 0.98, respectively, compared to manually annotated ground truth. These results demonstrate the potential of Classi4RPE to surpass human visual limitations and provide a robust tool for quantitative RPE analysis.

bioinformatics↗

bioRxiv2026

Enzyme-functionalized microparticles to open the vitreoretinal interface

The transport of therapeutics and gene carriers to their site of action is often hindered by biological barriers, such as cell layers and basement membranes. Among these, the inner limiting membrane (ILM) represents a major barrier within the eye, separating the vitreous body from the retina. The ILM must be crossed, if for instance gene carriers are to reach retinal target cells following intravitreal administration. However, the ILM is a densely cross-linked basement membrane barrier, allowing only the smallest nanoparticles to pass. Here, we demonstrate that active micro-colloids decorated with enzymes can locally open the ILM and thereby facilitate the diffusion of passive carriers into retinal tissue. We utilize an ex vivo porcine eye model to determine the membrane permeability threshold using fluorescent nanoprobes. We further show that collagenase-decorated silica microparticles can facilitate the transport of nanoparticles, while exhibiting excellent biocompatibility with no adverse morphological or functional retinal effects over a six-week in vivo evaluation in a porcine model. Overall, our findings introduce a biocompatible and minimally invasive strategy to facilitate the targeted nanoparticle transport across biological barriers, which we demonstrate for retinal delivery enabled by active colloids.

bioengineering↗

bioRxiv2024

Using muscle-tendon load limits to assess unphysiological musculoskeletal model deformation and Hill-type muscle parameter choice

Musculoskeletal simulations are a useful tool for improving our understanding of the human body. However, the physiological validity of predicted kinematics and forces is highly dependent upon the correct calibration of muscle parameters and the structural integrity of a models internal skeletal structure. In this study, we show how ill-tuned muscle parameters and unphysiological deformations of a models skeletal structure can be detected by using muscle elements as sensors with which modelling and parameterization inconsistencies can be identified through muscle and tendon strain injury assessment. To illustrate our approach, two modelling issues were recreated. First, a model repositioning simulation using the THUMS AM50 occupant model version 5.03 was performed to show how internal model deformations can occur during a change of model posture. Second, the muscle material parameters of the OpenSim gait2354 model were varied to illustrate how unphysiological muscle forces can arise if material parameters are inadequately calibrated. The simulations were assessed for muscle and tendon strain injuries using previously published injury criteria and a newly developed method to determine tendon strain injury threshold values. Muscle strain injuries in the left and right musculus pronator teres were detected during the model repositioning. This straining was caused by an unphysiologically large gap (12.92 mm) that had formed in the elbow joint. Similarly, muscle and tendon strain injuries were detected in the modified right-hand musculus gastrocnemius medialis of the gait2354 model where an unphysiological reduction of the tendon slack length introduced large pre-strain of the muscle-tendon-unit. The results of this work show that the proposed method can quantify the internal distortion behaviour of musculoskeletal human body models and the validity of Hill-type muscle parameter choice via strain injury assessment. Furthermore, we highlight possible actions to avoid the presented issues and inconsistencies in literature data concerning the material characteristics of human tendons.

biophysics↗

bioRxiv2022

Natural hybridisation reduces vulnerability to climate change

Understanding how species can respond to climate change is a major global challenge. Species unable to track their niche via range shifts are largely reliant on genetic variation to adapt and persist. Genomic vulnerability predictions are used to identify populations that lack the necessary variation, particularly at climate relevant genes. However, hybridization as a source of novel adaptive variation is typically ignored in genomic vulnerability studies. We estimated environmental niche models and genomic vulnerability for closely related species of rainbowfish (Melanotaenia spp.) across an elevational gradient in the Australian wet tropics. Hybrid populations between a widespread generalist and narrow range endemics exhibited reduced vulnerability to projected climates compared to pure narrow endemics. Overlaps between introgressed and adaptive genomic regions were consistent with a signal of adaptive introgression. Our findings highlight the often-underappreciated conservation value of hybrid populations and indicate that adaptive introgression may contribute to evolutionary rescue of species with narrow environmental ranges.

evolutionary biology↗

bioRxiv2022

Phosphorylation Determines Whether Neuroligin-3 is at Excitatory or Inhibitory Synapses in Different Regions of the Brain

Neuroligin-3 is a postsynaptic adhesion molecule involved in development, function, and pathologies of synapses in the brain. It is a genetic cause of autism and a potent component of the tumor microenvironment in gliomas. There are four Neuroligins that operate at distinct synapse types, selectively interacting with presynaptic adhesion and postsynaptic scaffold proteins. We investigated the subcellular localization and scaffold specificities of synaptic Neuroligin-3 and demonstrate an unexpected pattern of localization to excitatory synapses in cortical areas, and inhibitory synapses in subcortical areas. Using phosphoproteomics, we identify Neuroligin-3-specific serine phosphorylation in cortex and hippocampus that obstructs a key binding site for inhibitory synapse scaffolds. Using in utero CRISPR/Cas9 knockout and replacement with phosphomimetic mutants, we demonstrate that phosphorylation at this site determines excitatory versus inhibitory synapse localization of Neuroligin-3 in vivo. Our data reveal a mechanism that differentially regulates the balance of Neuroligin-3 between excitatory and inhibitory synapses, adding to our emerging understanding of their role in the development of brain connectivity and associated pathologies.

neuroscience↗

bioRxiv2021

Variation in intraspecific demography drives localised concordance but species-wide discordance in responses to Plio-Pleistocene climatic change

Understanding how species biology may facilitate resilience to climate change remains a critical factor in detecting and protecting species at risk of extinction. Many studies have focused on the role of particular ecological traits in driving species responses, but less so on demographic history and levels of standing genetic variation. We used environmental and genomic datasets to reconstruct the phylogeographic histories of two ecologically similar and largely co-distributed freshwater fishes to assess the degree of concordance in their responses to Plio-Pleistocene climatic changes. Although several co-occurring populations demonstrated concordant demographic histories, idiosyncratic population size changes were found at the range edges of the more spatially restricted species. Discordant responses between species were associated with low standing genetic variation in peripheral populations. This might have hindered adaptive potential, as documented in recent population declines and extinctions of the two species. Our results highlight both the role of spatial scale in the degree of concordance in species responses to climate change, and the importance of standing genetic variation in facilitating range shifts. Even when ecological traits are similar between species, long-term genetic diversity and historical population demography may lead to discordant responses to ongoing and future climate change.

evolutionary biology↗

bioRxiv2021

Micro-Meta App: an interactive software tool to facilitate the collection of microscopy metadata based on community-driven specifications

For the information content of microscopy images to be appropriately interpreted, reproduced, and meet FAIR (Findable Accessible Interoperable and Reusable) principles, they should be accompanied by detailed descriptions of microscope hardware, image acquisition settings, image pixel and dimensional structure, and instrument performance. Nonetheless, the thorough documentation of imaging experiments is significantly impaired by the lack of community-sanctioned easy-to-use software tools to facilitate the extraction and collection of relevant microscopy metadata. Here we present Micro-Meta App, an intuitive open-source software designed to tackle these issues that was developed in the context of nascent global bioimaging community organizations, including BioImaging North America (BINA) and QUAlity Assessment and REProducibility in Light Microscopy (QUAREP-LiMi), whose goal is to improve reproducibility, data quality and sharing value for imaging experiments. The App provides a user-friendly interface for building comprehensive descriptions of the conditions utilized to produce individual microscopy datasets as specified by the recently proposed 4DN-BINA-OME tiered-system of Microscopy Metadata model. To achieve this goal the App provides a visual guide for a microscope-user to: 1) interactively build diagrammatic representations of hardware configurations of given microscopes that can be easily reused and shared with colleagues needing to document similar instruments. 2) Automatically extracts relevant metadata from image files and facilitates the collection of missing image acquisition settings and calibration metrics associated with a given experiment. 3) Output all collected Microscopy Metadata to interoperable files that can be used for documenting imaging experiments and shared with the community. In addition to significantly lowering the burden of quality assurance, the visual nature of Micro-Meta App makes it particularly suited for training users that have limited knowledge of the intricacies of light microscopy experiments. To ensure wide-adoption by microscope-users with different needs Micro-Meta App closely interoperates with MethodsJ2 and OMERO.mde, two complementary tools described in parallel manuscripts.

bioinformatics↗

bioRxiv2021

Towards community-driven metadata standards for light microscopy: tiered specifications extending the OME model

1 -Digital light microscopy provides powerful tools for quantitatively probing the real-time dynamics of subcellular structures. While the power of modern microscopy techniques is undeniable, rigorous record-keeping and quality control are required to ensure that imaging data may be properly interpreted (quality), reproduced (reproducibility), and used to extract reliable information and scientific knowledge which can be shared for further analysis (value). Keeping notes on microscopy experiments and quality control procedures ought to be straightforward, as the microscope is a machine whose components are defined and the performance measurable. Nevertheless, to this date, no universally adopted community-driven specifications exist that delineate the required information about the microscope hardware and acquisition settings (i.e., microscopy "data provenance" metadata) and the minimally accepted calibration metrics (i.e., microscopy quality control metadata) that should be automatically recorded by both commercial microscope manufacturers and customized microscope developers. In the absence of agreed guidelines, it is inherently difficult for scientists to create comprehensive records of imaging experiments and ensure the quality of resulting image data or for manufacturers to incorporate standardized reporting and performance metrics. To add to the confusion, microscopy experiments vary greatly in aim and complexity, ranging from purely descriptive work to complex, quantitative and even sub-resolution studies that require more detailed reporting and quality control measures. To solve this problem, the 4D Nucleome Initiative (4DN) (1, 2) Imaging Standards Working Group (IWG), working in conjunction with the BioImaging North America (BINA) Quality Control and Data Management Working Group (QC-DM-WG) (3), here propose light Microscopy Metadata specifications that scale with experimental intent and with the complexity of the instrumentation and analytical requirements. They consist of a revision of the Core of the Open Microscopy Environment (OME) Data Model, which forms the basis for the widely adopted Bio-Formats library (4-6), accompanied by a suite of three extensions, each with three tiers, allowing the classification of imaging experiments into levels of increasing imaging and analytical complexity (7, 8). Hence these specifications not only provide an OME-based comprehensive set of metadata elements that should be recorded, but they also specify which subset of the full list should be recorded for a given experimental tier. In order to evaluate the extent of community interest, an extensive outreach effort was conducted to present the proposed metadata specifications to members of several core-facilities and international bioimaging initiatives including the European Light Microscopy Initiative (ELMI), Global BioImaging (GBI), and European Molecular Biology Laboratory (EMBL) - European Bioinformatics Institute (EBI). Consequently, close ties were established between our endeavour and the undertakings of the recently established QUAlity Assessment and REProducibility for Instruments and Images in Light Microscopy global community initiative (9). As a result this flexible 4DN-BINA-OME (NBO namespace) framework (7, 8) represents a turning point towards achieving community-driven Microscopy Metadata standards that will increase data fidelity, improve repeatability and reproducibility, ease future analysis and facilitate the verifiable comparison of different datasets, experimental setups, and assays, and it demonstrates the method for future extensions. Such universally accepted microscopy standards would serve a similar purpose as the Encode guidelines successfully adopted by the genomic community (10, 11). The intention of this proposal is therefore to encourage participation, critiques and contributions from the entire imaging community and all stakeholders, including research and imaging scientists, facility personnel, instrument manufacturers, software developers, standards organizations, scientific publishers, and funders.

bioinformatics↗

bioRxiv2020

Causal relations between cortical network oscillations and breathing frequency

Nasal breathing generates a rhythmic signal which entrains cortical network oscillations in widespread brain regions on a cycle-to-cycle time scale. It is unknown, however, how respiration and neuronal network activity interact on a larger time scale: are breathing frequency and typical neuronal oscillation patterns correlated? Is there any directionality or causal relationship? To address these questions, we recorded field potentials from the posterior parietal cortex of mice together with respiration during REM sleep. In this state, the parietal cortex exhibits prominent theta and gamma oscillations while behavioral activity is minimal, reducing confounding signals. We found that the instantaneous breathing rate strongly correlates with the instantaneous frequency and amplitude of both theta and gamma oscillations. Granger causality analysis revealed specific directionalities for different rhythms: changes in theta activity precede and cause changes in breathing rate, suggesting control of breathing frequency by the functional state of the brain. On the other hand, the instantaneous breathing rate Granger-causes changes in gamma oscillations, suggesting that gamma is influenced by a peripheral reafference signal. These findings show that breathing causally relates to different patterns of rhythmic brain activity, revealing new and complex interactions between elementary physiological functions and neuronal information processing. Significance StatementThe study of the interactions between respiration and brain activity has been focused on phase-entrainment relations, in which cortical networks oscillate phase-locked to breathing cycles. Here we discovered new and much broader interactions which link respiration rate (frequency) to different patterns of oscillatory brain activity. Specifically, we show that the instantaneous breathing rate strongly correlates with the instantaneous frequency and amplitude of theta and gamma oscillations, two major network patterns associated with cognitive functions. Interestingly, causality analyses reveal that changes in breathing rate follow theta, suggesting a central drive, while in contrast, gamma activity follows changes in breathing rate, suggesting the role of a reafferent signal. Our results reveal new mechanisms by which nasal breathing patterns may influence brain functions.

neuroscience↗

bioRxiv2020

Theta-gamma coupling depends on breathing rate

Temporal coupling between theta and gamma oscillations is a hallmark activity pattern of several cortical networks and becomes especially prominent during REM sleep. In a parallel approach, nasal breathing has been recently shown to generate phase-entrained network oscillations which also modulate gamma. Both slow rhythms (theta and respiration-entrained oscillations) have been suggested to aid large-scale integration but they differ in frequency, display low coherence, and modulate different gamma sub-bands. Respiration and theta are therefore believed to be largely independent. In the present work, however, we report an unexpected but robust relation between theta-gamma coupling and respiration in mice. Interestingly, this relation takes place not through the phase of individual respiration cycles, but through respiration rate: the strength of theta-gamma coupling exhibits an inverted V-shaped dependence on breathing rate, leading to maximal coupling at breathing frequencies of 4-6 Hz. Noteworthy, when subdividing sleep epochs into phasic and tonic REM patterns, we find that breathing differentially relates to theta-gamma coupling in each state, providing new evidence for their physiological distinctiveness. Altogether, our results reveal that breathing correlates with brain activity not only through phase-entrainment but also through rate-dependent relations with theta-gamma coupling. Thus, the link between respiration and other patterns of cortical network activity is more complex than previously assumed.

neuroscience↗

bioRxiv2020

Fluorescence Lifetimes and Spectra of RPE and sub-RPE Deposits in Histology of Normal and AMD Eyes

PurposeTo investigate autofluorescence lifetimes as well as spectral characteristics of drusen and retinal pigment epithelium (RPE) in age-related macular degeneration (AMD). MethodFluorescence lifetimes and spectra of five eyes with AMD and nine control eyes were analyzed in cryosections by means of two-photon excited fluorescence at 960 nm. Spectra were detected at 490 - 647nm. Lifetime was measured using time-correlated single photon counting in two spectral channels: 500-550nm and 550-700nm. The fluorescence decays over time were approximated by a series of three exponential functions and the amplitude-weighted mean fluorescence lifetime {tau}m was determined. Results196 sub-RPE deposits were identified (AMD n=76, healthy n=120) and 230 RPE sites recorded. The peak emission wavelength of drusen was significantly green-shifted compared to RPE (peak at 570nm vs. 610nm), but not different between patients and controls. Drusen showed considerably longer {tau}m than RPE: (ch1: 581 {+/-} 163 ps vs. 177 {+/-} 25 ps, ch2: 541 {+/-} 125 ps vs. 285 {+/-} 31 ps, p < 0.001). Drusen found in AMD eyes had longer lifetimes than drusen of controls (ch1: 650 {+/-} 167 ps vs. 537 {+/-} 145 ps, ch2: 600 {+/-} 125 ps vs. 504 {+/-} 111 ps, p < 0.001). In addition, drusen in AMD eyes showed a more homogenous fluorescence distribution and more drusen were larger than 63{micro}m than in control eyes. ConclusionsEx vivo fluorescence imaging of drusen in cross-sections enables the separation of their autofluorescence from that of over- or underlying structures. Our analysis showed considerable variability of drusen lifetimes but not spectra. This indicates that drusen consist of a variety of different fluorophores or expose the same fluorophores to different microenvironments. Changes in drusen lifetimes could be related to AMD progression.

cell biology↗

bioRxiv2020

The roles of aridification and sea level changes in the diversification and persistence of freshwater fish lineages

While the influence of Pleistocene climatic changes on divergence and speciation has been well-documented across the globe, complex spatial interactions between hydrology and eustatics over longer timeframes may also determine species evolutionary trajectories. Within the Australian continent, glacial cycles were not associated with changes in ice cover and instead largely resulted in fluctuations from moist to arid conditions across the landscape. Here, we investigate the role of hydrological and coastal topographic changes brought about by Plio-Pleistocene climatic changes on the biogeographic history of a small Australian freshwater fish, the southern pygmy perch Nannoperca australis. Using 7,958 ddRAD-seq (double digest restriction-site associated DNA) loci and 45,104 filtered SNPs, we combined phylogenetic, coalescent and species distribution analyses to investigate the relative roles of aridification, sea level and tectonics and their associated biogeographic changes across southeast Australia. Sea-level changes since the Pliocene and reduction or disappearance of large waterbodies throughout the Pleistocene were determining factors in strong divergence across the clade, including the initial formation and maintenance of a cryptic species, N. flindersi. Isolated climatic refugia and fragmentation due to lack of connected waterways maintained the identity and divergence of inter- and intraspecific lineages. Our historical findings suggest that predicted increases in aridification and sea level due to anthropogenic climate change might result in markedly different demographic impacts, both spatially and across different landscape types.

evolutionary biology↗

bioRxiv2019

A genome-wide genetic pleiotropy approach identified shared loci between multiple system atrophy and inflammatory bowel disease

We aimed to identify shared genetic background between multiple system atrophy (MSA) and autoimmune diseases by using the conjFDR approach. Our study showed significant genetic overlap between MSA and inflammatory bowel disease and identified DENND1B, C7, and RSP04 loci, which are linked to significant changes in methylation or expression levels of adjacent genes. We obtained evidence of enriched heritability involving immune/digestive categories. Finally, an MSA mouse model showed dysregulation of the C7 gene in the degenerating midbrain compared to wildtype mice. The results identify novel molecular mechanisms and implicate immune and gut dysfunction in MSA pathophysiology.

genetics↗

bioRxiv2018

Photon count estimation in single-molecule localization microscopy

Recently, Franke, Sauer and van de Linde1 introduced a way to estimate the axial position of single-molecules (TRABI). To this end, they compared the detected photon count from a temporal radial-aperture-based intensity estimation to the estimated count from Gaussian point-spread function (PSF) fitting to the data. Empirically they found this photometric ratio to be around 0.7-0.8 close to focus and decreasing away from it. Here, we explain this reported but unexplained discrepancy and furthermore show that the photometric ratio as indicator for axial position is susceptible even to typical optical aberrations.

molecular biology↗