Biology subjects

Schneider, W.

Publications and source records attributed to Schneider, W..

5 recordsLinked to original sources

bioRxiv2026

Genetic accessibility, not superior fitness, determines which anti-HBs antibody escape variants are selected in patients

Background and Aims: Genetic diversity within the hepatitis B virus (HBV) surface antigen (HBsAg) drives the emergence of immune escape variants, but studying this diversity in vivo has been limited by the difficulty of establishing infection from patient isolates. We aimed to perform unbiased in vivo screening of variants in the HBs antigenic loop for antibody escape and fitness, using synthetic DNA launch of HBV in human hepatocyte chimeric mice. Approach and Results: Direct intrahepatic injection (IHI) of barcoded, recombinant cccDNA established infection from up to 32,000 independent launch events of a genotype A strain and successfully launched clinical consensus sequences across the major genotypes. Using IHI, we performed deep mutational scanning of 4,608 codon variants across the HBs antigenic loop. Under control antibody treatment, diverse variants expanded, revealing differences in variant fitness in the absence of adaptive immune pressure. Anti-HBs monoclonal antibody treatment suppressed wild-type viremia while permitting breakthrough of resistant variants from the library. Sequencing identified strong enrichment for substitutions at G145 (G145Q, G145H, G145R). Notably, G145Q, rather than the clinically dominant G145R, showed the greatest fitness advantage, indicating that G145R's clinical prevalence does not reflect superior intrinsic fitness. Conclusions: Unbiased in vivo screening reveals that clinically observed HBV escape mutations reflect which substitutions are genetically accessible by single nucleotide change and compatible with the overlapping polymerase reading frame, rather than which confer optimal fitness. This approach establishes a platform for high-resolution mapping of viral variant fitness under antiviral selection pressure.

microbiology↗

bioRxiv2024

Time-division multiplexing (TDM) sequence removes bias in T2 estimation and relaxation-diffusion measurements

PurposeTo compare the performance of multi-echo (ME) and time-division multiplexing (TDM) sequences for accelerated relaxation-diffusion MRI (rdMRI) acquisition and to examine their reliability in estimating accurate rdMRI microstructure measures. MethodThe ME, TDM, and the reference single-echo (SE) sequences with six echo times (TE) were implemented using Pulseq with single-band (SB-) and multi-band 2 (MB2-) acceleration factors. On a diffusion phantom, the image intensities of the three sequences were compared, and the differences were quantified using the normalized root mean squared error (NRMSE). For the in-vivo brain scan, besides the image intensity comparison and T2-estimates, different methods were used to assess sequence-related effects on microstructure estimation, including the relaxation diffusion imaging moment (REDIM) and the maximum-entropy relaxation diffusion distribution (MaxEnt-RDD). ResultsTDM performance was similar to the gold standard SE acquisition, whereas ME showed greater biases (3-4x larger NRMSEs for phantom, 2x for in-vivo). T2 values obtained from TDM closely matched SE, whereas ME sequences underestimated the T2 relaxation time. TDM provided similar diffusion and relaxation parameters as SE using REDIM, whereas SB-ME exhibited a 60% larger bias in the map and on average 3.5x larger bias in the covariance between relaxation-diffusion coefficients. ConclusionOur analysis demonstrates that TDM provides a more accurate estimation of relaxation-diffusion measurements while accelerating the acquisitions by a factor of 2 to 3.

neuroscience↗

bioRxiv2021

Transcriptomic responses of the human kidney to acute injury at single cell resolution

BackgroundAcute kidney injury (AKI) occurs frequently in critically ill patients and is associated with adverse outcomes. Cellular mechanisms underlying AKI and kidney cell responses to injury remain incompletely understood. MethodsWe performed single-nuclei transcriptomics, bulk transcriptomics, molecular imaging studies, and conventional histology on kidney tissues from 8 individuals with severe AKI (stage 2 or 3 according to Kidney Disease: Improving Global Outcomes (KDIGO) criteria). Specimens were obtained within 1-2 hours after individuals had succumbed to critical illness associated with respiratory infections, with 4 of 8 individuals diagnosed with COVID-19. Control kidney tissues were obtained post-mortem or after nephrectomy from individuals without AKI. ResultsHigh-depth single cell-resolved gene expression data of human kidneys affected by AKI revealed enrichment of novel injury-associated cell states within the major cell types of the tubular epithelium, in particular in proximal tubules, thick ascending limbs and distal convoluted tubules. Four distinct, hierarchically interconnected injured cell states were distinguishable and characterized by transcriptome patterns associated with oxidative stress, hypoxia, interferon response, and epithelial-to-mesenchymal transition, respectively. Transcriptome differences between individuals with AKI were driven primarily by the cell type-specific abundance of these four injury subtypes rather than by private molecular responses. AKI-associated changes in gene expression between individuals with and without COVID-19 were similar. ConclusionThe study provides an extensive resource of the cell type-specific transcriptomic responses associated with critical illness-associated AKI in humans, highlighting recurrent disease-associated signatures and inter-individual heterogeneity. Personalized molecular disease assessment in human AKI may foster the development of tailored therapies.

molecular biology↗

bioRxiv2021

The desmin mutation R349P increases contractility and fragility of stem cell-generated muscle micro-tissues

Desminopathies comprise hereditary myopathies and cardiomyopathies caused by mutations in the intermediate filament protein desmin that lead to severe and often lethal degeneration of striated muscle tissue. Animal and single cell studies hinted that this degeneration process is associated with massive ultrastructural defects correlating with increased susceptibility of the muscle to acute mechanical stress. The underlying mechanism of mechanical susceptibility, and how muscle degeneration develops over time, however, has remained elusive. Here, we investigated the effect of a desmin mutation on the formation, differentiation, and contractile function of in vitro-engineered three-dimensional micro-tissues grown from muscle stem cells (satellite cells) isolated from heterozygous R349P desmin knock-in mice. Micro-tissues grown from desmin-mutated cells exhibited spontaneous unsynchronized contractions, higher contractile forces in response to electrical stimulation, and faster force recovery compared to tissues grown from wild-type cells. Within one week of culture, the majority of R349P desmin-mutated tissues disintegrated, whereas wild-type tissues remained intact over at least three weeks. Moreover, under tetanic stimulation lasting less than five seconds, desmin-mutated tissues partially or completely ruptured, whereas wild-type tissues did not display signs of damage. Our results demonstrate that the progressive degeneration of desmin-mutated micro-tissues is closely linked to extracellular matrix fiber breakage associated with increased contractile forces and unevenly distributed tensile stress. This suggests that the age-related degeneration of skeletal and cardiac muscle in patients suffering from desminopathies may be similarly exacerbated by mechanical damage from high-intensity muscle contractions. We conclude that micro-tissues may provide a valuable tool for studying the organization of myocytes and the pathogenic mechanisms of myopathies.

biophysics↗

bioRxiv2019

High-force magnetic tweezers with hysteresis-free force feedback

Magnetic tweezers based on solenoids with iron alloy cores are widely used to apply large forces (~100 nN) onto micron-sized (~5 m) superparamagnetic particles for mechanical manipulation or microrheological measurements at the cellular and molecular level. The precision of magnetic tweezers, however, is limited by the magnetic hysteresis of the core material, especially for time-varying force protocols. Here, we eliminate magnetic hysteresis by a feedback control of the magnetic induction, which we measure with a Hall sensor mounted to the distal end of the solenoid core. We find that the generated force depends on the induction according to a power-law relationship, and on the bead-tip distance according to a stretched exponential relationship. Together, both relationships allow for an accurate force calibration and precise force feedback with only 3 calibration parameters. We apply our method to measure the force-dependence of the viscoelastic and plastic properties of fibroblasts using a protocol with stepwise increasing and decreasing forces. We find that soft cells show an increasing stiffness but decreasing plasticity at higher forces, indicating a pronounced stress stiffening of the cytoskeleton. By contrast, stiff cells show no stress stiffening but an increasing plasticity at higher forces. These findings indicate profound differences between soft and stiff cells regarding their protection mechanisms against external mechanical stress. In summary, our method increases the precision, simplifies the handling and extends the applicability of magnetic tweezers. SIGNIFICANCEMagnetic tweezers are widely used, versatile tools to investigate the mechanical behavior of cells or to measure the strength of receptor-ligand bonds. A limitation of existing high-force magnetic tweezer setups, however, is caused by the magnetic hysteresis of the tweezer core material. This magnetic hysteresis requires that the tweezer core must be de-magnetized (de-Gaussed) prior to each measurement, and that flexible force protocols with decreasing forces are not possible. We describe how these limitations can be overcome with a force feedback though direct magnetic field measurement. We demonstrate the applicability of our setup by investigating the visco-elastic and plastic deformations of fibroblasts to forces of different amplitudes.

biophysics↗