Browsing by Author "Brock, Judith"

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Open Access
Acclimation to elevated CO2 affects the C/N balance by reducing de novo N‐assimilation
(2021) Krämer, Konrad; Kepp, Gabi; Brock, Judith; Stutz, Simon; Heyer, Arnd G.
Plants exposed to elevated atmospheric CO2 concentrations show an increased photosynthetic activity. However, after prolonged exposure, the activity declines. This acclimation to elevated CO2 is accompanied by a rise in the carbon‐to‐nitrogen ratio of the biomass. Hence, increased sugar accumulation and sequential downregulation of photosynthetic genes, as well as nitrogen depletion and reduced protein content, have been hypothesized as the cause of low photosynthetic performance. However, the reason for reduced nitrogen content in plants at high CO2 is unclear. Here, we show that reduced photorespiration at increased CO2‐to‐O2 ratio leads to reduced de novo assimilation of nitrate, thus shifting the C/N balance. Metabolic modeling of acclimated and non‐acclimated plants revealed the photorespiratory pathway to function as a sink for already assimilated nitrogen during the light period, providing carbon skeletons for de novo assimilation. At high CO2, low photorespiratory activity resulted in diminished nitrogen assimilation and eventually resulted in reduced carbon assimilation. For the hpr1‐1 mutant, defective in reduction of hydroxy‐pyruvate, metabolic simulations show that turnover of photorespiratory metabolites is expanded into the night. Comparison of simulations for hpr1‐1 with those for the wild type allowed investigating the effect of a perturbed photorespiration on N‐assimilation.
Open Access
Interaction of nitrate assimilation and photorespiration at elevated CO2
(2022) Krämer, Konrad; Brock, Judith; Heyer, Arnd G.
It has been shown repeatedly that exposure to elevated atmospheric CO2 causes an increased C/N ratio of plant biomass that could result from either increased carbon or - in relation to C acquisition - reduced nitrogen assimilation. Possible reasons for diminished nitrogen assimilation are controversial, but an impact of reduced photorespiration at elevated CO2 has frequently been implied. Using a mutant defective in peroxisomal hydroxy-pyruvate reductase (hpr1-1) that is hampered in photorespiratory turnover, we show that indeed, photorespiration stimulates the glutamine-synthetase 2 (GS) / glutamine-oxoglutarate-aminotransferase (GOGAT) cycle, which channels ammonia into amino acid synthesis. However, mathematical flux simulations demonstrated that nitrate assimilation was not reduced at elevated CO2, pointing to a dilution of nitrogen containing compounds by assimilated carbon at elevated CO2. The massive growth reduction in the hpr1-1 mutant does not appear to result from nitrogen starvation. Model simulations yield evidence for a loss of cellular energy that is consumed in supporting high flux through the GS/GOGAT cycle that results from inefficient removal of photorespiratory intermediates. This causes a futile cycling of glycolate and hydroxy-pyruvate. In addition to that, accumulation of serine and glycine as well as carboxylates in the mutant creates a metabolic imbalance that could contribute to growth reduction.
Open Access
Native mechano-regulative matrix properties stabilize alternans dynamics and reduce spiral wave stabilization in cardiac tissue
(2024) Erhardt, Julia; Ludwig, Sebastian; Brock, Judith; Hörning, Marcel
The stability of wave conduction in the heart is strongly related to the proper interplay between the electrophysiological activation and mechanical contraction of myocytes and extracellular matrix (ECM) properties. In this study, we statistically compare bioengineered cardiac tissues cultured on soft hydrogels ( E≃12kPa) and rigid glass substrates by focusing on the critical threshold of alternans, network-physiological tissue properties, and the formation of stable spiral waves that manifest after wave breakups. For the classification of wave dynamics, we use an improved signal oversampling technique and introduce simple probability maps to identify and visualize spatially concordant and discordant alternans as V- and X-shaped probability distributions. We found that cardiac tissues cultured on ECM-mimicking soft hydrogels show a lower variability of the calcium transient durations among cells in the tissue. This lowers the likelihood of forming stable spiral waves because of the larger dynamical range that tissues can be stably entrained with to form alternans and larger spatial spiral tip movement that increases the chance of self-termination on the tissue boundary. Conclusively, we show that a dysfunction in the excitation-contraction coupling dynamics facilitates life-threatening arrhythmic states such as spiral waves and, thus, highlights the importance of the network-physiological interplay between contractile myocytes and the ECM.
Open Access
Optimization of H9c2 differentiation leads to calcium-active and striated cardiac cells without addition of retinoic acid
(2024) Brock, Judith; Hörning, Marcel
As a reliable alternative to animal testing in cardiovascular research, it is crucial to improve differentiation of immortalized cell lines. In this study, we focused on optimizing the differentiation efficiency of the H9c2 cell line into cardiomyocytes using a high-throughput, automated image processing approach. While previous studies used protocols involving retinoic acid to enhance cardiac differentiation, we applied a simplified medium composition that results in higher differentiation rates. Along that line, we differentiated H9c2 cells into cardiomyocytes, which not only showed sarcomere-characteristic striation but also periodic intracellular calcium signaling for the first time. As a second step, we examined the potential application of polyacrylamide hydrogels ( E=12kPa) with defined fibronectin coating densities. The optimum fibronectin density of 2.6 μg/cm 2 found for single cells was investigated to further improve the differentiation efficiency. However, the differentiation and proliferation dynamics dominate the adhesion forces between the cells and the hydrogel, and thus, result in premature clustering and detachment. In conclusion, we identified an optimized differentiation protocol and provided a basis for the further investigation necessary to potentially use hydrogels as natural cell environment, aiming to raise the differentiation efficiency even more.
Open Access
Optimization of mechanosensitive cross-talk between matrix stiffness and protein density : independent matrix properties regulate spreading dynamics of myocytes
(2022) Brock, Judith; Erhardt, Julia; Eisler, Stephan A.; Hörning, Marcel
Cells actively sense differences in topology, matrix elasticity and protein composition of the extracellular microenvironment and adapt their function and morphology. In this study, we focus on the cross-talk between matrix stiffness and protein coating density that regulates morphology and proliferation dynamics of single myocytes. For this, C2C12 myocytes were monitored on L-DOPA functionalized hydrogels of 22 different elasticity and fibronectin density compositions. Static images were recorded and statistically analyzed to determine morphological differences and to identify the optimized extracellular matrix (ECM). Using that information, selected ECMs were used to study the dynamics before and after cell proliferation by statistical comparison of distinct cell states. We observed a fibronectin-density-independent increase of the projected cell area until 12 kPa. Additionally, changes in fibronectin density led to an area that was optimum at about 2.6 μg/cm2, which was confirmed by independent F-actin analysis, revealing a maximum actin-filament-to-cell-area ratio of 7.5%. Proliferation evaluation showed an opposite correlation between cell spreading duration and speed to matrix elasticity and protein density, which did not affect cell-cycle duration. In summary, we identified an optimized ECM composition and found that independent matrix properties regulate distinct cell characteristics.