Publications by authors named "Lars Barthel"

6 Publications

  • Page 1 of 1

Ca-Zn-Ag Alginate Aerogels for Wound Healing Applications: Swelling Behavior in Simulated Human Body Fluids and Effect on Macrophages.

Polymers (Basel) 2020 Nov 18;12(11). Epub 2020 Nov 18.

Department Food Chemistry and Toxicology, Institute of Food Technology and Food Chemistry, TU Berlin, Straße des 17. Juni 135, 10623 Berlin, Germany.

Chronic non-healing wounds represent a substantial economic burden to healthcare systems and cause a considerable reduction in quality of life for those affected. Approximately 0.5-2% of the population in developed countries are projected to experience a chronic wound in their lifetime, necessitating further developments in the area of wound care materials. The use of aerogels for wound healing applications has increased due to their high exudate absorbency and ability to incorporate therapeutic substances, amongst them trace metals, to promote wound-healing. This study evaluates the swelling behavior of Ca-Zn-Ag-loaded alginate aerogels and their metal release upon incubation in human sweat or wound fluid substitutes. All aerogels show excellent liquid uptake from any of the formulas and high liquid holding capacities. Calcium is only marginally released into the swelling solvents, thus remaining as alginate bridging component aiding the absorption and fast transfer of liquids into the aerogel network. The zinc transfer quota is similar to those observed for common wound dressings in human and animal injury models. With respect to the immune regulatory function of zinc, cell culture studies show a high availability and anti-inflammatory activity of aerogel released Zn-species in RAW 264.7 macrophages. For silver, the balance between antibacterial effectiveness versus cytotoxicity remains a significant challenge for which the alginate aerogels need to be improved in the future. An increased knowledge of the transformations that alginate aerogels undergo in the course of the fabrication as well as during wound fluid exposure is necessary when aiming to create advanced, tissue-compatible aerogel products.
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http://dx.doi.org/10.3390/polym12112741DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7699170PMC
November 2020

Universal law for diffusive mass transport through mycelial networks.

Biotechnol Bioeng 2021 Feb 24;118(2):930-943. Epub 2020 Nov 24.

School of Life Sciences Weihenstephan, Chair of Process Systems Engineering, Technical University of Munich, Freising, Germany.

Filamentous fungal cell factories play a pivotal role in biotechnology and circular economy. Hyphal growth and macroscopic morphology are critical for product titers; however, these are difficult to control and predict. Usually pellets, which are dense networks of branched hyphae, are formed during industrial cultivations. They are nutrient- and oxygen-depleted in their core due to limited diffusive mass transport, which compromises productivity of bioprocesses. Here, we demonstrate that a generalized law for diffusive mass transport exists for filamentous fungal pellets. Diffusion computations were conducted based on three-dimensional X-ray microtomography measurements of 66 pellets originating from four industrially exploited filamentous fungi and based on 3125 Monte Carlo simulated pellets. Our data show that the diffusion hindrance factor follows a scaling law with respect to the solid hyphal fraction. This law can be harnessed to predict diffusion of nutrients, oxygen, and secreted metabolites in any filamentous pellets and will thus advance the rational design of pellet morphologies on genetic and process levels.
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http://dx.doi.org/10.1002/bit.27622DOI Listing
February 2021

Vesicle transport and growth dynamics in Aspergillus niger: Microscale modeling of secretory vesicle flow and centerline extraction from confocal fluorescent data.

Biotechnol Bioeng 2020 09 27;117(9):2875-2886. Epub 2020 Jun 27.

Chair of Measurement and Control, Technische Universität Berlin, Berlin, Germany.

In this paper, we present a mathematical model to describe filamentous fungal growth based on intracellular secretory vesicles (SVs), which transport cell wall components to the hyphal tip. Vesicular transport inside elongating hyphae is modeled as an advection-diffusion-reaction equation with a moving boundary, transformed into fixed coordinates, and discretized using a high-order weighted essentially nonoscillatory discretization scheme. The model describes the production and the consumption of SVs with kinetic functions. Simulations are subsequently compared against distributions of SVs visualized by enhanced green fluorescent protein in young Aspergillus niger hyphae after germination. Intensity profile data are obtained using an algorithm scripted in ImageJ that extracts mean intensity distributions from 3D time-lapse confocal measurement data. Simulated length growth is in good agreement with the experimental data. Our simulations further show that a decrease of effective vesicle transport velocity towards the tip can explain the observed tip accumulation of SVs.
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http://dx.doi.org/10.1002/bit.27452DOI Listing
September 2020

From three-dimensional morphology to effective diffusivity in filamentous fungal pellets.

Biotechnol Bioeng 2019 12 8;116(12):3360-3371. Epub 2019 Oct 8.

Chair of Process Systems Engineering, Technical University of Munich, Freising, Germany.

Filamentous fungi are exploited as cell factories in biotechnology for the production of proteins, organic acids, and natural products. Hereby, fungal macromorphologies adopted during submerged cultivations in bioreactors strongly impact the productivity. In particular, fungal pellets are known to limit the diffusivity of oxygen, substrates, and products. To investigate the spatial distribution of substances inside fungal pellets, the diffusive mass transport must be locally resolved. In this study, we present a new approach to obtain the effective diffusivity in a fungal pellet based on its three-dimensional morphology. Freeze-dried Aspergillus niger pellets were studied by X-ray microcomputed tomography, and the results were reconstructed to obtain three-dimensional images. After processing these images, representative cubes of the pellets were subjected to diffusion computations. The effective diffusion factor and the tortuosity of each cube were calculated using the software GeoDict. Afterwards, the effective diffusion factor was correlated with the amount of hyphal material inside the cubes (hyphal fraction). The obtained correlation between the effective diffusion factor and hyphal fraction shows a large deviation from the correlations reported in the literature so far, giving new and more accurate insights. This knowledge can be used for morphological optimization of filamentous pellets to increase the yield of biotechnological processes.
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http://dx.doi.org/10.1002/bit.27166DOI Listing
December 2019

An X-ray microtomography-based method for detailed analysis of the three-dimensional morphology of fungal pellets.

Biotechnol Bioeng 2019 06 8;116(6):1355-1365. Epub 2019 Mar 8.

Technical University of Munich, School of Life Sciences Weihenstephan, Chair of Process Systems Engineering, Freising, Germany.

Filamentous fungi are widely used in the production of biotechnological compounds. Since their morphology is strongly linked to productivity, it is a key parameter in industrial biotechnology. However, identifying the morphological properties of filamentous fungi is challenging. Owing to a lack of appropriate methods, the detailed three-dimensional morphology of filamentous pellets remains unexplored. In the present study, we used state-of-the-art X-ray microtomography (µCT) to develop a new method for detailed characterization of fungal pellets. µCT measurements were performed using freeze-dried pellets obtained from submerged cultivations. Three-dimensional images were generated and analyzed to locate and quantify hyphal material, tips, and branches. As a result, morphological properties including hyphal length, tip number, branch number, hyphal growth unit, porosity, and hyphal average diameter were ascertained. To validate the potential of the new method, two fungal pellets were studied-one from Aspergillus niger and the other from Penicillium chrysogenum. We show here that µCT analysis is a promising tool to study the three-dimensional structure of pellet-forming filamentous microorganisms in utmost detail. The knowledge gained can be used to understand and thus optimize pellet structures by means of appropriate process or genetic control in biotechnological applications.
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http://dx.doi.org/10.1002/bit.26956DOI Listing
June 2019

Construction of an improved Aspergillus niger platform for enhanced glucoamylase secretion.

Microb Cell Fact 2018 Jun 16;17(1):95. Epub 2018 Jun 16.

Department Applied and Molecular Microbiology, Institute of Biotechnology, Technische Universität Berlin, Gustav-Meyer-Allee 25, 13355, Berlin, Germany.

Background: The lifestyle of filamentous fungi depends on the secretion of hydrolytic enzymes into the surrounding medium, which degrade polymeric substances into monomers that are then taken up to sustain metabolism. This feature has been exploited in biotechnology to establish platform strains with high secretory capacity including Aspergillus niger. The accepted paradigm is that proteins become mainly secreted at the tips of fungal hyphae. However, it is still a matter of debate if the amount of growing hyphal tips in filamentous fungi correlates with an increase in secretion, with previous studies showing either a positive or no correlation.

Results: Here, we followed a systematic approach to study protein secretion in A. niger. First, we put the glaA gene encoding for glucoamylase (GlaA), the most abundant secreted protein of A. niger, under control of the tunable Tet-on system. Regulation of glaA gene expression by omitting or adding the inducer doxycycline to cultivation media allowed us to study the effect of glaA under- or overexpression in the same isolate. By inducing glaA expression in a fluorescently tagged v-SNARE reporter strain expressing GFP-SncA, we could demonstrate that the amount of post-Golgi carriers indeed depends on and correlates with glaA gene expression. By deleting the racA gene, encoding the Rho-GTPase RacA in this isolate, we generated a strain which is identical to the parental strain with respect to biomass formation but produces about 20% more hyphal tips. This hyperbranching phenotype caused a more compact macromorphology in shake flask cultivations. When ensuring continuous high-level expression of glaA by repeated addition of doxycycline, this hyperbranching strain secreted up to four times more GlaA into the culture medium compared to its parental strain.

Conclusion: The data obtained in this study strongly indicate that A. niger responds to forced transcription of secretory enzymes with increased formation of post-Golgi carriers to efficiently accommodate the incoming cargo load. This physiological adaptation can be rationally exploited to generate hypersecretion platforms based on a hyperbranching phenotype. We propose that a racA deletion background serves as an excellent chassis for such hypersecretion strains.
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http://dx.doi.org/10.1186/s12934-018-0941-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6004097PMC
June 2018