Biology subjects

Hampl, A.

Publications and source records attributed to Hampl, A..

7 recordsLinked to original sources

bioRxiv2025

Phosphatidylcholine Metabolism Controls Alveolar Progenitor Renewal and Pulmonary Fibrosis

Idiopathic pulmonary fibrosis (IPF) is a progressive, fatal lung disease marked by alveolar type 2 (AT2) stem cell dysfunction and excessive matrix deposition, with no effective treatments. Recent advances have recognized that AT2 cells act as stem cells, in addition to their role in the production of pulmonary surfactants in the distal alveolar space. We and others have reported a failure of AT2 regeneration and a loss of AT2 cells in IPF. We recently further reported that there is a defect in lipid metabolism in IPF AT2 cells and we discovered a selective loss of lysophosphatidylcholine acyltransferase 1 (LPCAT1) in AT2 cells from IPF, as well as in AT2 cells from bleomycin-injured mice. Pharmacological and genetic experiments confirm that LPCAT1 is required for AT2 cell renewal in 3D organoid assays. AT2 cell-specific Lpcat1 deletion resulted in reduced AT2 renewal, spontaneous lung fibrosis, and heightened susceptibility to bleomycin-induced fibrosis in mice in vivo. Expression-based high-content drug screening with an LPCAT1 knock-in cell line identified several drug families that upregulated LPCAT1 expression. We further confirmed that anti-malarial artesunate and PLA2 inhibitor ONO-RS-082 increased LPCAT1 mRNA expression, promoted AT2 renewal, and attenuated bleomycin-induced lung fibrosis in mice in vivo. Our findings establish LPCAT1 as a critical regulator of AT2 renewal and lipid metabolism in IPF, suggesting that reactivation of LPCAT1 could offer a novel therapeutic strategy for restoring alveolar progenitor function and mitigating lung fibrosis.

cell biology↗

bioRxiv2023

Brain map of aging-induced alterations in membrane ganglioside pattern

Recent efforts to develop comprehensive metabolome and proteome brain atlases for animal models have yielded significant progress. However, the ganglioside (GSs) profile of these models remains largely unexplored. As essential components of the brain, GSs play a crucial role in neuronal function. To address this gap in knowledge, we conducted an in-depth analysis of the young and adult rat brain, as well as its major brain regions, using ultra-high performance liquid chromatography and tandem mass spectrometry in positive and negative ion modes. We also analyzed GSs in cerebrospinal fluid (CSF) and serum from matched samples. Our findings indicate a shift in the ratio of a-series to b-series GSs with age, along with region-specific changes accompanied by aging. This could improve our understanding of brain aging and neurodegenerative diseases. Our study complements existing brain atlases of lipidome and protein expression and highlights the importance of further investigating the mechanisms underlying these GSs changes and the potential therapeutic implications of our findings.

neuroscience↗

bioRxiv2023

FGF2-induced Redox Signaling: A Mechanism Regulating Pyruvate Dehydrogenase Driven Histone Acetylation and NANOG Upregulation

Precise control of pluripotency is a requirement for the safe and effective use of hPSCs in research and therapies. Here we report that pyruvate dehydrogenase upregulates histone H3 pan acetylation and levels of pluripotency marker NANOG in 5% O2. Pyruvate dehydrogenase (PDH) is an essential metabolic switch and a bottleneck for the glycolytic production of acetyl-CoA. Silencing of gene expression showed that PDH is regulated by the activity of its phosphatase PDP1. We show that PDP1 is sensitive to reactive oxygen species-mediated inactivation, leading to the downregulation of H3 pan acetylation and NANOG levels. Furthermore, we show that FGF2, a cytokine commonly used to maintain pluripotency activates pyruvate dehydrogenase through MEK1/2-ERK1/2 signaling pathway-mediated downregulation of ROS in 5% O2, thus promoting histone acetylation. Our results show the importance of pyruvate dehydrogenase in regulating energy metabolism and its connection to pluripotency. Furthermore, our data highlight the role of reactive oxygen species and redox homeostasis in pluripotency maintenance and differentiation. Highlights- PDP1-induced activation of PDH leads to increased histone H3 pan acetylation and NANOG levels in hPSCs - Reactive oxygen species (ROS) inactivate PDP1 and decrease histone H3 pan acetylation and NANOG levels in hPSCs - MEK1/2-ERK1/2 signaling-mediated downregulation of ROS in 5% O2 activates PDH in hPSCs Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=129 SRC="FIGDIR/small/524871v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@133f8dcorg.highwire.dtl.DTLVardef@1176d94org.highwire.dtl.DTLVardef@11b153corg.highwire.dtl.DTLVardef@10f2e4d_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2022

LuminoCell: a versatile and affordable luminometer platform for monitoring in-cell luciferase-based reporters

Luciferase reporter assays represent a simple and sensitive experimental system in cell and molecular biology to study multiple biological processes. However, the application of these assays is often limited by the costs of conventional luminometer instruments and the versatility of their use in different experimental conditions. Therefore, we aimed to develop a small, affordable luminometer allowing continuous measurement of luciferase activity, designed for inclusion into various kinds of tissue culture incubators. Here we introduce LuminoCell - an open-source platform for the construction of an affordable, sensitive, and portable luminometer capable of realtime monitoring in-cell luciferase activity. The LuminoCell costs $40, requires less than 1 hour to assemble, and it is capable of performing real-time sensitive detection of both magnitude and duration of the activity of major signalling pathways in cell cultures, including receptor tyrosine kinases (EGF, FGF), WNT/{beta}-catenin, and NF-{kappa}B. Additionally, we show that the LuminoCell is suitable to be used in cytotoxicity assays as well as for monitoring periodic circadian gene expression.

molecular biology↗

bioRxiv2021

TACSTD2 upregulation is an early reaction to lung infection

TACSTD2 encodes a transmembrane glycoprotein Trop2 commonly overexpressed in carcinomas. While the Trop2 protein was discovered already in 1981 and first antibody-drug conjugate targeting Trop2 were recently approved cancer therapy, the physiological role of Trop2 is still not fully understood. In this article, we show that TACSTD2/Trop2 expression is evolutionarily conserved in lungs of various vertebrates. By analysis of publicly available transcriptomic data we demonstrate that TACSTD2 level consistently increases in lungs infected with miscellaneous pathogens. Single cell and subpopulation based transcriptomic data revealed that the major source of TACSTD2 transcript are lung epithelial cells and their progenitors and that TACSTD2 is induced directly in lung epithelial cells following infection. This increase may represent a mechanism to maintain/restore epithelial barrier function and contribute to regeneration process in infected/damaged lungs.

cell biology↗

bioRxiv2021

High-resolution 3D reconstruction of human oocytes using FIB-SEM.

The oocyte plays a pivotal role in the reproduction of our species. Nevertheless, its biology remains poorly understood. Electron microscopy is traditionally used to inspect the ultrastructure of female gametes. However, two-dimensional micrographs contain only fragmentary information about the spatial organization of the complex oocyte cytoplasm. Here, we employed the Focused Ion Beam Scanning Electron Microscopy (FIB-SEM) to explore human oocyte intracellular morphology in three dimensions (3D). Volume reconstruction from high-resolution image stacks provided an unprecedented view of ooplasmic architecture. Organelle distribution patterns observed in 9 donor oocytes, representing 3 maturational stages, documented structural changes underlying the process by which the egg acquires developmental competence. 3D image segmentation was performed to extract information about distinct organelle populations. The quantitative analysis of the organelle abundance revealed that mitochondrion occupies ~ 4.26 % of the maturing oocyte cytoplasm. This proof-of-concept study demonstrates the potential of FIB-SEM imaging to study human oocyte morphology.

cell biology↗

bioRxiv2020

3D cell culture models demonstrate a role for FGF and WNT signaling in regulation of lung epithelial cell fate and morphogenesis

FGF signaling plays an essential role in lung development, homeostasis, and regeneration. Several FGF ligands were detected in the developing lungs, however, their roles have not been fully elucidated. We employed mouse 3D cell culture models and imaging to ex vivo study of a) the role of FGF ligands in lung epithelial morphogenesis and b) the interplay of FGF signaling with epithelial growth factor (EGF) and WNT signaling pathways. In non-adherent conditions, FGF signaling promoted formation of lungospheres from lung epithelial stem/progenitor cells (LSPCs). Based on their architecture, we defined three distinct phenotypes of lungospheres. Ultrastructural and immunohistochemical analyses showed that LSPCs produced more differentiated lung cell progeny. In 3D extracellular matrix, FGF2, FGF7, FGF9, and FGF10 promoted lung organoid formation with similar efficiency. However, FGF9 showed reduced capacity to promote lung organoid formation, suggesting that FGF9 has a reduced ability to sustain LSPCs survival and/or initial divisions. Analysis of lung organoid phenotypes revealed that FGF7 and FGF10 produce bigger organoids and induce organoid branching with higher frequency than FGF2 and FGF9. Higher FGF concentration and/or the use of FGF2 with increased stability and affinity to FGF receptors both increased lung organoid and lungosphere formation efficiency, respectively, suggesting that the level of FGF signaling is a crucial driver of LSPC survival and differentiation, and also lung epithelial morphogenesis. EGF signaling played a supportive but nonessential role in FGF-induced lung organoid formation. Moreover, analysis of tissue architecture and cell type composition confirmed that the lung organoids contained alveolar-like regions with cells expressing alveolar type I and type II cell markers, as well as airway-like structures with club cells and ciliated cells. WNT signaling enhanced the efficiency of lung organoid formation, but in the absence of FGF10 signaling, the organoids displayed limited branching and less differentiated phenotype. In summary, we present lung 3D cell culture models as useful tools to study the role and interplay of signaling pathways in lung development and we reveal roles for FGF ligands in regulation of mouse lung morphogenesis ex vivo.

developmental biology↗