Biology subjects

Nüsse, O.

Publications and source records attributed to Nüsse, O..

4 recordsLinked to original sources

bioRxiv2025

Adenovirus phagocytosis by neutrophils triggers a pro-inflammatory response

Adenoviruses are common pathogens that have been engineered and used for medical purposes. Their recognition by innate immune cells such as macrophages and dendritic cells has been widely studied, but very little information is available on adenovirus interaction with neutrophils. Using cytometry and confocal and electron microscopy, we showed that neutrophils bind to antibody-coated adenoviruses and engulf them in a phagosome. Using a single-cell transcriptomic approach, we found that adenovirus phagocytosis activates a specific transcriptional program in neutrophils. It also triggers calcium entry into neutrophils, reactive oxygen species production in the phagosome, and CXCL8 release. Moreover, 4 hours after adenovirus incubation, 50% of neutrophils undergo calcium- and RIPK3-dependent cell death, accompanied by Neutrophil Extracellular Trap emission. Because of this rapid cell death, engulfed adenoviruses are not fully degraded after 3 hours, and the remaining viruses are still capable of expressing their genome in target cells. Thus, our data suggest that, during adenoviral infection, the neutrophil response may promote a pro-inflammatory environment that could damage host tissues. Author SummaryAdenoviruses are common viruses that trigger mild infections, although they can lead to more severe diseases such as gastroenteritis or respiratory tract infections in immunocompromised patients. Non-replicative adenovirus-derived vectors are widely used in medicine for vaccination and antitumor therapy. In this study, we investigated how adenoviruses affect neutrophils, the first immune cells recruited to the site of infection and crucial players in the immune system. We used various methods to explore this interaction. We found that adenoviruses bind to neutrophils, and that this binding is enhanced by human serum containing antibodies against the viruses. This binding is followed by adenovirus internalization, and we showed that it triggers several responses. These responses include the activation of specific genes, the entry of calcium into the cells, the production of reactive oxygen species (molecules that fight infections), and the release of signaling molecules to attract other immune cells. However, this process also leads to rapid neutrophil death and the release of structures called Neutrophil Extracellular Traps. Because of this quick death, adenoviruses are not fully broken down and can still infect other cells. We suggest that this neutrophil response might create an inflammatory environment that could potentially harm the bodys tissues.

immunology↗

bioRxiv2025

Stiffening cells with light

Fluorescence microscopy is widely used to observe structures and dynamic processes in living cells and organisms and is often used as if it were purely innocuous to the cells or structures of interest. However, it can lead to phototoxicity, which can affect the cellular behavior and lead to erroneous interpretations of the observations. The major cause of cell damage through phototoxicity is the production of reactive oxygen species (ROS), which can form crosslinks between intracellular molecules, including proteins and nucleic acids. By using profile microindentation and atomic force microscopy, we demonstrate that the excitation of various fluorescent probes leads to a large increase in the stiffness of several cell types within seconds of illumination. The stiffening exhibits a dose-dependent response, where longer exposure times to exciting light are correlated with larger stiffening. This photostiffening effect explains why T cells loaded with the Fluo-4 Calcium probe stop emitting a protrusion within seconds after the excitation light is turned on. We observed photostiffening in different cell types and fluorophores. We showed that repeated cell indentation alone led to cell stiffening as well as excitation with blue or UV light in the absence of a fluoroph ore. However, in the latter case, the stiffening was much smaller than that when the fluorophore was excited. We used both sharp and blunt indenters to show that stiffening occurred not only at the cell cortex level but also deeper in the cell interior and that photostiffening is independent of actin cytoskeleton organization. We correlated the increase in cell stiffness with the production of intracellular reactive oxygen species and reproduced cell stiffening by incubating cells with a ROS inducer, H2O2. The excitation of the photosensitizer Pheophorbide a, which induces a specific type of reactive oxygen species, namely singlet oxygen, also led to cell stiffening. This study reminds the experimentalists that it is crucial to perform controls when using fluorescence. It further allows us to propose exploiting photostiffening as a new method for rapidly quantifying phototoxicity. SignificanceThis study reveals a direct relationship between fluorescence excitation and cell mechanical properties. The generality of th is phenomenon across diverse fluorophores and cell types highlights the importance of controlling phototoxicity in fluorescence experiments, in particular in complex, quantitative cell biology and biophysical experiments, but also reveals an unexpected rol e of intracellular reactive oxygen species production and identifies cell stiffness as a proxy for assessing the efficacy of photodynamic therapy.

biophysics↗

bioRxiv2021

Consequences of the constitutive NOX2 activity in living cells: cytosol acidification, apoptosis, and localized lipid peroxidation

The phagocyte NADPH oxidase (NOX2) is a key enzyme of the innate immune system generating superoxide anions (O2*-), precursors of reactive oxygen species. The NOX2 protein complex is composed of six subunits: two membrane proteins (gp91phox and p22phox) forming the catalytic core, three cytosolic proteins (p67phox, p47phox and p40phox) and a small GTPase Rac. The sophisticated activation mechanism of the NADPH oxidase relies on the assembly of cytosolic subunits with the membrane-bound components. A chimeric protein, called Trimera, composed of the essential domains of the cytosolic proteins p47phox (aa 1-286), p67phox (aa 1-212) and full-length Rac1Q61L, enables a constitutive and robust NOX2 activity in cells without the need of any stimulus. We employed Trimera as a single activating protein of the phagocyte NADPH oxidase in living cells and examined the consequences on the cell physiology of this continuous and long-term NOX activity. We showed that the sustained high level of NOX activity causes acidification of the intracellular pH, triggers apoptosis and leads to local peroxidation of lipids in the membrane. These local damages to the membrane correlate with the strong tendency of the Trimera to clusterize in the plasma membrane observed by FRET-FLIM microscopy. HighlightsO_LITrimera is a tool to trigger a continuous ROS production in living cells C_LIO_LIContinuous NOX2 activity causes cytosol acidification and apoptosis C_LIO_LIROS overproduction leads to localized oxidation of the membrane lipids C_LIO_LITrimera tends to clusterize in the plasma membrane of COSNOX and COS-7 cells C_LI

biophysics↗

bioRxiv2019

Single-cell immuno-mechanics: rapid viscoelastic changes are a hallmark of early leukocyte activation.

To accomplish their critical task of removing infected cells and fighting pathogens, leukocytes activate by forming specialized interfaces with other cells. Using an innovative micropipette rheometer, we show in three different cell types that when stimulated by microbeads mimicking target cells, leukocytes become up to ten times stiffer and more viscous. These mechanical changes initiate within seconds after contact and evolve rapidly over minutes. Remarkably, leukocyte elastic and viscous properties evolve in parallel, preserving a well-defined ratio that constitutes a mechanical signature specific to each cell type. The current results indicate that simultaneously tracking both elastic and viscous properties during an active cell process provides a new way to investigate cell mechanical processes. Our findings also suggest that dynamic immuno-mechanical measurements provide an identifier of leukocyte type and an indicator of the cells state of activation.

biophysics↗