Biology subjects

Szafranska, K.

Publications and source records attributed to Szafranska, K..

4 recordsLinked to original sources

bioRxiv2023

Highly oxidized albumin is mainly cleared by mouse liver sinusoidal endothelial cells via the receptors stabilin-1 and -2

BackgroundOxidized albumin (oxHSA) is elevated in several pathological conditions, especially those involving the liver, such as decompensated cirrhosis, acute on chronic liver failure and liver mediated renal failure. Patient derived oxidized albumin was previously shown to be an inflammatory mediator in cultured endothelial cells and leukocytes. The removal from circulation of oxidized albumins is therefore essential for maintenance of homeostasis. Normal serum levels of oxidized albumin are low, implying it is constantly eliminated. Liver sinusoidal endothelial cells (LSEC) are prominent scavenger cells in the body, specializing in the removal of macromolecules e.g. hyaluronan, denatured collagen, modified albumins, bacterial endotoxin (LPS) and oxidized lipoprotein. Given that oxidized albumin is mainly cleared by the liver, we hypothesize the LSEC are the site of uptake in the liver. Furthermore the stabilins -1 and -2 are the most prominent candidates for oxHSA uptake receptors, given their expression pattern and uptake of other ligands. MethodsIn vivo biodistribution, hepatocellular distribution and in vitro uptake studies on isolated liver cell populations or receptor expressing cell lines. ResultsIn vivo oxHSA was cleared rapidly (t1/2 <90seconds) by the liver (47% of uptake) and distributed to mainly the LSEC. In in vitro studies LSEC endocytosed oxHSA much more than other cell populations isolated from the liver. Furthermore, it was shown that the uptake was mediated by the stabilins, by inhibiting uptake in LSEC with other stabilin ligands and showing uptake in HEK cells overexpressing stabilin-1 or 2. oxHSA also inhibited the uptake of other stabilin ligands. ConclusionsLSEC and their stabilins are vital for the clearance of oxidized albumin, and therefore play a pivotal role in maintaining homeostasis.

molecular biology↗

bioRxiv2023

High-speed TIRF and 2D super-resolution structured illumination microscopy with large field of view based on fiber optic components

Super-resolved structured illumination microscopy (SR-SIM) is among the most flexible, fast, and least perturbing fluorescence microscopy techniques capable of surpassing the optical diffraction limit. Current custom-built instruments are easily able to deliver two-fold resolution enhancement at video-rate frame rates, but the cost of the instruments is still relatively high, and the physical size of the instruments based on the implementation of their optics is still rather large. Here, we present our latest results towards realizing a new generation of compact, cost-efficient, and high-speed SR-SIM instruments. Tight integration of the fiber-based structured illumination microscope capable of multi-color 2D- and TIRF-SIM imaging, allows us to demonstrate SR-SIM with a field of view of up to 150 x 150 m2 and imaging rates of up to 44 Hz while maintaining highest spatiotemporal resolution of less than 100 nm. We discuss the overall integration of optics, electronics, and software that allowed us to achieve this, and then present the fiberSIM imaging capabilities by visualizing the intracellular structure of rat liver sinusoidal endothelial cells, in particular by resolving the structure of their trans-cellular nanopores called fenestrations.

biophysics↗

bioRxiv2023

Effect of caffeine and other xanthines on liver sinusoidal endothelial cell ultrastructure

Xanthines such as caffeine and theobromine are among the most consumed psychoactive stimulants in the world, either as natural components of coffee, tea and chocolate, or as food additives. The present study assessed if xanthines affect liver sinusoidal endothelial cells (LSEC). Cultured primary rat LSEC were challenged with xanthines at concentrations typically obtained from normal consumption of xanthine-containing beverages, food or medicines; and at higher concentrations below the in vitro toxic limit. The fenestrated morphology of LSEC were examined with scanning electron and structured illumination microscopy. All xanthine challenges had no toxic effects on LSEC ultrastructure as judged by LSEC fenestration morphology, or function as determined by endocytosis studies. All xanthines in high concentrations (150 g/mL) increased fenestration frequency but at physiologically relevant concentrations, only theobromine (8 g/mL) showed an effect. LSEC porosity was influenced only by high caffeine doses which also shifted the fenestration distribution towards smaller pores. Moreover, a dose-dependent increase in fenestration number was observed after caffeine treatment. If these compounds induce similar changes in vivo, age-related reduction of LSEC porosity can be reversed by oral treatment with theobromine or with other xanthines using targeted delivery.

cell biology↗

bioRxiv2022

Dual-color single molecule localization microscopy on transparent polymer waveguide chips

Photonic waveguide chips offer near-field excitation of biological samples, which enables cost-effective, large field-of-view super-resolution microscopy without the need for high numerical aperture (NA) objective lenses. Single molecule localization based super-resolution microscopy that requires high illumination intensities is currently limited to solid state photonic waveguide chips composed of hard-coated, high NA planar waveguides deposited on opaque substrates. These platforms do not permit epi-detection of fluorescence through the substrate, which limits the use of photonic waveguide chips to the upright configuration. Additionally, the detection efficiency is reduced because the majority of the fluorescence emission is directed towards the high refractive index substrate. A low cost waveguide chip based on a polymer core material deposited on common #1.5 coverslips that is easy to produce was recently demonstrated. Here, a platform that is capable of performing single-molecule localization microscopy (SMLM) of biological samples using polymer-based photonic waveguide chips is presented, enabling super-solution microscopy in the inverted microscope configuration. Super-resolved imaging of two different structures of the cytoskeleton in primary liver sinusoidal endothelial cells (LSECs) by two popular SMLM methods, dSTORM and DNA-PAINT, down to 23 nm is demonstrated.

biophysics↗