Towards understanding the dynamic structure of fenestrations in living liver sinusoidal endothelial cells

Liver Sinusoidal Endothelial Cells(LSECs) contribute to maintain transportation of lipoproteins and chylomicrons between bloodstream and a space of Disse. Filtration is provided by the presence of trans-membrane pores with... [ view full abstract ]

Liver Sinusoidal Endothelial Cells(LSECs) contribute to maintain transportation of lipoproteins and chylomicrons between bloodstream and a space of Disse. Filtration is provided by the presence of trans-membrane pores with diameter in range of 80–200 nm, called fenestrations[1]. The most of research on this structure has been performed on fixed cells using electron microscopy[1]. In our work we investigate a dynamic rearrangement of the cell actin cytoskeleton connected with formation and closing of fenestrations in living LSECs using atomic force microscopy (AFM). LSECs were isolated from murine liver and seeded on glass coverslips. AFM measurements were conducted within 24h to avoid loss of fenestrations[2]. We apply different AFM work modes that yield different kind
of information on both topography and nanomechanical properties of LSECs[3]. Obtained data allowed for confirmation of the dynamic rearrangement of the structure of fenestrations. Moreover, quantitative imaging provides an insight into the process of fenestration formation and cytoskeletal alteration after exposition 
to factors influencing actin polymerization: cytochalasin, jasplakinolide, antimycin. We also identify Fenestration Forming Centers(FFC) responsible for increasing the number of pores. The results indicate that due to the recent development of AFM imaging, investigation of liver sinusoidal endothelial cells could go beyond the standard analysis capabilities of the electron microscopy. This AFM-based tool provides new possibilities to significantly improve studies on both impact of different factors on LSECs morphology and detailed studies of the fenestration dynamic structure.

Bibliography:

[1]F.Braet and E.Wisse,Comparative Hepatology1:1,2002

[2]DeLeve et al., Am J Physiol Gastrointest Liver Physiol.287(4),2004

[3]Zapotoczny et al.,J Mol Recogne2610,2017