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Investigation of liver sinusoidal endothelial cells - characterisation and application of new transgenic mouse models

Zierow, Johanna

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Abstract

Liver sinusoidal endothelial cells (LSECs) constitute organ-specific microvascular endothelial cells (ECs) with unique phenotypical and functional characteristics. They form a discontinuous endothelial sheet at the interface between the blood stream and the hepatocytes and are the most permeable vascular barrier of the mammalian body. LSECs express endocytosis receptors that clear blood borne waste molecules and ensure thereby tissue homeostasis. Under pathological conditions they participate in development and progression of chronic liver diseases or promote liver regeneration through secretion of angiocrine factors. So far, most of LSEC research performed with mouse models relies on transgenic mice that demonstrated off-target effects in a variety of vascular beds. In vitro cell culture techniques only enable short-term cultivation of LSECs due to the subsequent dedifferentiation and are therefore of limited utility. To target the microvasculature of the liver more specifically transgenic mouse models were generated in which the bacteriophage-derived recombinase Cre is controlled by promoter elements of LSEC characteristic proteins. Two different founders of the Stab2-cre mouse and one founder of the Clec4g-cre mouse were characterised by reporter mouse models. All of three transgenic mouse lines demonstrated recombination in LSECs, but also in arterial and venous liver ECs. Apart from the liver, the Stab2- and Clec4g-cre mice target sinusoidal endothelial beds of other organs. A transgene activity in subpopulations of haematopoietic cells has been observed in two of three mouse lines. The pattern of Cre activity was timedependent. Stab2-cre founder 2 mouse for example showed sinusoidal EC activity during development that broadened to most ECs in other organs during adulthood. Therefore, the transgenic mouse models can be advantageous when manipulating LSEC specific protein expression or using them for cell fate tracking. An appropriate model should be selected depending on the gene of interest and the developmental period to be investigated. In comparison to conventional and inducible endothelial transgenic mouse lines these mouse models exhibit reduced off-target effects in ECs of other organs and homogenous Cre activity in LSECs, respectively. GATA4 was identified in a cluster of transcription factors (TFs) overexpressed in rat LSECs compared to rat lung microvascular ECs. GATA4 has a fundamental role during cardiac development and in cardiac ECs precluding the analysis of GATA4 in LSECs by conventional endothelial Tek- or Cdh5-cre mice. Consequently, investigation of the function of GATA4 in LSECs required a transgenic mouse line that would induce GATA4 loss in LSECs but not in the developing heart. Therefore, Stab2-cre founder 2 mouse was utilised to generate a conditional GATA4 knockout in LSECs. The knockout mouse demonstrated embryonic lethality with a reduced size of the fetal liver (FL) and a impaired hepatic vasculature. The FL endothelium transformed from discontinuous liver sinusoids to continuous capillaries. The capillarized ECs impaired the immigration of haematopoietic stem and progenitor cells (HSPCs) into the FL leading to lethal anaemia. Together, the data identify GATA4 as a master regulator for liver sinusoidal EC specification and for acquisition of organ-specific vascular competence supporting liver development. Finally, the results establish a role of the FL sinusoidal endothelium on maturation of HSPCs.

Item Type: Dissertation
Supervisor: Sleeman, Prof. Dr. Jonathan
Place of Publication: Heidelberg
Date of thesis defense: 22 June 2018
Date Deposited: 26 Jul 2018 11:58
Date: 2018
Faculties / Institutes: The Faculty of Bio Sciences > Dean's Office of the Faculty of Bio Sciences
Medizinische Fakultät Mannheim > Hautklinik
Subjects: 570 Life sciences
Controlled Keywords: Leber, Endothelzellen
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