COMPARATIVE ATOMIC-FORCE AND SCANNING ELECTRON-MICROSCOPY - AN INVESTIGATION ON FENESTRATED ENDOTHELIAL-CELLS IN-VITRO

Rat liver sinusoidal endothelial cells (LEG) contain fenestrae, which are clustered in sieve plates, Fenestrae control the exchange of fluid s, solutes and particles between the sinusoidal blood and the space of Disse, which at its back side is flanked by the microvillous surface of the parenchymal cells. The surface of LEC can optimally be imaged b y scanning electron microscopy (SEM), and SEM images can be used to st udy dynamic changes in fenestrae by comparing fixed specimens subjecte d to different experimental conditions. Unfortunately, the SEM allows only investigation of fixed, dried and coated specimens, Recently, the use of atomic force microscopy (AFM) was introduced for analysing the cell surface, independent of complicated preparation techniques, We u sed the AFM for the investigation of cultured LEC surfaces and the stu dy of morphological changes of fenestrae, SEM served as a conventional reference. AFM images of LEC show structures that correlate well with SEM images. Dried-coated, dried-uncoated and wet-fixed LEC show a cen tral bulging nucleus and flat fenestrated cellular processes, It was a lso possible to obtain height information which is not available in SE M, After treatment with ethanol or serotonin the diameters of fenestra e increased (+6%) and decreased (-15%), respectively, The same alterat ions of fenestrae could be distinguished by measuring AFM images of dr ied-coated, dried-uncoated and wet-fixed LEG. Comparison of dried-coat ed (SEM) and wet-fixed (AFM) fenestrae indicated a mean shrinkage of 2 0% in SEM preparations, In conclusion, high-resolution imaging with AF M of the cell surface of cultured LEC can be performed on dried-coated , dried-uncoated and wet-fixed LEG, which was hitherto only possible w ith fixed, dried and coated preparations in SEM and transmission elect ron microscopy (TEM).