Liliane Dubuisson

Hepatocyte ultrastructure in rats with portacaval shunt

The principal reported morphological consequence of portacaval shunt in the rat is liver atrophy. The present study was designed to investigate ultrastructural changes in hepatocytes using electron microscopy morphometry. Two weeks following portacaval shunt, rat livers were fixed by perfusion and hepatocyte organelles from the two opposite zones of the acinus (zones 1 and 3) were quantified. Liver weight/body weight decreased by 50%, hepatocyte-specific volume decreased by 30% (28% in zone 1 and 35% in zone 3). Estimated sinusoidal space increased, and estimated number of hepatocytes decreased by 50%. Hepatocytes had a normal ultrastructure except for mitochondria. Smooth endoplasmic reticulum-specific surface area was reduced by 65% (zone 3), and rough endoplasmic reticulum surface density was increased in zone 1 only. Mitochondriaspecific volume was unchanged but decreased inner and outer membrane-specific surface area in zone 3 suggests in this zone a change in their conformation and possibly their number. Golgi-rich area surface density increased but not significantly. Hepatocyte loss and atrophy and rearrangement of organelles represent a new ultrastructural steady state following portacaval shunt that may help explain the new functional steady state.

Liver arterialization improves hepatocytes ultrastructure in rats with portacaval shunts

The effect of arterialization (ART) of the distal stump of the portal vein after portacaval shunt (PCS) on bile formation and liver ultrastructure was assessed. ART using the left gastric artery was performed in male Wistar rats. Animals were sacrificed 3 weeks later. ART prevented body and liver atrophy. However, the liver weight to body weight ratio was significantly decreased when compared to sham PCS (2.5±0.36 vs 3.22±0.15). Reduction in total bile secretion (μl/min) seen following PCS is reversed by ART. ART partly corrected hepatocyte size atrophy and the major ultrastructural abnormalities, namely the irregularity of the nucleus and the dilatation of the nuclear envelope and of the rough endoplasmic reticulum appearing after PCS. However, mitochondria remained swollen, deformed, and enlarged with scission figures. No lesions in connection with ART were seen. This result confirms, at the ultrastructural level, the beneficial effect of ART in PCS.

Benign Recurrent Cholestasis: A Light and Electron Microscopic Study with Emphasis on Sinusoidal Cells

A liver biopsy was performed on a patient with benign recurrent cholestasis. Cholestasis was mainly centrolobular with infiltration by sinusoidal macrophages. There was no necrosis. All the classic and specific ultrastructural criteria of cholestasis were observed in hepatocytes under electron microscopy. Perfusion-fixation of the biopsy allowed in addition a good visualization of sinusoids and sinusoidal cells. Numerous macrophages (Kupffer cells) with intense phagocytic activity were present in the lumen; some formed the sinusoidal barrier or were infiltrated in the Disse space. Endothelial cells contained numerous dense bodies and had few fenestrae. Cellular debris of hepatocytic origin which were not phagocytized in the Disse space were extruded in the lumen either through enlarged endothelial pores or by progressive invagination in the endothelial wall followed by outpouching in the sinusoid. In an enlarged Disse space containing amorphous material and collagen fibrils some perisinusoidal cells were transitional cells. These results indicate that pure cholestasis leads not only to hepatocyte injury with intense phagocytic activity but also to some degree of sinusoidal cell damage and extracellular matrix changes.

The sinusoidal barrier in rats with portacaval anastomosis: a morphometric study.

Shunting of portal blood in the rat leads to liver atrophy and to an increase in arterial blood flow with microcirculatory disturbances. The aim of this study was to investigate the effects of these disturbances on the liver sinusoidal barrier (endothelial and perisinusoidal cells) using morphometric techniques. Rats with portacaval anastomosis (PCA) and sham operated pair-fed controls were studied 3 months after the shunt. Sinusoidal volume density in PCA increased but not significantly and the volume density (Vv) of total endothelial (EC) and perisinusoidal cells (PSC) increased by 104.54% compared to sham operated pair-fed rats. The increase of EC Vv was not associated with an increase in surface density (Sv) suggesting a fall in the number of small fenestrations and an increase in cell thickness. This interpretation supports the morphological observations. The increase of PSC Vv was mainly related to the increase in their subendothelial processes Vv and not to that of the cell body Vv. Lipids Vv and RER Sv expressed per sinusoidal cells remained unchanged suggesting that the balance between the 2 hypothetical functions of the PSC, namely fibrogenesis and storage of vitamin A, was maintained. In conclusion, changes of EC and PSC after PCA result mainly in thickening of the sinusoidal barrier. This increase may impair exchanges between the sinusoidal lumen and Disse space and contribute to functional abnormalities.

Quasi-real time digital stereology in electron microscopy

Summary— We investigated the possibility of applying classical stereological methods to digitized images in quasi‐real time. Stereological lattices are superimposed on the image of an analyser fitted to a transmission electron microscope via a video camera. Estimations of stereological parameters can be obtained in a very short time even with a complicated object contour.

Removal of cellular debris formed in the Disse space in patients with cholestasis.

Using electron microscopy, we investigated how cellular debris, formed in the Disse space during cholestasis, was cleared. Ten patients with cholestasis of varied origin and severity were studied and compared with 10 controls without liver disease. In cholestatic patients, sinusoidal cells contained variable amounts of amylase PAS-positive material. In clean perfusion-fixed sinusoids the endothelial cells often appeared swollen and active, with few fenestrations. Hepatocyte blebs and cellular debris were sometimes seen in the Disse space. Two mechanisms were apparently involved in the clearing process: phagocytosis by macrophages either infiltrated into the Disse space, or forming the barrier; and the passage of debris from the Disse space into the sinusoidal lumen through the endothelial wall. Debris was either forced through enlarged pores or through the wall, with a progressive invagination followed by an outpouching in the lumen. The force, possibly provided by endothelial massage, may not be sufficient to push out cellular debris from the Disse space; morphological data seemed to indicate that endothelial damage may be a necessary factor. Debris present in the lumen was phagocytized by numerous active macrophages. Cellular debris was not observed in the Disse space of control patients.

Trans-Resveratrol and the Liver: Deactivation of Liver Myofibroblasts and Inhibition of Tumor Cell Invasion

In several situations, mesenchymal fibroblasts acquire a myofibroblastic phenotype and express characteristic cytoskeletal proteins, such as smooth muscle α-actin (α—SMA). Myofibroblasts are present for a limited time in normal wound healing, but persist when tissue repair is abnormal, i.e., during tissue fibrosis and in the stromal reaction to tumors1. Myofibroblasts are believed to be essential actors of tissue fibrosis. They are involved in tissue contraction, synthesis of extracellular matrix and synthesise cytokines. In the context of cancer, there are less data, but strong evidence point to a role of infiltrating myofibroblasts in invasion2.