Gene regulation of melatonin and dopamine receptors during eye development.

To gain insight into the role of melatonin and dopamine in retinal development, gene expression of two melatonin receptors, MT1 and MT2, as well as five dopamine receptors, D1, D2, D3, D4 and D5, in the rat eye was analyzed by reverse transcription-polymerase chain reaction across various developmental stages. MT1 transcript levels reached maximum levels at embryonic day (E) 16 and then decreased gradually until reaching adult levels by postnatal day (P) 14. MT2 transcript levels similarly peaked at E16, but then decreased dramatically until birth to its lowest levels, which were maintained throughout the postnatal period. Thus, gene expression of both the MT1 and MT2 receptors showed a striking inverse correlation with maturation of the eye. In contrast to melatonin receptors, gene expression of all dopamine receptor subtypes, except for D3, showed only an increase as development proceeds with highest levels in adulthood. The D3 message was not detected throughout the developmental period examined. Gene expression of D1-like receptors, D1 and D5, showed a substantial increase to adult levels during the fetal period at E16 and E20, respectively. Transcript levels of D2-like receptors, D2 and D4, on the other hand, were not detected before birth but increased significantly to adult levels by P7 and P14, respectively. The present findings suggest the presence of unique developmental mechanisms by which transcription of various G protein-coupled receptors are regulated in the eye.

High doses of intravenous immunoglobulin G enhance Kupffer cell phagocytic function during the late phase of sepsis and endotoxemia in rats.

The effect of intravenous immunoglobuln G (ivIG) on the hepatic microvascular inflammatory response during the late phase of sepsis and endotoxemia in rats was studied by using in vivo microscopy. One hour after administration of a clinically relevant dose of ivIG (0.5 g/kg body weight, Sandoglobulin), rats were subjected to polymicrobial sepsis induced by cecal ligation and puncture (CLP) or were injected intravenously with lipopolysaccharide (LPS, 0.1 mg/kg body weight). Twenty-four hours after CLP or LPS, the number of leukocytes adhering to the sinusoidal wall was increased 11.0-fold in CLP-treated animals and 5.6-fold in LPS-treated animals, respectively, compared with the controls. Concomitantly, the numbers of swollen sinusoidal endothelial cells were increased 4.2-fold and 3.2-fold. The number of perfused sinusoids was decreased by 35% and by 24%. These responses were minimized by pretreatment with high doses of ivIG. Kupffer cell phagocytic activity in the periportal sinusoids in CLP-treated animals was decreased by 41%, whereas that in the centrilobular sinusoids in LPS-treated animals was increased by 72%. IvIG significantly elevated this activity in both CLP- and LPS-treated animals and the number of ED2-positive Kupffer cells in tissue sections. The results suggest that ivIG limits the hepatic microvascular inflammatory response during the late phase of sepsis and endotoxemia by affecting Kupffer cell function.

Effect of intravenous immunoglobulin G on the TNFalpha-mediated hepatic microvascular inflammatory response.

The effect of intravenous immunoglobulin G (ivIG) on the hepatic microvascular inflammatory response elicited by tumor necrosis factor alpha (TNFalpha) in rats was studied by means of in vivo microscopy and histological examination. One hour after the portal infusion of TNFalpha, the average number of leukocytes adhering to the sinusoidal endothelium was increased sevenfold, and the average number of the perfused sinusoids was decreased by 15% when compared with controls. Concomitantly, the expression of intercellular adhesion molecule-1 (ICAM-1) on the hepatic sinusoidal endothelium and that of the central vein was increased. The phagocytic activity of Kupffer cells in centrilobular sinusoids was increased by 54%, as were the number of ED2-positive Kupffer cells in tissue sections. Pretreatment with a clinically relevant high dose of ivIG (1 g/kg body weight, Sandoglobulin) minimized these responses by reducing leukocyte-endothelial interactions and Kupffer cell phagocytic function. The results suggest that high doses of ivIG limit the hepatic microvascular inflammatory response by inhibiting the action of the proinflammatory cytokine TNFalpha.

The stellate cells phenotypic transformation in the CCl4- injured liver fibrosis of ICR mice: their desmin immunoreactivity and vitamin A storage.

To elucidate the process of phenotypic transformation of stellate cells which were responsible in the production of collagen fibers, ICR mice were injected subcutaneously with CCl4 twice weekly for 5 weeks and the livers were examined by light and electron microscopy every week. Stellate cells were detected by desmin immunoreaction and vitamin A-autofluorescence. In the first week, following cell necrosis in the pericentral areas, granulomas were formed. Two distinct stellate cell phenotypes were developed i.e. desmin-negative cells in the granuloma and a row of intense desmin-positive cells in its boundaries. Both phenotypes seemed to be derived mostly from perisinusoidal stellate cells and involved in collagen synthesis as evidenced by the presence of reticulin meshwork within the granulomas. The meshwork formation was the initial stage of a new septum formation since in the second week the intense desmin-positive cell rows were closer to the central vein concomitant with the growth of several protrusions to form thin layer septa towards neighboring central or portal veins. The myofibroblast-like cells in the septa with features interposed between stellate cells and myofibroblasts were likely derived from the stellate cells phenotypes. During the transformation of stellate cells, the decline of vitamin A storage was accompanied by the formation of connective tissue septa. The desmin immunoreactivity and vitamin A storage in the stellate cell phenotypes during liver fibrogenesis described in this study may reveal their phenotypic transformation in association with the collagen synthesis.

Granuloma formation in the liver of Balb/c mice intoxicated with carbon tetrachloride.

Hepatic granulomas induced by a single or several subcutaneous injections of carbon tetrachloride (CCl4) in Balb/c mice were examined electronmicroscopically and immunocytochemically. Stellate cells (fat-storing cells; lipocytes; Ito cells) were identified by the detection of cytoplasmic desmin, while T-lymphocytes and monocytes/macrophages were identified with monoclonal antibodies Thy 1.2 and MOMA-2, respectively. Following pericentral necrosis induced with CCl4, clear foci containing lymphocytes, monocytes, macrophages and perisinusoidal stellate cells occurred in the surrounding hepatic parenchyma on day 5. These clear foci developed to granulomas with increasing numbers of macrophages and stellate cells. Mitotic and apoptotic figures in randomly distributed macrophages, and direct contacts between macrophages and stellate cells were frequently seen within the granulomas. The stellate cells were characterized by a well-developed granular endoplasmic reticulum and Golgi complex. Collagen fibrils were closely applied to the stellate cells and connective tissue septa extended between neighboring granulomas and/or the pericentral necrotic areas after several injections of CCl4. CCl4-induced hepatic granulomas provide a model for investigating paracrine and/or autocrine modulation within a well-organized microenvironment during progressive hepatic inflammation and fibrosis.