125I-Insulin is preferentially internalized in regions of the hepatocytes rich in lysosomal and Golgi structures.

125I-Insulin initially localizes to the plasma membrane of isolated rat hepatocytes but is subsequently internalized and preferentially associates with lysosomal structures. In the present study, we show that this preferential association to lysosomes occurs in regions of the cell rich in lysosomal and Golgi structures.     

Intramitochondrial complexes of atypical structures in hepatocytes ofTriturus alpestris (Laurenti)

Résumé Des structures lamellaires sont décrites dans la matrice mitochondriale du foie de triton (Triturus alpestris).     

Analysis of a sub-proteome which co-purifies with and is phosphorylated by the Golgi casein kinase

In an attempt to gain information about the identity of the Golgi apparatus casein kinase(s) (G-CK), responsible for the phosphorylation of caseins in lactating mammary gland, the proteins present in fractions enriched in G-CK activity eluted from DEAE-Sepharose and heparin-Sepharose columns were resolved by two-dimensional electrophoresis and analyzed by mass spectrometry. This led to the identification of 47 proteins altogether, none of which is a bona fide protein kinase. At least 9 of the identified proteins however, are readily phosphorylated by co-purifying G-CK activity, and 7 are physically associated with it to give supramolecular complex(es) of about 500 kDa as judged from Superdex S200 gel fitration and glycerol gradient ultracentrifugation experiments. In contrast, the apparent molecular weight of G-CK estimated from an in gel activity assay after SDSPAGE and renaturation is about 41 kDa. Many of the proteins phosphorylated by and/or associated with G-CK belong to the category of chaperonines, including HSP90, GRP-94 and −78, and various isoforms of protein disulfide isomerases, suggesting a global role of this kinase in the modulation of protein folding. Received 21 October 2005; received after revision 30 November 2005; accepted 6 December 2005 †These authors contributed equally to this work.     

Heparanase is preferentially expressed in human psoriatic lesions and induces development of psoriasiform skin inflammation in mice

Heparanase is the sole mammalian endoglycosidase that selectively degrades heparan sulfate, the key polysaccharide associated with the cell surface and extracellular matrix of a wide range of tissues. Extensively studied for its capacity to promote cancer progression, heparanase enzyme was recently implicated as an important determinant in several inflammatory disorders as well. Applying immunohistochemical staining, we detected preferential expression of heparanase by epidermal keratinocytes in human psoriatic lesions. To investigate the role of the enzyme in the pathogenesis of psoriasis, we utilized heparanase transgenic mice in a model of 12-O-tetradecanoyl phorbol 12-myristate 13-acetate-induced cutaneous inflammation. We report that over-expression of the enzyme promotes development of mouse skin lesions that strongly recapitulate the human disease in terms of histomorphological appearance and molecular/cellular characteristics. Importantly, heparanase of epidermal origin appears to facilitate abnormal activation of skin-infiltrating macrophages, thus generating psoriasis-like inflammation conditions, characterized by induction of STAT3, enhanced NF-κB signaling, elevated expression of TNF-α and increased vascularization. Taken together, our results reveal, for the first time, involvement of heparanase in the pathogenesis of psoriasis and highlight a role for the enzyme in facilitating abnormal interactions between immune and epithelial cell subsets of the affected skin. Heparanase inhibitors (currently under clinical testing in malignant diseases) could hence turn highly beneficial in psoriatic patients as well.     

Autophagy and lysosomal proteolysis in the liver

Summary Autophagy is defined as any process whereby cellular macromolecules destined for degradation gain access to the lysosomes. A review is presented on the physiological significance, mechanisms and regulation of autophagy in hepatocytes, concentrating on the issue of regulation. The article ends by discussing techniques available for future research.     

Autophagy and lysosomal proteolysis in the liver

Autophagy is defined as any process whereby cellular macromolecules destined for degradation gain access to the lysosomes. A review is presented on the physiological significance, mechanisms and regulation of autophagy in hepatocytes, concentrating on the issue of regulation. The article ends by discussing techniques available for future research.     

Alpha 1-adrenergic stimulation of ketogenesis and fatty acid oxidation is associated with inhibition of lipogenesis in rat hepatocytes

The effect of norepinephrine on fatty acid synthesis (3H2O incorporation into fatty acids), on fatty acid oxidation to CO2 and on ketogenesis was studied in isolated hepatocytes of fed rats. After incubation with norepinephrine (50 microM), lipogenesis was lower (5.7 +/- 1.1 nmoles 3H2O incorporated into fatty acids/mg dry weight/30 min) than in controls (7.5 +/- 1.7; n = 6, p less than 0.02). In contrast, (1-14C) palmitate conversion into total ketone bodies was increased to 10.9 +/- 1.8 nmoles/mg/30 min with norepinephrine, vs 8.5 +/- 1.6 in controls (p less than 0.05), and more (1-14C) palmitate was converted to 14CO2 with norepinephrine than in controls (1.48 +/- 0.10 nmoles/mg/30 min vs 1.06 +/- 0.11, p less than 0.05). The inhibitory effect of norepinephrine on lipogenesis was abolished by addition of the alpha 1-receptor blocker prazosin, but not by alpha 2 or beta-blockers. The results demonstrate that the ketogenic effect of norepinephrine is coupled with an inhibitory effect on lipogenesis which may be explained by diminished activity of acetyl-CoA carboxylase, diminished formation of malonyl-CoA and decreased activity of carnitine palmitoyl transferase I.     

Localization of cholesterol in the Golgi apparatus of cardiac muscle cells

Summary Filipin (a polyene) interacts with cholesterol in membranes, producing distinctive deformations that can be detected by freeze-fracture. The distribution of filipin-induced deformations in the Golgi apparatus of cardiac myocytes suggests a role for this organelle in the transformation of cholesterol-poor membrane to cholesterol-rich membrane.Acknowledgments. This work was supported by a grant (No. 779) from the British Heart Foundation. I thank Dr E. Massey and Mr A. Slade for their help. Filipin was generously donated by Upjohn Ltd, Crawley, Sussex, U.K.     

BMP9 is produced by hepatocytes and circulates mainly in an active mature form complexed to its prodomain

Bone Morphogenetic Protein 9 (BMP9) has been recently found to be the physiological ligand for the activin receptor-like kinase 1 (ALK1), and to be a major circulating vascular quiescence factor. Moreover, a soluble chimeric ALK1 protein (ALK1-Fc) has recently been developed and showed powerful anti-tumor growth and anti-angiogenic effects. However, not much is known concerning BMP9. This prompted us to investigate the human endogenous sources of this cytokine and to further characterize its circulating form(s) and its function. Analysis of BMP9 expression reveals that BMP9 is produced by hepatocytes and intrahepatic biliary epithelial cells. Gel filtration analysis combined with ELISA and biological assays demonstrate that BMP9 circulates in plasma (1) as an unprocessed inactive form that can be further activated by furin a serine endoprotease, and (2) as a mature and fully active form (composed of the mature form associated with its prodomain). Analysis of BMP9 circulating levels during mouse development demonstrates that BMP9 peaks during the first 3 weeks after birth and then decreases to 2 ng/mL in adulthood. We also show that circulating BMP9 physiologically induces a constitutive Smad1/5/8 phosphorylation in endothelial cells. Taken together, our results argue for the role of BMP9 as a hepatocyte-derived factor, circulating in inactive (40%) and active (60%) forms, the latter constantly activating endothelial cells to maintain them in a resting state.     

Endocrine control of lysosomal alterations in rat liver during the perinatal period

Summary Within hours of birth, some physical properties of liver lysosomes are modified. These alterations, which may be related to the autophagic vacuoles formation known to occur during this period, were inhibited by insulin administration. Glucagon, a potent inducer of autophagy in adult rat liver, did not anticipate this process in fetal liver. Our results suggest that the decrease of plasma insulin immediately after birth is a important factor in the development of hepatic autophagy.     

Endocrine control of lysosomal alterations in rat liver during the perinatal period

Within hours of birth, some physical properties of liver lysosomes are modified. These alterations, which may be related to the autophagic vacuoles formation known to occur during this period, were inhibited by insulin administration. Glucagon, a potent inducer of autophagy in adult rat liver, did not anticipate this process in fetal liver. Our results suggest that the decrease of plasma insulin immediately after birth is an important factor in the development of hepatic autophagy.     

Biosynthesis of phosphatidylethanolamine from CDP-ethanolamine by the Golgi complex of rat liver in vitro

Summary A Golgi-rich fraction from rat liver has been shown to synthesize phosphatidylethanolamine from CDP-ethanolamine in vitro. The implications of the existence of such a pathway for the membrane flow hypothesis are discussed.Acknowledgments. This study was supported by the Medical Research Council of Canada.     

Comparative studies on the125I uptake of the thyroid and thymus

Zusammenfassung Es wird gezeigt, dass an erwachsenen und neugeborenen Ratten die Jodaufnahme des Thymus bei den letzteren viel höher ist als die Jodaufnahme der Thyroidea. Dies unterstützt die These, wonach die Jodspeicherkapazität der Thyroidea das Resultat einer Differenzierung darstellt, indem auf früherer ontogenetischer Stufe für den ganzen Vorderdarm eine Jodspeicherkapazität besteht.