Decrease of mRNA levels and biosynthesis of sucrase-isomaltase but not dipeptidylpeptidase IV in forskolin or monensin-treated Caco-2 cells

Treatment for 48 h of differentiated, confluent Caco-2 cells with 2.5 10^?5 M forskolin or 10^?6 M monensin, which produces a significant decrease of the de novo biosynthesis of sucrase-isomaltase, does not change quantitatively the de novo biosynthesis of dipeptidylpeptidase IV. Western blot analysis and silver nitrate staining indicate that neither drug induces any modification in the steady state expression of these two brush border hydrolases. Northern blot analysis shows that the level of dipeptidylpeptidase IV mRNA does not change in treated as compared to control Caco-2 cells. In contrast, forskolin and monensin dramatically decrease the level of sucrase-isomaltase mRNA. These observations suggest a separate regulation of biosynthesis for sucrase-isomaltase and dipeptidylpeptidase IV in intestinal cells. The mechanisms responsible for such a difference are discussed. Among them, the role of glucose metabolism, which is perturbed by both drugs, appears to be of crucial importance.     

Histochemical evidence for the presence of dipeptidylpeptidase IV in the Schwann cells of skin uymyelinated axons

Summary DPP IV activity was localized in the nerve fascicles of cat glabrous skin at light and electron microscope levels. The observation that the DPP IV end product was restricted to the axon-Schwann cell interface suggests that this enzyme may be involved in the interactions between unmyelinated axons and their Schwann cells.17 November 1986     

Histochemical evidence for the presence of dipeptidylpeptidase IV in the Schwann cells of skin unmyelinated axons

DPP IV activity was localized in the nerve fascicles of cat glabrous skin at light and electron microscope levels. The observation that the DPP IV end product was restricted to the axon-Schwann cell interface suggests that this enzyme may be involved in the interactions between unmyelinated axons and their Schwann cells.     

Decrease of cholesterol and free fatty acids in cortisone-resistant lymphoid cells incubated with allogeneic tumor cells.

A marked decrease of cholesterol and free fatty acids was found in the cortisone-resistant lymphoid cells from thymus or spleen of mice immunized with Ehrlich carcinoma cells when incubated with the tumor cells.     

Activation-induced cellular accumulation of histamine in immature but not mature murine mast cells

Mast cell activation involves the rapid release of inflammatory mediators, including histamine, from intracellular granules. The cells are capable of regranulation and multiple rounds of activation. The goal of this study was to determine if there are changes in the content of pre-formed mast cell mediators after a round of activation. After 24 h, the histamine content of bone marrow-derived mast cells (BMMC), but not that of peritoneal mast cells, exceeded the amount in resting cells. Accumulation of histamine in BMMC peaked at 72 h of activation, and returned toward preactivation levels by 96 h. The increase in histamine content was accompanied by an increase in the gene expression of histidine decarboxylase. No increases in beta hexosaminidase or murine mast cell protease-6 were observed. These findings indicate that BMMC respond to activation by increasing total cell-associated histamine content. This increase may be important to the response of these cells upon subsequent exposure to antigens.     

Glycoprotein biosynthesis in splenic cells. Purification of a microsomal galactosyl-transferase.

One part of the microsomal galactosyl-transferase activity of splenic cells of rats can by solubilized by the action of Triton X-100 and Tween 20. Its purification on a Sephadex G-200 column and by electrophoresis on a polyacrylamide gel leads to a solution of high specific enzymic activity.     

Glycoprotein biosynthesis in splenic cells. Purification of a microsomal galactosyl-transferase

Summary One part of the microsomal galactosyl-transferase activity of splenic cells of rats can by solubilized by the action of Triton X-100 and Tween 20. Its purification on a Sephadex G-200 column and by electrophoresis on a polyacrylamide gel leads to a solution of high specific enzymic activity.This work has benefited from the help of the Centre National de la Recherche Scientifique, the Direction des Recherches et Moyens d'Essais, the Institut National de la Santé et de la Recherche Médicale, the Fondation pour la Recherche Médicale Française the Délégation Générale à la Recherche Scientifique et Technique and the Université de Lyon (UER Lyon-Sud et Biologie Humaine).     

ERK activation by Ca2+ ionophores depends on Ca2+ entry in lymphocytes but not in platelets, and does not conduct membrane scrambling

Rapid Ca²⁺-dependent phospholipid (PL) reorganization (scrambling) at the plasma membrane is a mechanism common to hematopoietic cells exposing procoagulant phosphatidylserine (PS). The aim of this research was to determine whether activation of the extracellular signal-regulated kinase (ERK) pathway was required for PL scrambling, based on a single report analyzing both responses induced by Ca²⁺ ionophores in megakaryoblastic HEL cells. Ca²⁺ ionophore-stimulated ERK phosphorylation was induced in platelets without external Ca²⁺, whereas exogenous Ca²⁺ entry was crucial for ERK activation in Jurkat T cells. In both cells, membrane scrambling only occurred following Ca²⁺ entry and was not blocked by inhibiting ERK phosphorylation. Furthermore, ERK proteins are strongly phosphorylated in transformed B lymphoblastic cell lines, which do not expose PS in their resting state. Overall, the data demonstrated that ERK activation and membrane scrambling are independent mechanisms. A. Arachiche, I. Badirou: These authors contributed equally to this work. Received 18 June 2008; received after revision 24 September 2008; accepted 1 October 2008     

Decrease of cholesterol and free fatty acids in cortisone-resistant lymphoid cells incubated with allogeneic tumor cells

Summary A marked decrease of cholesterol and free fatty acids was found in the cortisone-resistant lymphoid cells from thymus or spleen of mice immunized with Ehrlich carcinoma cells when incubated with the tumor cells.     

Morphine but not fentanyl and methadone affects mitochondrial membrane potential by inducing nitric oxide release in glioma cells

We have observed that treatment of human glioma cells with morphine in the nanomolar range of concentration affects the mitochondrial membrane potential. The effect is specific to morphine and is mediated by naloxone-sensitive receptors, and is thus better observed on glioma cells treated with desipramine; moreover, the mitochondrial impairment is not inducible by fentanyl or methadone treatment and is prevented by the nitric oxide (NO) synthase inhibitor L-NAME. We conclude that in cultured glioma cells, the morphine-induced NO release decreases the mitochondrial membrane potential, as one might expect based on the rapid inhibition of the respiratory chain by NO. The identification of new intra-cellular pathways involved in the mechanism of action of morphine opens additional hypotheses, providing a novel rationale relevant to the therapy and toxicology of opioids.Received 19 August 2004; received after revision 16 September 2004; accepted 7 October 2004     

Silencing of RB1 but not of RB2/P130 induces cellular senescence and impairs the differentiation potential of human mesenchymal stem cells

Stem cell senescence is considered deleterious because it may impair tissue renewal and function. On the other hand, senescence may arrest the uncontrolled growth of transformed stem cells and protect organisms from cancer. This double function of senescence is strictly linked to the activity of genes that the control cell cycle such as the retinoblastoma proteins RB1, RB2/P130, and P107. We took advantage of the RNA interference technique to analyze the role of these proteins in the biology of mesenchymal stem cells (MSC). Cells lacking RB1 were prone to DNA damage. They showed elevated levels of p53 and p21^cip1 and increased regulation of RB2/P130 and P107 expression. These cells gradually adopted a senescent phenotype with impairment of self-renewal properties. No significant modification of cell growth was observed as it occurs in other cell types or systems. In cells with silenced RB2/P130, we detected a reduction of DNA damage along with a higher proliferation rate, an increase in clonogenic ability, and the diminution of apoptosis and senescence. Cells with silenced RB2/P130 were cultivated for extended periods of time without adopting a transformed phenotype. Of note, acute lowering of P107 did not induce relevant changes in the in vitro behavior of MSC. We also analyzed cell commitment and the osteo-chondro-adipogenic differentiation process of clones derived by MSC cultures. In all clones obtained from cells with silenced retinoblastoma genes, we observed a reduction in the ability to differentiate compared with the control clones. In summary, our data show evidence that the silencing of the expression of RB1 or RB2/P130 is not compensated by other gene family members, and this profoundly affects MSC functions.     

High levels of free fatty acids and their esters in lymphoid cells resistant to cortisone or cyclophosphamide

Summary The lymphoid cells from thymus, slpeen or mesenteric lymph node of mice treated with hydrocortisone or cyclophosphamide contained the significantly high levels of free fatty acids, triglycerides and cholesterol esters as compared to the corresponding cells from untreated animals.     

The phosphoinositide PI(3,5)P2 mediates activation of mammalian but not plant TPC proteins: functional expression of endolysosomal channels in yeast and plant cells

Two-pore channel proteins (TPC) encode intracellular ion channels in both animals and plants. In mammalian cells, the two isoforms (TPC1 and TPC2) localize to the endo-lysosomal compartment, whereas the plant TPC1 protein is targeted to the membrane surrounding the large lytic vacuole. Although it is well established that plant TPC1 channels activate in a voltage- and calcium-dependent manner in vitro, there is still debate on their activation under physiological conditions. Likewise, the mode of animal TPC activation is heavily disputed between two camps favoring as activator either nicotinic acid adenine dinucleotide phosphate (NAADP) or the phosphoinositide PI(3,5)P₂. Here, we investigated TPC current responses to either of these second messengers by whole-vacuole patch-clamp experiments on isolated vacuoles of Arabidopsis thaliana. After expression in mesophyll protoplasts from Arabidopsis tpc1 knock-out plants, we detected the Arabidopsis TPC1-EGFP and human TPC2-EGFP fusion proteins at the membrane of the large central vacuole. Bath (cytosolic) application of either NAADP or PI(3,5)P₂ did not affect the voltage- and calcium-dependent characteristics of AtTPC1-EGFP. By contrast, PI(3,5)P₂ elicited large sodium currents in hTPC2-EGFP-containing vacuoles, while NAADP had no such effect. Analogous results were obtained when PI(3,5)P₂ was applied to hTPC2 expressed in baker’s yeast giant vacuoles. Our results underscore the fundamental differences in the mode of current activation and ion selectivity between animal and plant TPC proteins and corroborate the PI(3,5)P₂-mediated activation and Na⁺ selectivity of mammalian TPC2.     

Decrease of mast cells in regional lymph nodes in response to allogeneic antigens and syngeneic tumor antigens

Summary The absolute number of mast cells in regional lymph nodes decreases on the 5th day after stimulation by allogeneic lymphocytes and semisyngeneic leukaemic cells, despite the enlargement of stimulated lymph nodes. It is postulated that the reaction of lymphatic mast cells could be a sensitive test for tissue incompatibility, and probably also for the presence of tumor associated antigens.     

Localization of ceruloplasmin biosynthesis in human and monkey liver cells and its copper regulation

Zusammenfassung Aus Affenserum (Macacus rhesus) ist das Zeruloplasmin in Form einer homogenen Eiweisssubstanz isoliert und in ihren physikalisch-chemischen Konstanten näher bestimmt worden. Der Vergleich der Eigenschaften des Zeruloplasmins des Menschen und des Affen ergibt eine Ähnlichkeit dieser beiden Substanzen. Bei der Untersuchung mittels spezifischer Präzipitation im Agar haben sich beim Antiserum des Kaninchens Beweisgründe für eine antigene Identität beider Zeruloplasmine ergeben. An Gewebeschnitten der Affenleber inkubiert bei aktiv oxydativer Phosphorylierung ergab sich die Bindung des Zeruloplasmins in vitro. Resynthetisierte Eiweißsubstanz bildet einen spezifischen Niederschlag mit dem Antiserum des Kaninchens, welches gegen das Zeruloplasmin des Menschen immunisiert ist. Mit Hilfe lumineszierender Antikörper ist die spezifische Lumineszenz des Zeruloplasmins nur in parenchimatösen Leberzellen gefunden worden. Die Biosynthese des Zeruloplasmins ist in der Leber lokalisiert. Aus In-vitro-Versuchen an Gewebeschnitten der Leber beim Affen und entsprechenden In-vivo-Versuchen ergibt sich, dass Kupfersalze in geringen Konzentrationen die Biosynthese verstärken, in grossen Konzentrationen hingegen die Biosynthese des Zeruloplasmins hemmen.     

Change in levels of cholesterol and free fatty acids of lymphoid cells during tumor growth.

Growth of Ehrlich's ascitic carcinoma in mice resulted in increase of free cholesterol and free fatty acids in lymphoid cells from thymus, spleen and cervical lymph node, but decrease of these lipids in the cells from mesenteric lymph node.