Urogastrone-epidermal growth factor is trophic to the intestinal epithelium of parenterally fed rats

The weight of the stomach, small intestine and colon and the mucosal crypt cell production rate of these tissues were significantly decreased after 10 days on an isocaloric TPN diet when compared to orally fed controls. Continuous infusion of recombinant beta urogastrone, at a dose below that needed to inhibit gastric acid secretion, largely prevented this decrease in crypt cell production rate and gastrointestinal tissue weights.     

Amyloid β-degrading cryptidases: insulin degrading enzyme, presequence peptidase, and neprilysin

The accumulation of aggregates of amyloidogenic peptides is associated with numerous human diseases. One well studied example is the association between deposition of amyloid beta (Abeta) and Alzheimer's disease. Insulin degrading enzyme and neprilysin are involved in the clearance of Abeta, and presequence peptidase is suggested to play a role in the degradation of mitochondrial Abeta. Recent structural analyses reveal that these three peptidases contain a catalytic chamber (crypt) that selectively encapsulates and cleaves amyloidogenic peptides, hence the name cryptidase. The substrate selectivity of these cryptidases is determined by the size and charge distribution of their crypt as well as the conformational flexibility of substrates. The interaction of Abeta with the catalytic core of these cryptidases is controlled by conformational changes that make the catalytic chambers accessible for Abeta binding. These new structural and biochemical insights into cryptidases provide potential therapeutic strategies for the control of Abeta clearance.     

Effect of the type of diet on the distribution of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase in rat small intestine

Summary The jejunoileal gradient for the HMG-CoA reductase activity in the microsomal fraction of the epithelial cells of the small intestine of rats given commercial pellets was reserved within a few days after changing the ration to a semipurified diet. The response of the reductase was essentially the same in villus and crypt cells.     

Changes in brush-border enzyme activities of intestinal epithelial cells isolated from the villus-crypt axis during the early phase of alloxan diabetes in rats

Summary The sucrase activity in enterocytes isolated from the villus crypt axis was found to increase in all regions of the villus from day 2 after induction of diabetes, and the increase continued until day 4. In contrast, alkaline phosphatase activity increased mainly in the apical one-third of the villus-crypt column, and the increase occurred abruptly on day 4 with increase in food intake.     

Changes in brush-border enzyme activities of intestinal epithelial cells isolated from the villus-crypt axis during the early phase of alloxan diabetes in rats

The sucrase activity in enterocytes isolated from the villus crypt axis was found to increase in all regions of the villus from day 2 after induction of diabetes, and the increase continued until day 4. In contrast, alkaline phosphatase activity increased mainly in the apical one-third of the villus-crypt column, and the increase occurred abruptly on day 4 with increase in food intake.     

Calmodulin in epithelial intestinal cells during rat development

Summary Calmodulin was immunocytochemically localized in the brush borders of rat intestinal epithelial cells from the tip to the base of the villi, from day 18 of fetal life up to the adult stage. The early (14th day) fetal cells, like the adult crypt cells, were not immunoreactive, although their calmodulin content was equal to that of the mature cells from the tips of the villi.     

Calmodulin in epithelial intestinal cells during rat development

Calmodulin was immunocytochemically localized in the brush borders of rat intestinal epithelial cells from the tip to the base of the villi, from day 18 of fetal life up to the adult stage. The early (14th day) fetal cells, like the adult crypt cells, were not immunoreactive, although their calmodulin content was equal to that of the mature cells from the tips of the villi.     

Beta-catenin/TCF4 transactivates miR-30e during intestinal cell differentiation

The Wnt/beta-catenin/TCF4 pathway plays critical roles in the maintenance of small intestinal epithelium; however, downstream targets of the beta-catenin/TCF4 complex are not extensively characterized. We identified miR-30e as an immediate target activated by the beta-catenin/TCF4 complex. miR-30e was detected in the peri-nuclear region of the intestinal crypt IEC-6 cells. Bioinformatics analysis revealed clustered beta-catenin/TCF4 binding sites within the miR-30e promoter region. This promoter region was cloned into pGL3-control luciferase reporter vector, with the enhancer region removed. Transfection of pCMV-SPORT6-beta-catenin expression vector dose-dependently increased luciferase activity, and co-transfection of pCMV-SPORT6-TCF4 expression vector further enhanced the promoter activity. Dexamethasone-induced IEC-6 cells differentiation caused a 2.5-fold increase in miR-30e expression, and upon beta-catenin siRNA transfection, miR-30e increased 1.3-fold. Electrophoretic mobility shift assay and chromatin immunoprecipitation assay confirmed the binding between beta-catenin/TCF4 complexes from IEC-6 nuclear extracts and the putative sequences in the miR-30e promoter. These results demonstrate that beta-catenin/TCF4 transactivates miR-30e during intestinal cell differentiation.     

Metabolic engineering of Saccharomyces cerevisiae: a key cell factory platform for future biorefineries

Metabolic engineering is the enabling science of development of efficient cell factories for the production of fuels, chemicals, pharmaceuticals, and food ingredients through microbial fermentations. The yeast Saccharomyces cerevisiae is a key cell factory already used for the production of a wide range of industrial products, and here we review ongoing work, particularly in industry, on using this organism for the production of butanol, which can be used as biofuel, and isoprenoids, which can find a wide range of applications including as pharmaceuticals and as biodiesel. We also look into how engineering of yeast can lead to improved uptake of sugars that are present in biomass hydrolyzates, and hereby allow for utilization of biomass as feedstock in the production of fuels and chemicals employing S. cerevisiae. Finally, we discuss the perspectives of how technologies from systems biology and synthetic biology can be used to advance metabolic engineering of yeast.     

A quick and efficient method to generate mammalian stable cell lines based on a novel inducible alphavirus DNA/RNA layered system

We report a new method to generate high-expressing mammalian cell lines in a quick and efficient way. For that purpose, we developed a master cell line (MCL) containing an inducible alphavirus vector expressing GFP integrated into the genome. In the MCL, recombinant RNA levels increased >4,600-fold after induction, due to a doxycycline-dependent RNA amplification loop. The MCL maintained inducibility and expression during 50 passages, being more efficient for protein expression than a conventional cell line. To generate new cell lines, mutant LoxP sites were inserted into the MCL, allowing transgene and selection gene exchange by Cre-directed recombination, leading to quick generation of inducible cell lines expressing proteins of therapeutic interest, like human cardiotrophin-1 and oncostatin-M at several mg/l/24 h. These proteins contained posttranslational modifications, showed bioactivity, and were efficiently purified. Remarkably, this system allowed production of toxic proteins, like oncostatin-M, since cells able to express it could be grown to the desired amount before induction. These cell lines were easily adapted to growth in suspension, making this methodology very attractive for therapeutic protein production.     

Metabolic changes during B cell differentiation for the production of intestinal IgA antibody

To sustain the bio-energetic demands of growth, proliferation, and effector functions, the metabolism of immune cells changes dramatically in response to immunologic stimuli. In this review, I focus on B cell metabolism, especially regarding the production of intestinal IgA antibody. Accumulating evidence has implicated not only host-derived factors (e.g., cytokines) but also gut environmental factors, including the possible involvement of commensal bacteria and diet, in the control of B cell metabolism during intestinal IgA antibody production. These findings yield new insights into the regulation of immunosurveillance and homeostasis in the gut.     

Protecting the boundary: the sentinel role of host defense peptides in the skin

The skin is our primary shield against microbial pathogens and has evolved innate and adaptive strategies to enhance immunity in response to injury or microbial insult. The study of antimicrobial peptide (AMP) production in mammalian skin has revealed several of the elegant strategies that AMPs use to prevent infection. AMPs are inducible by both infection and injury and protect the host by directly killing pathogens and/or acting as multifunctional effector molecules that trigger cellular responses to aid in the anti-infective and repair response. Depending on the specific AMP, these molecules can influence cytokine production, cell migration, cell proliferation, differentiation, angiogenesis and wound healing. Abnormal production of AMPs has been associated with the pathogenesis of several cutaneous diseases and plays a role in determining a patient’s susceptibility to pathogens. This review will discuss current research on the regulation and function of AMPs in the skin and in skin disorders.     

Blocking of melatonin synthesis and MT1 receptor impairs the activation of Jurkat T cells

Melatonin has been proposed as regulating the immune system by affecting cytokine production in immunocompetent cells, enhancing the production of several T helper (Th)1 cytokines. To further investigate the melatonin’s role in IL-2/IL-2R system, we established an inducible T-REx expression system in Jurkat cells in which the protein levels of HIOMT enzyme or MT₁ receptor were significantly down-regulated upon tetracycline incubation. We found that T-REx Jurkat cells with lower levels of HIOMT activity, and consequently lower content of endogenous melatonin, showed IL-2 production decrease after activation with lectin. Likewise, tetracycline-inducible stable cell line expressing MT₁ antisense produced decreased amounts of IL-2 (mRNA and protein levels) after stimulation. Moreover, in T-Rex-MT₁ cells incubated with tetracycline, a sub-optimal PHA dose failed to induce the early activation marker CD25 on the cell surface. The results shown here support the relevance of endogenous melatonin and its signaling in T cell activation.     

Molecular basis of parathyroid hormone receptor signaling and trafficking: a family B GPCR paradigm

The parathyroid hormone (PTH) receptor type 1 (PTHR), a G protein-coupled receptor (GPCR), transmits signals to two hormone systems—PTH, endocrine and homeostatic, and PTH-related peptide (PTHrP), paracrine—to regulate different biological processes. PTHR responds to these hormonal stimuli by activating heterotrimeric G proteins, such as G_S that stimulates cAMP production. It was thought that the PTHR, as for all other GPCRs, is only active and signals through G proteins on the cell membrane, and internalizes into a cell to be desensitized and eventually degraded or recycled. Recent studies with cultured cell and animal models reveal a new pathway that involves sustained cAMP signaling from intracellular domains. Not only do these studies challenge the paradigm that cAMP production triggered by activated GPCRs originates exclusively at the cell membrane but they also advance a comprehensive model to account for the functional differences between PTH and PTHrP acting through the same receptor.     

DNA-dependent-DNA-polymerase: possible limiting influence on cell reproduction during viral leukemogenesis

Evidence is presented that during viral leukemogenesis spleen cell nuclei show an increase in labelling index and mean grain count, that is not accompanied by any changes in the nuclear level of DNA-polymerase-alpha. It is suggested that polymerase production remains under the control of the normal cell mechanisms and the virus may affect cell proliferation by altering the primer-template levels.     

The effect of mouse erythrocyte rosette forming lymphocytes on lymphokine production in T-cell cultures

An enhancement effect of mouse erythrocyte rosette forming (MERF) cells on the production of migration inhibitory factor, chemotactic factor for neutrophils and skin reactive factor in T-lymphocyte cultures stimulated with the purified protein derivative of tuberculin was observed. We consider it likely that the MERF cells, possessing the appropriate cell surface constituents to construct an immunogenic moiety, present antigen on their surfaces to elicit lymphokine production.