Impaired development of T lymphoid precursors from pluripotent hematopoietic stem cells in New Zealand Black mice.

Bone marrow cells from autoimmune-prone New Zealand Black (NZB) mice are less efficient at colonizing fetal thymic lobes than cells from normal strains. This study demonstrates that the reduced capacity of NZB bone marrow cells to repopulate the thymus does not result from their inability to migrate to or enter the thymus. Rather, the T lymphopoietic defect of NZB mice is due to an impaired ability of pluripotent hematopoietic stem cells (PHSCs) to generate more committed lymphoid progeny, which could include common lymphoid precursors and/or other T cell-committed progenitors. Although PHSCs from NZB mice were not as efficient at thymic repopulation as comparable numbers of PHSCs from control strains, the ability of common lymphoid precursors from NZB mice to repopulate the thymus was not defective. Similarly, more differentiated NZB T cell precursors included in the intrathymic pool of CD4(-)CD8(-) cells also exhibited normal T lymphopoietic potential. Taken together, the results identify an unappreciated defect in NZB mice and provide further evidence that generation of lymphoid progeny from the PHSCs is a regulated event.     

Raising the list of SirT7 targets to a new level

Sirtuins are crucial proteins involved in sensing and coordinating the response to different forms of stress, mainly through NAD⁺‐dependent deacetylation of proteins. For that reason, sirtuins are directly involved in many human pathologies including cancer, diabetes, cardiovascular and neurodegenerative diseases. SirT7, one of the less well‐understood sirtuins, has been associated with ribosome biogenesis, gene expression, metabolism and cancer. Despite the wide range of these functions, only a handful of targets for SirT7 have so far been described. In this issue, Zhang et al. report the first proteomic screening of SirT7 substrates. Using stable isotope labeling with amino acids in cell culture (SILAC), coupled with quantitative mass spectrometry, they have identified a comprehensive list of candidates involved in a variety of functions, ranging from maintenance of chromatin architecture to gene silencing and metabolism. A selected group of these candidates has been validated by in vitro co‐immunoprecipitation and deacetylation experiments. Predictive tools have identified additional candidates. The identification of these novel targets not only suggests new ways of understanding the physiological role of SirT7, but also provides new evidence to add to our existing knowledge of the global impact of sirtuins in cell homeostasis.     

Loss of SIRT2 leads to axonal degeneration and locomotor disability associated with redox and energy imbalance

Sirtuin 2 (SIRT2) is a member of a family of NAD⁺‐dependent histone deacetylases (HDAC) that play diverse roles in cellular metabolism and especially for aging process. SIRT2 is located in the nucleus, cytoplasm, and mitochondria, is highly expressed in the central nervous system (CNS), and has been reported to regulate a variety of processes including oxidative stress, genome integrity, and myelination. However, little is known about the role of SIRT2 in the nervous system specifically during aging. Here, we show that middle‐aged, 13‐month‐old mice lacking SIRT2 exhibit locomotor dysfunction due to axonal degeneration, which was not present in young SIRT2 mice. In addition, these Sirt2^?/? mice exhibit mitochondrial depletion resulting in energy failure, and redox dyshomeostasis. Our results provide a novel link between SIRT2 and physiological aging impacting the axonal compartment of the central nervous system, while supporting a major role for SIRT2 in orchestrating its metabolic regulation. This underscores the value of SIRT2 as a therapeutic target in the most prevalent neurodegenerative diseases that undergo with axonal degeneration associated with redox and energetic dyshomeostasis.     

Quantitative electroencephalography reveals different physiological profiles between benign and remitting-relapsing multiple sclerosis patients

Background  

A possible method of finding physiological markers of multiple sclerosis (MS) is the application of EEG quantification (QEEG) of brain activity when the subject is stressed by the demands of a cognitive task. In particular, modulations of the spectral content that take place in the EEG of patients with multiple sclerosis remitting-relapsing (RRMS) and benign multiple sclerosis (BMS) during a visuo-spatial task need to be observed.     

Flow cytometric analysis of cytokine production by normal human peripheral blood dendritic cells and monocytes: comparative analysis of different stimuli, secretion-blocking agents and incubation periods

In this paper, we comparatively analyze the effects of the following different stimuli on the production and intracellular accumulation of the interleukin (IL)-1 beta, IL-6, IL-12, tumor necrosis factor-alpha (TNF-alpha), and IL-8 inflammatory cytokines in both normal human peripheral blood (PB) dendritic cell (DC) subsets and monocytes: lipopolysaccharide (LPS) versus Staphylococcus aureus cowan I (SAC) in the presence or absence of interferon-(IFN)-gamma-, cytokine secretion-blocking agents (brefeldin A alone versus brefeldin A plus monensin), and incubation periods (6, 12, and 24 h). For this purpose, a four-color multiple-staining direct immunofluorescence technique analyzed by flow cytometry was systematically used in all experiments (n = 19). Our results show that after stimulation, an important proportion of each of the two CD33(+) myeloid DC subsets as well as the monocytes produce significant amounts of all cytokines analyzed under each of the experimental conditions assayed. In contrast, CD33(-/+lo) lymphoplasmocytoid DC failed to produce detectable levels of any of the above-mentioned cytokines under the same stimulatory conditions. Upon comparing the different stimuli used, LPS was associated with higher percentages of cytokine-producing cells compared with SAC, especially within the CD33(hi) DC subset; interestingly, the addition of IFN-gamma enhanced the response of monocytes to both LPS and SAC. As regards the secretion-blocking agents, brefeldin A alone was superior to the combination of brefeldin A and monensin. This is because it was frequently associated with both a higher percentage of cytokine-positive cells and greater amounts of detectable cytokines per cell. Sequential analysis of cytokine production by PB DC and monocytes after 6, 12, and 24 h of cell culture showed that after 6 h, an increased cell death rate existed among DC, which became even undetectable at 24 h, in the absence of a significant increase in cytokine secretion. In summary, our results show that from the experimental conditions assayed in this paper, to induce cytokine production by normal human DC and monocytes, maximum response is obtained once PB samples are stimulated for 6 h with LPS (with or without IFN-gamma) in the presence of brefeldin A alone.     

Interchromatin granules in plant nuclei

The interchromatin granules (IG) are well characterized subnuclear structures in animal cell nuclei which form a part of the internal nuclear matrix and are supposed to correspond to accummulation sites of snRNPs and/or maturation and transport of rRNPs; but similar structures have not yet been characterized in plant nuclei. Using the bismuth impregnation technique of Locke and Huie, preferential for the staining of IG, we describe here a kind of positive granules 20–25 nm in diameter in plants. The putative plant IG have a series of common characteristics with animal IG, which are not found as a whole in any other subnuclear structures. These are: size, morphology and ultrastructural organization, presence in groups, EDTA and bismuth oxynitrate positive reactions, resistance to double digestion with proteases and RNase I and their high content in phosphorylated proteins. For all the above and also because they are ubiquitous structures in the plant nuclei, we identified these granules with the IG described in animal cells.     

Immunomodulation by Yersinia enterocolitica: comparison of live and heat-killed bacteria

This study compared the immunomodulating properties of viable and killed Yersinia enterocolitica O9 in BALB/c mice. At 10 days after infection by the intragastric route, ex vivo assays showed a suppression of spleen cell proliferation in response to Salmonella lipopolysaccharide, concanavalin A and heat-killed yersiniae. Mice infected with Y. enterocolitica O9 for 10 days resisted the challenge with a lethal dose of Listeria monocytogenes. In contrast, intravenous administration of heat-killed yersiniae did not modify the ability of spleen cells to proliferate in response to lipopolysaccharide or concanavalin A, and proliferation in response to killed yersiniae was significantly increased. By 3 days after administration of a single dose of heat-killed yersiniae, the resistance of mice to L. monocytogenes challenge was significantly increased. Our findings show profound differences in immunomodulation by viable and heat-killed yersiniae, but suggest that killed yersiniae retain interesting immunomodulating properties.     

Cereal crops as viable production and storage systems for pharmaceutical scFv antibodies

This report describes the stable expression of a medically important antibody in the staple cereal crops rice and wheat. We successfully expressed a single-chain Fv antibody (ScFvT84.66) against carcinoembryonic antigen (CEA), a well characterized tumor-associated marker antigen. scFv constructs were engineered for recombinant antibody targeting to the plant cell apoplast and ER. Up to 30 g/g of functional recombinant antibody was detected in the leaves and seeds of wheat and rice. We confirmed that transgenic dry seeds could be stored for at least five months at room temperature, without significant loss of the amount or activity of scFvT84.66. Our results represent the first transition from model plant expression systems, such as tobacco and Arabidopsis, to widely cultivated cereal crops, such as rice and wheat, for expression of an antibody molecule that has already shown efficacy in clinical applications. Thus, we have established that molecular pharming in cereals can be a viable production system for such high-value pharmaceutical macromolecules. Our findings provide a strong foundation for exploiting alternative uses of cereal crops both in industrialized and developing countries.     

Tocotrienols: balancing the mitochondrial crosstalk between apoptosis and autophagy

Tocotrienols are a group of natural vitamin E compounds with patent antitumoral effects, mostly based on their ability to induce apoptosis in cancer cells. In activated pancreatic stellate cells (PSCs) we have determined that tocotrienols elicit a dramatic mitochondrial destabilization followed by initiation of non-necrotic forms of programmed cell death, namely apoptosis and autophagy. PSCs are the main cell type involved in the generation of pancreatic fibrosis, and their removal is critical to limit the fibrogenic process. Noteworthy, tocotrienol death-promoting actions are exclusively directed to activated PSCs, but not to their quiescent counterparts nor to terminally differentiated acinar cells. Here, we hypothesize that the transformed phenotype of PSCs may include "activated" mitochondria, which can be used by tocotrienols to trigger autophagic and apoptotic signaling. We propose that mitochondria are the cornerstone of cell sensitivity to tocotrienols, and suggest possible mechanisms, that may be interconnected, on how tocotrienols may govern mitochondrial death pathways.