The modulation of hela cells secretory patterns by invasive Shigella spp. and enteroinvasive E. coli bacterial cells and their soluble components

Considering the important role of cytokines in the initiation and evolution of the inflammatory process induced by Shigella and EIEC strains, the purpose of this study was the characterization of the secretory patterns of HeLa cells induced by Shigella ssp. and EIEC strains and to link the obtained results with the invasiveness level of bacterial strains on this cellular line. During this study there were analyzed two EIEC strains and 12 strains of the following Shigella species: 2 Sh. flexneri 2a, 2 Sh. flexneri 3a, 2 Sh. flexneri 4a, 2 Sh. boydii, 2 Sh. sonnei strains isolated in Romania during 2005 from children with dysentery and diarrhoea and confirmed for their invasive ability by Sereny test. The level of the main pro and anti-inflammatory cytokines, IL-1 beta, IL-6, IL-10, IL-13, IL-17 and TNF-alpha induced by whole bacterial cultures as well as by their soluble mediators was determined by an ELISA test. Our results showed that HeLa cells can be used not only for the qualitative and quantitative assessment of Shigella and EIEC strains invasion ability, but also as a simple work procedure for the investigation of an in vitro complex crosstalk communication mechanisms that involves physical interactions between bacterial cells and epithelial cells (adhesins and complementary receptors) and pro- and anti-inflammatory molecules regulation.The majority of the analyzed Shigella serogroups, with the exception of Shigella sonnei and EIEC strains, inhibited the inflammatory response by reducing the expression of majority of pro-inflammatory cytokines, i.e., IL-1 beta, IL-6, TNF-alpha and IL-17. The reduced cost of the in vitro procedure, the possibility of results interpretation and the strict regulations concerning the use of animals for experimental purposes are the main reasons that support the implementation of such an in vitro test in the research labs.     

Effects of aging and gender on the spatial organization of nuclei in single human skeletal muscle cells

The skeletal muscle fibre is a syncitium where each myonucleus regulates the gene products in a finite volume of the cytoplasm, i.e., the myonuclear domain (MND). We analysed aging‐ and gender‐related effects on myonuclei organization and the MND size in single muscle fibres from six young (21–31 years) and nine old men (72–96 years), and from six young (24–32 years) and nine old women (65–96 years), using a novel image analysis algorithm applied to confocal images. Muscle fibres were classified according to myosin heavy chain (MyHC) isoform expression. Our image analysis algorithm was effective in determining the spatial organization of myonuclei and the distribution of individual MNDs along the single fibre segments. Significant linear relations were observed between MND size and fibre size, irrespective age, gender and MyHC isoform expression. The spatial organization of individual myonuclei, calculated as the distribution of nearest neighbour distances in 3D, and MND size were affected in old age, but changes were dependent on MyHC isoform expression. In type I muscle fibres, average NN‐values were lower and showed an increased variability in old age, reflecting an aggregation of myonuclei in old age. Average MND size did not change in old age, but there was an increased MND size variability. In type IIa fibres, average NN‐values and MND sizes were lower in old age, reflecting the smaller size of these muscle fibres in old age. It is suggested that these changes have a significant impact on protein synthesis and degradation during the aging process.     

The nuclear basket proteins Mlp1p and Mlp2p are part of a dynamic interactome including Esc1p and the proteasome

The basket of the nuclear pore complex (NPC) is generally depicted as a discrete structure of eight protein filaments that protrude into the nucleoplasm and converge in a ring distal to the NPC. We show that the yeast proteins Mlp1p and Mlp2p are necessary components of the nuclear basket and that they also embed the NPC within a dynamic protein network, whose extended interactome includes the spindle organizer, silencing factors, the proteasome, and key components of messenger ribonucleoproteins (mRNPs). Ultrastructural observations indicate that the basket reduces chromatin crowding around the central transporter of the NPC and might function as a docking site for mRNP during nuclear export. In addition, we show that the Mlps contribute to NPC positioning, nuclear stability, and nuclear envelope morphology. Our results suggest that the Mlps are multifunctional proteins linking the nuclear transport channel to multiple macromolecular complexes involved in the regulation of gene expression and chromatin maintenance.     

Lattice Boltzmann simulations of the time evolution of living multicellular systems

Embryonic tissues and multicellular aggregates of adult cells mimic the behavior of highly viscous liquids. The liquid analogy helps to understand morphogenetic phenomena, such as cell sorting and tissue fusion, observed in developmental biology and tissue engineering. Tissue fusion is vital in tissue printing, an emergent technique based on computer-controlled deposition of tissue fragments and biocompatible materials. Computer simulations proved useful in predicting post-printing shape changes of tissue constructs. The simulation methods available to date, however, are unable to describe the time evolution of living systems made of millions of cells. The Lattice Boltzmann (LB) approach allows the implementation of interaction forces between the constituents of the system and yields time evolution in terms of distribution functions. With tissue engineering applications in mind, we have developed a finite difference Lattice Boltzmann model of a multicellular system and applied it to simulate the sidewise fusion of two contiguous cylinders made of cohesive cells and embedded in a medium (hydrogel). We have identified a biologically relevant range of model parameters. The proposed LB model may be extended to describe the time evolution of more complex multicellular structures such as sheets or tubes produced by tissue printing.     

Immunological aspects in viral hepatitis B and C infection

Worldwide, viral hepatitis chronic infections are a serious health problem and a very interesting topic for both clinicians and researchers. Viral hepatitis has a variety of clinical forms: mild, inactive or severe and with a slow evolution, whose architectural structure of the hepatic tissue evolves towards cirrhosis or hepatocellular carcinoma. Sometimes, the virally induced hepatic injury evolves spectacularly and rapidly leads to exitus. The factors that generate this evolution pattern depend on the immune response of the host and equally on the viral survival and immune surveillance avoidance strategies. This paper aims to resume new discoveries in the field of immunology of the B and C viral hepatitis infection, from the perspective of the complex interactions between virus and host.     

Romanian experience in child celiac disease diagnosis

Last consensus in celiac disease in 2008 conducted under the aegis of the European Society of Pediatric Gastroenterology, Hepatology and Nutrition jointly with North American Society of Pediatric Gastroenterology, Hepatology and Nutrition reveals the following: "celiac disease is a chronic immune-mediated enteropathy characterized by sensitization to gluten. That can affect any organ or system, with a wide range of clinical manifestations of variable severity". Thus, in recent years, clinical picture of celiac disease has changed the old paradigm--bowel disease with villous atrophy and malnutrition, being replaced with the new paradigm--multi-organ autoimmune disease, affecting many organs and systems throughout but with more less specific symptoms, which undiagnosed leads to delayed diagnosis, at a late-onset disease and long-term major complications as the risk of cancer. According to this consensus "the serological diagnosis of celiac disease is based on high sensitivity and specificity tests", but in line with changing clinical features of celiac disease, its diagnosis has undergone significant changes in recent years. These changes in the diagnosis of celiac disease, we have decided to analyze them.     

Sulforaphane Preconditioning Sensitizes Human Colon Cancer Cells towards the Bioreductive Anticancer Prodrug PR-104A

The chemoprotective properties of sulforaphane (SF), derived from cruciferous vegetables, are widely acknowledged to arise from its potent induction of xenobiotic-metabolizing and antioxidant enzymes. However, much less is known about the impact of SF on the efficacy of cancer therapy through the modulation of drug-metabolizing enzymes. To identify proteins modulated by a low concentration of SF, we treated HT29 colon cancer cells with 2.5 μM SF. Protein abundance changes were detected by stable isotope labeling of amino acids in cell culture. Among 18 proteins found to be significantly up-regulated, aldo-keto reductase 1C3 (AKR1C3), bioactivating the DNA cross-linking prodrug PR-104A, was further characterized. Preconditioning HT29 cells with SF reduced the EC₅₀ of PR-104A 3.6-fold. The increase in PR-104A cytotoxicity was linked to AKR1C3 abundance and activity, both induced by SF in a dose-dependent manner. This effect was reproducible in a second colon cancer cell line, SW620, but not in other colon cancer cell lines where AKR1C3 abundance and activity were absent or barely detectable and could not be induced by SF. Interestingly, SF had no significant influence on PR-104A cytotoxicity in non-cancerous, immortalized human colonic epithelial cell lines expressing either low or high levels of AKR1C3. In conclusion, the enhanced response of PR-104A after preconditioning with SF was apparent only in cancer cells provided that AKR1C3 is expressed, while its expression in non-cancerous cells did not elicit such a response. Therefore, a subset of cancers may be susceptible to combined food-derived component and prodrug treatments with no harm to normal tissues.