Transforming growth factor-β1 (TGF-β1) is an important regulator of fibrogenesis in heart disease. In many other cellular systems, TGF-β1 may also induce autophagy, but a link between its fibrogenic and autophagic effects is unknown. Thus we tested whether or not TGF-β1-induced autophagy has a regulatory function on fibrosis in human atrial myofibroblasts (hATMyofbs). Primary hATMyofbs were treated with TGF-β1 to assess for fibrogenic and autophagic responses. Using immunoblotting, immunofluorescence and transmission electron microscopic analyses, we found that TGF-β1 promoted collagen type Iα2 and fibronectin synthesis in hATMyofbs and that this was paralleled by an increase in autophagic activation in these cells. Pharmacological inhibition of autophagy by bafilomycin-A1 and 3-methyladenine decreased the fibrotic response in hATMyofb cells. ATG7 knockdown in hATMyofbs and ATG5 knockout (mouse embryonic fibroblast) fibroblasts decreased the fibrotic effect of TGF-β1 in experimental versus control cells. Furthermore, using a coronary artery ligation model of myocardial infarction in rats, we observed increases in the levels of protein markers of fibrosis, autophagy and Smad2 phosphorylation in whole scar tissue lysates. Immunohistochemistry for LC3β indicated the localization of punctate LC3β with vimentin (a mesenchymal-derived cell marker), ED-A fibronectin and phosphorylated Smad2. These results support the hypothesis that TGF-β1-induced autophagy is required for the fibrogenic response in hATMyofbs.

Eukaryotic chromosomes are folded into hierarchical domains, forming functional compartments. Nuclear periphery and nucleolus are two nuclear landmarks contributing to repressive chromosome architecture. However, while the role of nuclear lamina (NL) in genome organization has been well documented, the function of the nucleolus remains under-investigated due to the lack of methods for the identification of nucleolar associated domains (NADs). Here we have established DamID- and HiC-based methodologies to generate accurate genome-wide maps of NADs in embryonic stem cells (ESCs) and neural progenitor cells (NPCs), revealing layers of genome compartmentalization with distinct, repressive chromatin states based on the interaction with the nucleolus, NL, or both. NADs show higher H3K9me2 and lower H3K27me3 content than regions exclusively interacting with NL. Upon ESC differentiation into NPCs, chromosomes around the nucleolus acquire a more compact, rigid architecture with neural genes moving away from nucleoli and becoming unlocked for later activation. Further, histone modifications and the interaction strength within A and B compartments of NADs and LADs in ESCs set the choice to associate with NL or nucleoli upon dissociation from their respective compartments during differentiation. The methodologies here developed will make possible to include the nucleolar contribution in nuclear space and genome function in diverse biological systems.

The nucleolus is the largest compartment of the eukaryotic cell's nucleus. It acts as a ribosome factory, thereby sustaining the translation machinery. The nucleolus is also the subnuclear compartment with the highest transcriptional activity in the cell, where hundreds of ribosomal RNA (rRNA) genes transcribe the overwhelming majority of RNAs. The structure and composition of the nucleolus change according to the developmental state. For instance, in embryonic stem cells (ESCs), rRNA genes display a hyperactive transcriptional state and open chromatin structure compared with differentiated cells. Increasing evidence indicates that the role of the nucleolus and rRNA genes might go beyond the control of ribosome biogenesis. One such role is linked to the genome architecture, since repressive domains are often located close to the nucleolus. This review highlights recent findings describing how the nucleolus is regulated in ESCs and its role in regulating ribosome biogenesis and genome organization for the maintenance of stem cell identity.

The antimicrobial effects of garlic, ginger, carrot and turmeric pastes against Escherichia coli O157:H7 in laboratory buffer and model food system were investigated. Turmeric paste, fresh carrot, ginger and garlic pastes from roots, and commercial ginger and garlic paste were heated alone or with buffered peptone water (BPW) or ground beef at 70 degrees C for 7 min. All samples were inoculated with a three strain cocktail of overnight cultures of E. coli O157: H7 and stored at 4 degrees C and 8 degrees C for 2 weeks. Each paste exhibited different antimicrobial effects alone and in ground beef or BPW at 4 degrees C and 8 degrees C for 2 weeks. Commercial ginger paste and fresh garlic paste showed the strongest antimicrobial activity with complete inactivation of E. coli O157:H7 in the paste at 3 days at 4 degrees C and 8 degrees C. Carrot and turmeric pastes did not show any antimicrobial activity both at 4 degrees C and 8 degrees C. Commercial garlic showed antimicrobial activity at both 4 degrees C and 8 degrees C (about 1 log CFU/g reduction) in the paste. However, fresh ginger paste showed antimicrobial activity only at 8 degrees C. Only commercial ginger paste had antimicrobial activity in BPW at 4 degrees C for 2 weeks. However, commercial ginger paste showed antimicrobial activity in ground beef at 3 days and after (about 1-2 log CFU/g) compared to control samples at 8 degrees C for 2 weeks. Fresh garlic paste showed antimicrobial activity only in BPW (1.3 log CFU/g) at 8 degrees C. These results indicate that the antimicrobial activity of these pastes is decreased in ground beef and laboratory buffer.