Functional Recellularization of Acellular Rat Liver Scaffold by Induced Pluripotent Stem Cells: Molecular Evidence for Wnt/B-Catenin Upregulation.

BACKGROUND: Liver transplantation remains the only viable therapy for liver failure but has a severely restricted utility. Here, we aimed to decellularize rat livers to form acellular 3D bio-scaffolds suitable for seeding with induced pluripotent cells (iPSCs) as a tool to investigate the role of Wnt/β-catenin signaling in liver development and generation. METHODS: Dissected rat livers were randomly divided into three groups: I (control); II (decellularized scaffolds) and III (recellularized scaffolds). Liver decellularization was established via an adapted perfusion procedure and assessed through the measurement of extracellular matrix (ECM) proteins and DNA content. Liver recellularization was assessed through histological examination and measurement of transcript levels of Wnt/β-catenin pathway, hepatogenesis, liver-specific microRNAs and growth factors essential for liver development. Adult rat liver decellularization was confirmed by the maintenance of ECM proteins and persistence of growth factors essential for liver regeneration. RESULTS: iPSCs seeded rat decellularized livers displayed upregulated transcript expression of Wnt/β-catenin pathway-related, growth factors, and liver specification genes. Further, recellularized livers displayed restored liver-specific functions including albumin secretion and urea synthesis. CONCLUSION: This establishes proof-of-principle for the generation of three-dimensional liver organ scaffolds as grafts and functional re-establishment

Deep-Sequencing Analysis of the Mouse Transcriptome Response to Infection with Brucella melitensis Strains of Differing Virulence

Brucella melitensis is an important zoonotic pathogen that causes brucellosis, a disease that affects sheep, cattle and occasionally humans. B. melitensis strain M5-90, a live attenuated vaccine cultured from B. melitensis strain M28, has been used as an effective tool in the control of brucellosis in goats and sheep in China. However, the molecular changes leading to attenuated virulence and pathogenicity in B. melitensis remain poorly understood. In this study we employed the Illumina Genome Analyzer platform to perform genome-wide digital gene expression (DGE) analysis of mouse peritoneal macrophage responses to B. melitensis infection. Many parallel changes in gene expression profiles were observed in M28- and M5-90-infected macrophages, suggesting that they employ similar survival strategies, notably the induction of anti-inflammatory and antiapoptotic factors. Moreover, 1019 differentially expressed macrophage transcripts were identified 4 h after infection with the different B. melitensis strains, and these differential transcripts notably identified genes involved in the lysosome and mitogen-activated protein kinase (MAPK) pathways. Further analysis employed gene ontology (GO) analysis: high-enrichment GOs identified endocytosis, inflammatory, apoptosis, and transport pathways. Path-Net and Signal-Net analysis highlighted the MAPK pathway as the key regulatory pathway. Moreover, the key differentially expressed genes of the significant pathways were apoptosis-related. These findings demonstrate previously unrecognized changes in gene transcription that are associated with B. melitensis infection of macrophages, and the central signaling pathways identified here merit further investigation. Our data provide new insights into the molecular attenuation mechanism of strain M5-90 and will facilitate the generation of new attenuated vaccine strains with enhanced efficacy

The Twin-Arginine Translocation Pathway in α-Proteobacteria Is Functionally Preserved Irrespective of Genomic and Regulatory Divergence

The twin-arginine translocation (Tat) pathway exports fully folded proteins out of the cytoplasm of Gram-negative and Gram-positive bacteria. Although much progress has been made in unraveling the molecular mechanism and biochemical characterization of the Tat system, little is known concerning its functionality and biological role to confer adaptive skills, symbiosis or pathogenesis in the α-proteobacteria class. A comparative genomic analysis in the α-proteobacteria class confirmed the presence of tatA, tatB, and tatC genes in almost all genomes, but significant variations in gene synteny and rearrangements were found in the order Rickettsiales with respect to the typically described operon organization. Transcription of tat genes was confirmed for Anaplasma marginale str. St. Maries and Brucella abortus 2308, two α-proteobacteria with full and partial intracellular lifestyles, respectively. The tat genes of A. marginale are scattered throughout the genome, in contrast to the more generalized operon organization. Particularly, tatA showed an approximately 20-fold increase in mRNA levels relative to tatB and tatC. We showed Tat functionality in B. abortus 2308 for the first time, and confirmed conservation of functionality in A. marginale. We present the first experimental description of the Tat system in the Anaplasmataceae and Brucellaceae families. In particular, in A. marginale Tat functionality is conserved despite operon splitting as a consequence of genome rearrangements. Further studies will be required to understand how the proper stoichiometry of the Tat protein complex and its biological role are achieved. In addition, the predicted substrates might be the evidence of role of the Tat translocation system in the transition process from a free-living to a parasitic lifestyle in these α-proteobacteria

Study on Surface Permeability of Concrete under Immersion

In this paper, concrete specimens are immersed in ultrapure water, to study the evolutions of surface permeability, pore structure and paste microstructure following the prolonging of immersion period. According to the results, after 30-day immersion, the surface permeability of concrete becomes higher as compared with the value before immersion. However, further immersion makes the surface permeability decrease, so that the value measured after 150-day immersion is only half that measured after 30-day immersion. The early increase in surface permeability should be mainly attributed to the leaching of calcium hydroxide, while the later decrease to the refinement of pore structure due to hydration. The two effects work simultaneously and compete throughout the immersion period. The proposed mechanisms get support from microscopic measurements and observations