Fusion of bone marrow-derived cells with renal tubules contributes to renal dysfunction in diabetic nephropathy

Although diabetic nephropathy (DN) is a major cause of end-stage renal disease, the mechanism of dysfunction has not yet been clarified. We previously reported that in diabetes proinsulin-producing bone marrow-derived cells (BMDCs) fuse with hepatocytes and neurons. Fusion cells are polyploidy and produce tumor necrosis factor (TNF)-α, ultimately causing diabetic complications. In this study, we assessed whether the same mechanism is involved in DN. We performed bone marrow transplantation from male GFP-Tg mice to female C57BL/6J mice and produced diabetes by streptozotocin (STZ) or a high-fat diet. In diabetic kidneys, massive infiltration of BMDCs and tubulointerstitial injury were prominent. BMDCs and damaged tubular epithelial cells were positively stained with proinsulin and TNF-α. Cell fusion between BMDCs and renal tubules was confirmed by the presence of Y chromosome. Of tubular epithelial cells, 15.4% contain Y chromosomes in STZ-diabetic mice, 8.6% in HFD-diabetic mice, but only 1.5% in nondiabetic mice. Fusion cells primarily expressed TNF-α and caspase-3 in diabetic kidney. These in vivo findings were confirmed by in vitro coculture experiments between isolated renal tubular cells and BMDCs. It was concluded that cell fusion between BMDCs and renal tubular epithelial cells plays a crucial role in DN.     

Coexisting <Emphasis Type="Italic">Curtobacterium</Emphasis> Bacterium Promotes Growth of White-Rot Fungus <Emphasis Type="Italic">Stereum</Emphasis> sp.

White-rot basidiomycetes are the main decomposers of woody biomass in forest ecosystems. Little is known, however, about the interactions between white-rot fungi and other microorganisms in decayed wood. A wood-rotting fungus, Stereum sp. strain TN4F, was isolated from a fruit body, and its coexisting cultivable bacteria were isolated from its substrate; natural white-rot decayed wood. The effects of bacteria on fungal growth were examined by confrontational assay in vitro. A growth-promoting bacterium for this Stereum strain was identified as Curtobacterium sp. TN4W-19, using 16SrRNA sequencing. A confrontational assay revealed that Curtobacterium sp. TN4W-19 significantly promoted the mycelial growth of Stereum sp. TN4F in the direction of the bacterial colony, without direct contact between the mycelium and bacterial cells. This is the first report of a positive interaction between a white-rot fungus and a coexisting bacterial strain in vitro.     

Negative Regulation of Human Astrocytes by Interferon (IFN) α in Relation to Growth Inhibition and Impaired Glucose Utilization

The present study assessed the direct effects of IFNs on human astrocytes. Human astrocytes were exposed to human recombinant IFNs, and the proliferation of cells was measured. Type I IFN receptor mRNA and protein expression, the phosphoprotein levels of signaling molecules including JNK, ERK1/2, IκB, p38MAPK, Stat3, and the expression of cytokines were determined respectively. In addition, cellular glucose consumption was measured as well as Glut-1 protein and activation of GSK-3β/mTOR signal were determined. The expression of Type I IFN receptor was detected in cultured human astrocytes. 2?IU/ml IFNα2a and IFNα2b significantly decreased the proliferation of human astrocytes respectively, compared to control. IFNβ had no significant effect on the proliferation of the cells. The phosphorylation of JNK stimulated by all IFNs detected was more pronounced and sustained than ERK1/2 and IκB. No effects were observed on the activation of p38MAPK and Stat3. Moreover, Treatment with IFNα, especially with IFNα2b, decreased glucose consumption and stimulated phosphorylation of GSK-3β and mTOR, but decreased the expression of Glut-1. In contrast, IFNβ had no significant effect on either glucose consumption or activation of GSK-3β/mTOR signals. INFα2b significantly decreased the levels of IL-8 whereas the levels of GM-CSF were increased. The present study demonstrates direct inhibitory effects of IFNα on cell proliferation, cell signaling and glucose utilization in human astrocytes.     

Thirteen novel deoxynivalenol-degrading bacteria are classified within two genera with distinct degradation mechanisms

The mycotoxin deoxynivalenol (DON), a secondary metabolite produced by species of the plant pathogen Fusarium, causes serious problems in cereal crop production because of its toxicity towards humans and livestock. A biological approach for the degradation of DON using a DON-degrading bacterium (DDB) appears to be promising, although information about DDBs is limited. We isolated 13 aerobic DDBs from a variety of environmental samples, including field soils and wheat leaves. Of these 13 strains, nine belonged to the Gram-positive genus Nocardioides and other four to the Gram-negative genus Devosia. The degradation phenotypes of the two Gram types were clearly different; all washed cells of the 13 strains degraded 100 μg mL(-1) DON to below the detection limit (0.5 μg mL(-1)), but the conditions inducing the DON-degrading activities differed between the two Gram types. The HPLC profiles of the DON metabolites were also distinct between the two genera, although all strains produced 3-epi-deoxynivalenol. The Gram-positive strains showed DON assimilation in media containing DON as a carbon source, whereas the Gram-negatives did not. Our results suggest that aerobic DDBs are distributed within at least two phylogenetically restricted genera, suggesting independent evolution of the DON-degradation mechanisms.     

Differential requirement for the N-terminal catalytic domain of the DNA polymerase ε p255 subunit in the mitotic cell cycle and the endocycle

In Drosophila, the 255kDa catalytic subunit (dpolεp255) and the 58kDa subunit of DNA polymerase ε (dpolεp58) have been identified. The N-terminus of dpolεp255 carries well-conserved six DNA polymerase subdomains and five 3'→5' exonuclease motifs as observed with Polε in other species. We here examined roles of dpolεp255 during Drosophila development using transgenic fly lines expressing double stranded RNA (dsRNA). Expression of dpolεp255 dsRNA in eye discs induced a small eye phenotype and inhibited DNA synthesis, indicating a role in the G1-S transition and/or S-phase progression of the mitotic cycle. Similarly, expression of dpolεp255 dsRNA in the salivary glands resulted in small size and endoreplication defects, demonstrating a critical role in endocycle progression. In the eye disc, defects induced by knockdown of dpolεp255 were rescued by overexpression of the C-terminal region of dpolεp255, indicating that the function of this non-catalytic domain is conserved between yeast and Drosophila. However, this was not the case for the salivary gland, suggesting that the catalytic N-terminal region is crucial for endoreplication and its defect cannot be complemented by other DNA polymerases. In addition, several genetic interactants with dpolεp255 including genes related to DNA replication such as RFC, DNA primase, DNA polη, Mcm10 and Psf2 and chromatin remodeling such as Iswi were also identified.     

Structural and biochemical characterization of glycoside hydrolase family 79 β-glucuronidase from Acidobacterium capsulatum

We present the first structure of a glycoside hydrolase family 79 β-glucuronidase from Acidobacterium capsulatum, both as a product complex with β-D-glucuronic acid (GlcA) and as its trapped covalent 2-fluoroglucuronyl intermediate. This enzyme consists of a catalytic (β/α)(8)-barrel domain and a β-domain with irregular Greek key motifs that is of unknown function. The enzyme showed β-glucuronidase activity and trace levels of β-glucosidase and β-xylosidase activities. In conjunction with mutagenesis studies, these structures identify the catalytic residues as Glu(173) (acid base) and Glu(287) (nucleophile), consistent with the retaining mechanism demonstrated by (1)H NMR analysis. Glu(45), Tyr(243), Tyr(292)-Gly(294), and Tyr(334) form the catalytic pocket and provide substrate discrimination. Consistent with this, the Y292A mutation, which affects the interaction between the main chains of Gln(293) and Gly(294) and the GlcA carboxyl group, resulted in significant loss of β-glucuronidase activity while retaining the side activities at wild-type levels. Likewise, although the β-glucuronidase activity of the Y334F mutant is ~200-fold lower (k(cat)/K(m)) than that of the wild-type enzyme, the β-glucosidase activity is actually 3 times higher and the β-xylosidase activity is only 2.5-fold lower than the equivalent parameters for wild type, consistent with a role for Tyr(334) in recognition of the C6 position of GlcA. The involvement of Glu(45) in discriminating against binding of the O-methyl group at the C4 position of GlcA is revealed in the fact that the E45D mutant hydrolyzes PNP-β-GlcA approximately 300-fold slower (k(cat)/K(m)) than does the wild-type enzyme, whereas 4-O-methyl-GlcA-containing oligosaccharides are hydrolyzed only 7-fold slower.