Expression of fibrosis-related genes in liver and kidney fibrosis in comparison to inflammatory bowel diseases

Fibrosis is an important feature of inflammatory bowel diseases (IBD), but its pathogenesis is incompletely understood. Our aim was to identify genes important for fibrosis in IBD by comparison with kidney and liver fibrosis. First, we performed bioinformatics analysis of Gene Expression Omnibus datasets of liver and kidney fibrosis and identified CXCL9, THBS2, MGP, PTPRC, CD52, GZMA, DPT and DCN as potentially important genes with altered expression in fibrosis. We then performed qPCR analysis of the selected genes’ expression on samples of fibrotic kidney, liver, Crohn’s disease (CD) with and without fibrosis and ulcerative colitis (UC), in comparison to corresponding normal tissue. We found significantly altered expression in fibrosis for all selected genes. A significant difference for some genes was observed in CD with fibrosis in comparison to CD without fibrosis and UC. We conclude that similar changes in the expression of selected genes in liver, kidney fibrosis and IBD provide further evidence that fibrosis in IBD might share common mechanisms with other organs, supporting the hypothesis that fibrosis is the common pathway in diseases of various organs. Some genes were already active in IBD with inflammation without fibrosis, suggesting the early activation of profibrotic pathways or overlapping function in fibrosis and inflammation

Expression of cytokine-coding genes BMP8B, LEFTY1 and INSL5 could distinguish between ulcerative colitis and Crohn\u27s disease

Ulcerative colitis (UC) and Crohn’s disease (CD) are characterized by an imbalance between pro-inflammatory and anti-inflammatory cytokines, interfering with the resolution of inflammation. Due to the crucial role of cytokines, new insights into their profiles in UC and CD would help to improve our understanding of pathogenesis and enable the development of new treatment modalities. We provide an expression profile of cytokines in UC and CD, using bioinformatics approach, and experimental validation of expression of the selected genes. We retrieved data and analyzed the cytokine gene expression profiles of UC and CD. From ten genes with inverse expression, common to CD and UC, BMP8B, LEFTY1 and INSL5 were selected for gene expression experimental validation. Experimentally, BMP8B and INSL5 were down-regulated in both CD and UC but followed the bioinformatics trend. The expression of genes LEFTY1 and BMP8B was statistically significant when comparing UC and CD in colon and the expression of gene LEFTY1 showed statistical significance when CD in ileum and colon were compared. Using the bioinformatics approach and experimental validation, we found differences in expression profiles between UC and CD for INSL5, LEFTY1 and BMP8B. These three promising candidate genes need to be further explored at different levels, such as DNA methylation and protein expression, to provide more evidence on their potential diagnostic role in CD and UC

Engineered combinatorial cell device for wound healing and bone regeneration

Growth factors are the key regulators that promote tissue regeneration and healing processes. While the effects of individual growth factors are well documented, a combination of multiple secreted growth factors underlies stem cell–mediated regeneration. To avoid the potential dangers and laborintensive individual approach of stem cell therapy while maintaining their regeneration-promoting effects based on multiple secreted growth factors, we engineered a “mix-and-match” combinatorial platform based on a library of cell lines producing growth factors. Treatment with a combination of growth factors secreted by engineered mammalian cells was more efficient than with individual growth factors or even stem cell–conditioned medium in a gap closure assay. Furthermore, we implemented in a mouse model a device for allogenic cell therapy for an in situ production of growth factors, where it improved cutaneous wound healing. Augmented bone regeneration was achieved on calvarial bone defects in rats treated with a cell device secreting IGF, FGF, PDGF, TGF-β, and VEGF. In both in vivo models, the systemic concentration of secreted factors was negligible, demonstrating the local effect of the regeneration device. Finally, we introduced a genetic switch that enables temporal control over combinations of trophic factors released at different stages of regeneration mimicking the maturation of natural wound healing to improve therapy and prevent scar formation

Pathology of fibrosis in Crohn\u27s disease - contribution to understanding its pathogenesis

Background: Despite significant progress in the research of fibrosis in various organs, fibrosis remains a poorly understood complication of Crohn\u27s disease (CD). We analyzed pathologic features of fibrosis and inflammation in CD and compared them with the normal bowel, aiming to clarify whether fibrosis in CD pathogenetically resembles fibrosis in other organs. Methods: Resection specimens from 30 patients with CD were included. Normal bowel from resection specimens of colorectal carcinoma was used for comparison. Trichrome Masson staining, immunohistochemistry for α-smooth muscle actin, fibroblast activation protein, CD34 and erg, in situ hybridization for TGF-β1 and analysis of selected fibrosis-related microRNAs were performed. Results: In normal bowel, CD34-positive fibroblasts/pericytes were detected in the submucosa and subserosa, particularly around blood vessels. In CD, fibrosis prevailed in the submucosa and subserosa, together with proliferation of myofibroblasts and disappearance of CD34-positive fibroblasts/pericytes. TGF-β1 was present in the lamina propria in normal bowel and CD, and in deeper parts of the bowel wall in CD. MicroRNAs miR-29c, miR-155 miR-150, and miR-155, which have been demonstrated to contribute to fibrosis in various organs, showed significant deregulation in CD. Conclusions: Distribution of fibroblasts/pericytes in the submucosa and subserosa of normal bowel, their disappearance in fibrosis in CD, together with the appearance of myofibroblasts, suggest that fibroblasts/pericytes are the most likely source of myofibroblasts in CD. Furthemore, fibrosis-related microRNAs showed deregulation in fibrotic areas. Pathogenesis of fibrosis in CD is thus comparable to fibrosis in other organs, in which myofibroblasts are the key effector cells, and pericytes have emerged as the main origin of myofibroblasts. Fibrosis in CD should be regarded as a result of (over)response of the bowel wall to the presence of inflammation in deep structures of the bowel wall, presenting another example of a common pathogenetic pathway of fibrosis development

Interactions of archaeal chromatin proteins Alba1 and Alba2 with nucleic acids.

BACKGROUND: Architectural proteins have important roles in compacting and organising chromosomal DNA. There are two potential histone counterpart peptide sequences (Alba1 and Alba2) in the Aeropyrum pernix genome (APE1832.1 and APE1823). METHODOLOGY/PRINCIPAL FINDINGS: THESE TWO PEPTIDES WERE EXPRESSED AND THEIR INTERACTIONS WITH VARIOUS DNAS WERE STUDIED USING A COMBINATION OF VARIOUS EXPERIMENTAL TECHNIQUES: surface plasmon resonance, UV spectrophotometry, circular dichroism-spectropolarimetry, gel-shift assays, and isothermal titration calorimetry. CONCLUSIONS/SIGNIFICANCE: Our data indicate that there are significant differences in the properties of the Alba1 and Alba2 proteins. Both of these Alba proteins can thermally stabilise DNA polynucleotides, as seen from UV melting curves. Alba2 and equimolar mixtures of Alba1/Alba2 have greater effects on the thermal stability of poly(dA-dT).poly(dA-dT). Surface plasmon resonance sensorgrams for binding of Alba1, Alba2, and equimolar mixtures of Alba1/Alba2 to DNA oligonucleotides show different binding patterns. Circular dichroism indicates that Alba2 has a less-ordered secondary structure than Alba1. The secondary structures of the Alba proteins are not significantly influenced by DNA binding, even at high temperatures. Based on these data, we conclude that Alba1, Alba2, and equimolar mixtures of Alba1/Alba2 show different properties in their binding to various DNAs

UV melting curves of GC-DNA.

<p>The molar ratios of protein:DNA base pairs were at 1∶30 to 1∶2 for Alba1 (a), at 1∶30 to 1∶10 for Alba2 (b), and at 1∶30 to 1∶20 for the Alba1/Alba2 complex (c).</p

Molecular modelling of the Alba protein dimers.

<p>Left: Distances between the β3–β4 hairpins on the Alba1/Alba1 homodimer (top), the Alba1/Alba2 heterodimer (middle), and the Alba2/Alba2 homodimer (bottom). Right: Alba2 dimers covering the DNA duplex with maximal binding density of one dimer per six base pairs.</p

CD spectra of Alba1/Alba2 complex.

<p>Molar ellipticity, [Θ], in the far-UV range (200–250 nm) at different temperatures (as indicated), for the Alba1/Alba2 complex without bound DNA (a), with CT-DNA (b), with AT-DNA (c), and with GC-DNA (d). Molar ratio of Alba:DNA per base pair was 1∶5 at pH 7.0 (50 mM NaH₂PO₄).</p

CD spectra of Alba1.

<p>Molar ellipticity, [Θ], in the far-UV range (200–250 nm) at different temperatures (as indicated), for Alba1 without bound DNA (a), with CT-DNA (b), with AT-DNA (c), and with GC-DNA (d). The molar ratio of Alba:DNA per base pair was 1∶5 at pH 7.0 (50 mM NaH₂PO₄).</p

UV melting curves of AT-DNA.

<p>The molar ratios of protein:DNA base pairs were at 1∶30 to 1∶5 for Alba1 (a) and at 1∶30 to 1∶2 for Alba2 (b) and the Alba1/Alba2 complex (c).</p