The aggregatibacter actinomycetemcomitans heat shock protein GroEL interacts directly with human peripheral blood T cells

Heat shock family protein GroEL of Aggregatibacter actinomycetemcomitans (Aa) has antigenic properties. We previously demonstrated that A. actinomycetemcomitans GroEL-like protein affects human CD4 T cells by converting them into IL-10 and IFNg double cytokine producing Tbet+ Th1 cells. The objective of this study was to investigate whether or not AaGroEL communicates with T cells directly. To do this, sorted cells from peripheral blood mononuclear cells were stimulated with AaGroEL for 48 h. Flow cytometry was used to measure soluble and intracellular cytokine expression in the cell cultures and detect TLR2 expression on the surface of T cells. Expression of six different soluble cytokines was evaluated by CBA assay. To determine whether AaGroEL affects CD3+ T cells directly or not, purified CD3+ T cells or CD14+ cells were cultured with AaGroEL separately, and the quantity of soluble cytokine was measured. Results showed that sorted CD3+ cells produced soluble IL-6, TNFα-and IFNγ cytokines. Additionally, the intracellular cytokine staining data showed that AaGroEL-stimulated CD3+ cells were also TNFα-and IFNγ-positive. Moreover, AaGroEL-responsive T cells slightly increased their TLR2 expression. These findings suggest that CD3+ T cells produce cytokines in response to AaGroEL protein without requirements for other cells, such as CD14+ monocytes.Scientific and Technological Research Council of Turkey (TUBITAK 106T417

Pharmacological Induction of Transforming Growth Factor-Beta1 in Rat Models Enhances Radiation Injury in the Intestine and the Heart

<div><p>Radiation therapy in the treatment of cancer is dose limited by radiation injury in normal tissues such as the intestine and the heart. To identify the mechanistic involvement of transforming growth factor-beta 1 (TGF-β1) in intestinal and cardiac radiation injury, we studied the influence of pharmacological induction of TGF-β1 with xaliproden (SR 57746A) in rat models of radiation enteropathy and radiation-induced heart disease (RIHD). Because it was uncertain to what extent TGF-β induction may enhance radiation injury in heart and intestine, animals were exposed to irradiation schedules that cause mild to moderate (acute) radiation injury. In the radiation enteropathy model, male Sprague-Dawley rats received local irradiation of a 4-cm loop of rat ileum with 7 once-daily fractions of 5.6 Gy, and intestinal injury was assessed at 2 weeks and 12 weeks after irradiation. In the RIHD model, male Sprague-Dawley rats received local heart irradiation with a single dose of 18 Gy and were followed for 6 months after irradiation. Rats were treated orally with xaliproden starting 3 days before irradiation until the end of the experiments. Treatment with xaliproden increased circulating TGF-β1 levels by 300% and significantly induced expression of TGF-β1 and TGF-β1 target genes in the irradiated intestine and heart. Various radiation-induced structural changes in the intestine at 2 and 12 weeks were significantly enhanced with TGF-β1 induction. Similarly, in the RIHD model induction of TGF-β1 augmented radiation-induced changes in cardiac function and myocardial fibrosis. These results lend further support for the direct involvement of TGF-β1 in biological mechanisms of radiation-induced adverse remodeling in the intestine and the heart.</p></div

Histopathological manifestations of radiation enteropathy.

<p>Xaliproden significantly reduced mucosal surface area (A) at 12 weeks after local irradiation of the small intestine, and enhanced radiation injury score (B), intestinal wall thickness (C) and serosal thickness (D) at 2 weeks and 12 weeks after irradiation. Average ± SEM, n = 5 (sham-irradiated) or 13–15 (irradiated). *p<0.05, ^†p<0.001. Representative micrographs of histopathological stainings are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070479#pone.0070479.s003" target="_blank">Figure S3</a>.</p

Activation of apoptosis pathways in mucosa and whole gut.

<p>Stacked bar charts demonstrate IPA-generated activated apoptosis pathways in mucosa and whole gut at 4 h, 24 h, and 3.5 d after irradiation. The height of the bars indicates the percentage of genes that changed in the particular pathway. Red bar: up-regulated. Green bar: down-regulated. Pathways with p-value (yellow dot) above the threshold (dashed line) are significantly activated. Heatmaps demonstrate the change of the genes in the selected signaling pathway before (left column) and after radiation (right column). Blue: decreased, red: increased.</p

Echocardiographic parameters (average ± SEM) at 3 and 6 months after local heart irradiation.

#<p>Significant difference with sham-irradiated animals (p<0.05).</p>*<p>Significant difference with vehicle-treated animals (p<0.05).</p

Venn diagram demonstrating radiation- induced differential transcriptional profiles in mucosa and whole gut.

<p>Venn diagram demonstrating radiation- induced differential transcriptional profiles in mucosa and whole gut.</p

<i>Ex vivo</i> cardiac function analysis at 6 months after local heart irradiation and xaliproden treatment.

<p>Xaliproden significantly enhanced the effects of radiation on diastolic wall stress (A), systolic wall stress (B), and coronary pressure (C) as measured in Langendorff isolated perfused hearts at 6 months after local heart irradiation. Average ± SEM, n = 3–6. *Significant difference with sham-irradiated vehicle-treated animals (p<0.05).</p

Left ventricular collagen area (average ± SEM) and scores for α-SMC actin containing myofibroblasts at 6 months after local heart irradiation.

#<p>Significant difference with sham-irradiated animals (p<0.05).</p>*<p>Significant difference with all other groups (p<0.05).</p><p>Representative micrographs of histopathological stainings are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070479#pone.0070479.s002" target="_blank">Figure S2</a>.</p

Activation of cell cycle control pathways in mucosa and whole gut.

<p>Stacked bar charts demonstrate IPA-generated activated cell cycle control pathways in mucosa and whole gut at 4 h, 24 h, and 3.5 d after irradiation. The height of the bars indicates the percentage of genes that changed in the particular pathway. Red bar: up-regulated. Green bar: down-regulated. Pathways with p-value (yellow dot) above the threshold (dashed line) are significantly activated. Heatmaps demonstrate the change of the genes in the selected signaling pathway before (left column) and after radiation (right column). Blue: decreased, red: increased.</p

Expression of Caspase14 mRNA and protein in the small intestines.

<p><b>A</b>: Immunohistochemistry demonstrating the expression of Caspase14 in goblet cells in baseline (control) and irradiated intestine at 4 h, 24 h and 3.5 d post irradiation. <b>B</b>: Expression of Caspase14 mRNA in mucosa and whole gut tissue plotted by the raw signal from microarray. <b>C</b>: A higher magnification power demonstrating goblet cells secreting Caspase14 at 24 h post irradiation. Red: Caspase14; Green: E-cadherin; Blue: DAPI.</p