Acta de Renovación de Convenio de cooperación académica y científica entre la Università della Calabria (UNICAL, Italia) y la Universidad Nacional de la Plata, (UNLP, Argentina)

La Università della Calabria (UNICAL, Italia) y la Universidad Nacional de la Plata, (UNLP, Argentina) acuerdan renovar en todos sus términos, y por un período de vigencia de cinco (5) años, el Convenio de Colaboración Científica y Cultural suscripto entre las partes el 1 del mes de septiembre de 2008 en el cual convinieron en programar y ejecutar conjuntamente acciones de cooperación en los campos de la Docencia, Extensión, Investigación y Postgrado, así como el intercambio de información, de recursos humanos y materiales, a través de proyectos comunes de asistencia técnica.Universidad Nacional de La Plat

DNA demethylation-dependent enhancement of toll-like receptor-2 gene expression in cystic fibrosis epithelial cells involves SP1-activated transcription-2

PGL3-T2P (-120), SP1 binding site-mutant pGL3-T2P (-120) SPm, pGL3-T2P (-12), or pGL3-null plasmids were each transiently transfected into HeLa cells. Luciferase activity is expressed as the percent activity of the pGL3-T2P (-120) construct. (B) Assessment of the effect of mutating the ETS binding site within the TLR2 -120/+110 promoter region. The pGL3-T2P (-120), ETS binding site-mutant pGL3-T2P (-120) ETSm, pGL3-T2P (-12), or pGL3-null plasmids were each transiently transfected into HeLa cells. Luciferase activity is expressed as the percent activity of the pGL3-T2P (-120) construct. (C) Assessment of the effect of mutating the ETS binding site within TLR2 -60/+110 promoter region. The pGL3-T2P (-60), mutant ETS binding site- pGL3-T2P (-60) ETSm construct, the pGL3-T2P (-12) construct, or the pGL3-null plasmid were transiently transfected into HeLa cells. Luciferase activity is expressed as the percent activity of the pGL3-T2P (-60) construct. Data are presented as the mean ± S.E.M. from three independent experiments performed in triplicate. n.s. = not significant as assessed by Student's test.<p><b>Copyright information:</b></p><p>Taken from "DNA demethylation-dependent enhancement of toll-like receptor-2 gene expression in cystic fibrosis epithelial cells involves SP1-activated transcription"</p><p>http://www.biomedcentral.com/1471-2199/9/39</p><p>BMC Molecular Biology 2008;9():39-39.</p><p>Published online 21 Apr 2008</p><p>PMCID:PMC2387165.</p><p></p

DNA demethylation-dependent enhancement of toll-like receptor-2 gene expression in cystic fibrosis epithelial cells involves SP1-activated transcription-0

D as +1.<p><b>Copyright information:</b></p><p>Taken from "DNA demethylation-dependent enhancement of toll-like receptor-2 gene expression in cystic fibrosis epithelial cells involves SP1-activated transcription"</p><p>http://www.biomedcentral.com/1471-2199/9/39</p><p>BMC Molecular Biology 2008;9():39-39.</p><p>Published online 21 Apr 2008</p><p>PMCID:PMC2387165.</p><p></p

Mild Electrical Stimulation and Heat Shock Ameliorates Progressive Proteinuria and Renal Inflammation in Mouse Model of Alport Syndrome

<div><p>Alport syndrome is a hereditary glomerulopathy with proteinuria and nephritis caused by defects in genes encoding type IV collagen in the glomerular basement membrane. All male and most female patients develop end-stage renal disease. Effective treatment to stop or decelerate the progression of proteinuria and nephritis is still under investigation. Here we showed that combination treatment of mild electrical stress (MES) and heat stress (HS) ameliorated progressive proteinuria and renal injury in mouse model of Alport syndrome. The expressions of kidney injury marker neutrophil gelatinase-associated lipocalin and pro-inflammatory cytokines interleukin-6, tumor necrosis factor-α and interleukin-1β were suppressed by MES+HS treatment. The anti-proteinuric effect of MES+HS treatment is mediated by podocytic activation of phosphatidylinositol 3-OH kinase (PI3K)-Akt and heat shock protein 72 (Hsp72)-dependent pathways <em>in vitro</em> and <em>in vivo</em>. The anti-inflammatory effect of MES+HS was mediated by glomerular activation of c-jun NH₂-terminal kinase 1/2 (JNK1/2) and p38-dependent pathways <em>ex vivo</em>. Collectively, our studies show that combination treatment of MES and HS confers anti-proteinuric and anti-inflammatory effects on Alport mice likely through the activation of multiple signaling pathways including PI3K-Akt, Hsp72, JNK1/2, and p38 pathways, providing a novel candidate therapeutic strategy to decelerate the progression of patho-phenotypes in Alport syndrome.</p> </div

DNA demethylation-dependent enhancement of toll-like receptor-2 gene expression in cystic fibrosis epithelial cells involves SP1-activated transcription-4

Cted with pGL3-null and pcDNA3.1 as control. Luciferase activity is expressed as the percent activity of the pGL3-T2P (-120) plasmid in pcDNA3.1-co-transfected, methylase-untreated cells. Data are presented as the mean ± S.E.M. from three independent experiments performed in triplicate. *p < 0.0001, assessed by Student's -test.<p><b>Copyright information:</b></p><p>Taken from "DNA demethylation-dependent enhancement of toll-like receptor-2 gene expression in cystic fibrosis epithelial cells involves SP1-activated transcription"</p><p>http://www.biomedcentral.com/1471-2199/9/39</p><p>BMC Molecular Biology 2008;9():39-39.</p><p>Published online 21 Apr 2008</p><p>PMCID:PMC2387165.</p><p></p

MES+HS directly suppresses the expressions of pro-inflammatory cytokines via PI3K-Akt or Hsp72-independent pathway in isolated glomeruli of Alport mice ex vivo.

<p>(<i>a</i>) Bright field image indicates isolated glomeruli from Alport mice in culture medium. (<i>b, c</i>) Glomeruli were isolated from Alport mice by mesh sieving followed by <i>ex vivo</i> MES+HS treatment for 10 min. Four hours after treatment, total RNA was extracted from glomeruli and subjected to Q-PCR analysis to assess the cytokines mRNA expression. (<i>d</i>–<i>f</i>) Isolated glomeruli from Alport mice were pre-treated with LY294002 (20 µM) or quercetin (200 µM) for 2 hr, and treated with MES+HS for 10 min. Thirty minutes after the treatment, proteins were isolated, and immunoblotted with p-Akt and total Akt antibodies (<i>d</i>). Five hours after the treatment, protein lysates were analyzed for Hsp72 expression (<i>e</i>), or total RNA was isolated and subjected to Q-PCR analysis to assess the expression of IL-6 (<i>f</i>). Values are the mean ± S.E. (n = 3 for b, <i>c</i> & <i>f</i>). *<i>P</i><0.05, **<i>P</i><0.01 assessed by unpaired <i>t</i>-test. n.s., not significant.</p

MES+HS suppresses the expressions of pro-inflammatory cytokines in kidneys of Alport mice in vivo.

<p>(<i>a–j</i>) Forty-eight hours after the last treatment, total RNA was isolated from kidneys (<i>a</i>–<i>f</i>) or glomeruli (<i>g</i>–<i>j</i>) of Alport mice sham-treated (CON) or treated with MES+HS for 8 weeks. Q-PCR analysis was performed to assess the expression of the indicated cytokines (<i>a</i>–<i>e, g</i>–<i>j</i>) and renal injury marker NGAL (<i>f</i>). Values are the mean ± S.E. (n = 6). *<i>P</i><0.05, *** <i>P</i><0.001 assessed by unpaired <i>t</i>-test. n.s., not significant.</p

Primer sequences for Real-time quantitative RT-PCR.

<p>Primer sequences for Real-time quantitative RT-PCR.</p

MES+HS activates Akt and induces Hsp72 expression in podocytes of Alport mice in vivo.

<p>(<i>a</i>, <i>c & e</i>) One hour after the final treatment, whole kidney protein lysates or glomeruli protein lysates were isolated from 8 weeks-MES+HS-treated and control Alport mice, and immunoblotted to detect p-Akt and total Akt proteins. Actin was used as loading control (<i>a, c</i>). Renal cryosections were subjected to immunohistochemical analysis to assess phosphorylated Akt. Arrowheads indicate phosphorylated Akt in podocytes. Cryosections were counterstained with DAPI (<i>e</i>). (<i>b, d</i> & <i>f</i>) Eight hours (<i>b</i> & <i>f</i>) or five hours (<i>d</i>) after the final treatment, whole kidney or glomeruli protein lysates were isolated from MES+HS-treated (8 weeks treated for <i>b</i> & <i>f</i>, or 4 weeks treated for <i>d</i>) and control Alport mice, and immunoblotted to assess Hsp72 and HSC70 expression. α-Tubulin or actin was used as loading control (<i>b, d</i>). Renal cryosections were subjected to immunohistochemical analysis to assess Hsp72. Arrowheads indicate up-regulated Hsp72 in podocytes. Cryosections were counterstained with DAPI (<i>f</i>). (<i>e</i> & <i>f</i>) Representative glomeruli images of three independent experiments are shown. Renal sections were immunostained with anti-synaptopodin antibody to assess podocyte localization (<i>lower panels</i>). Sections were counterstained with DAPI. White bars; 10 µm.</p

MES and HS synergistically activate PI3K-Akt signaling, induce Hsp72 expression and regulate permeability activity in podocytes.

<p>(<i>a</i>) Mouse podocytes were treated with MES for 10 min using various pulse durations as indicated. Whole cell proteins were isolated 30 min after MES treatment. Phosphorylated Akt (p-Akt) and total Akt proteins were analyzed by immunoblotting. Actin was used as loading control. (<i>b</i>) Relative amount of p-Akt in cells treated with MES (0.1 ms pulse duration) was quantified and normalized to total Akt. (<i>c</i>) Cells were treated with 10 µM LY294002 for 1 hr, and stimulated with MES for 10 min. Whole cell protein lysates were isolated after 30 min. (<i>d</i>) Whole cell proteins were isolated 5 hr after HS treatment. Hsp72 protein expression was analyzed by Western blotting. (<i>e</i>) Relative amount of Hsp72 was quantified with Image Gauge Software and normalized to HSC70. (<i>f</i>) Cells were treated with HS for 10 min. Total RNA was isolated after 1 hr and Hsp72 mRNA expression was analyzed by Q-PCR. (<i>g</i> & <i>h</i>) Cells were co-treated with MES and HS for 10 min. Thirty minutes or 5 hr after the treatment, proteins were isolated and analyzed with p-Akt and total Akt (<i>g</i>) or with Hsp72 (<i>h</i>) antibodies. (<i>i</i> & <i>j</i>) Cells were plated onto Transwell chamber and cultured for 8 days. Cells were pre-treated with LY294002 (10 µM) or quercetin (100 µM) for 1 hr and treated with MES+HS for 10 min. (<i>i</i>) After treatment, 40 mg/ml BSA in RPMI-1640 solution was added to the bottom chamber. Permeability activity of podocytes was analyzed by measuring the protein concentration in the upper chamber 5 hr after adding the BSA solution. (<i>j</i>) Thirty minutes or 5 hr after the treatment, proteins were isolated and immunoblotted respectively with p-Akt and total Akt (<i>left</i>) or with Hsp72 (<i>right</i>) antibodies. For (<i>b</i>, <i>e</i>, <i>f</i> & <i>i</i>), values are the mean ± S.E. (n = 3). *<i>P</i><0.05, *** <i>P</i><0.001 assessed by unpaired <i>t</i>-test. n.s.; not significant.</p