36 research outputs found
Additional file 2: Table S1. of Relationship of clusterin with renal inflammation and fibrosis after the recovery phase of ischemia-reperfusion injury
Fibrosis-related gene expression in CLU KO kidneys compared to WT controls after 30ΓΒ days of IRI. (DOCX 21 kb
Additional file 1: Figure S1. of Relationship of clusterin with renal inflammation and fibrosis after the recovery phase of ischemia-reperfusion injury
Tubular dilatation and degeneration in outer area of renal cortex of some WT kidneys. The sections of WT kidneys (Γ’ΒΔ½ 1.5 of fibrosis score) were stained with MT (top) or H&E (middle and bottom). Data were presented as a typical microscopic image of renal cortex in each type of stain (MT or H&E stained sections), showing the tubular dilatation and degeneration in the squared area of the renal cortex. (TIFF 9328 kb
BIRC6 Protein, an Inhibitor of Apoptosis: Role in Survival of Human Prostate Cancer Cells
<div><h3>Background</h3><p>BIRC6 is a member of the Inhibitors of Apoptosis Protein (IAP) family which is thought to protect a variety of cancer cells from apoptosis. The main objective of the present study was to investigate whether BIRC6 plays a role in prostate cancer and could be useful as a novel therapeutic target.</p> <h3>Methods</h3><p>BIRC6 expression in cell lines was assessed using Western blot analysis and in clinical samples using immunohistochemistry of tissue microarrays. The biological significance of BIRC6 was determined by siRNA-induced reduction of <em>BIRC6</em> expression in LNCaP cells followed by functional assays.</p> <h3>Results</h3><p>Elevated BIRC6 protein expression was found in prostate cancer cell lines and clinical specimens as distinct from their benign counterparts. Increased BIRC6 expression was associated with Gleason 6β8 cancers and castration resistance. Reduction of BIRC6 expression in LNCaP cells led to a marked reduction in cell proliferation which was associated with an increase in apoptosis and a decrease in autophagosome formation. Doxorubicin-induced apoptosis was found to be coupled to a reduction in BIRC6 protein expression.</p> <h3>Conclusion</h3><p>The data suggest a role for BIRC6 in prostate cancer progression and treatment resistance, and indicate for the first time that the <em>BIRC6</em> gene and its product are potentially valuable targets for treatment of prostate cancers.</p> </div
Reduction of BIRC6 expression decreases prostate cancer cell viability and proliferation.
<p><b>A</b>, treatment of LNCaP cell cultures with siRNAs targeting BIRC6 leads to reduction of BIRC6 protein expression. BIRC6 protein expression in untreated LNCaP cells (Lane 1; at 78 h) and in LNCaP cells incubated with lipofectamine only (Lanes 2, 6, 10), siRNA-1 targeting BIRC6 (Lanes 3, 7, 11), siRNA-2 targeting BIRC6 (Lanes 4, 8, 12) or non-targeting siRNA (Lanes 5, 9, 13) for 30, 54 and 78 h after transfection. The results are representative of three independent experiments. <b>B</b>, treatment of LNCaP cell cultures with siRNA-2 targeting BIRC6 leads to reduced cell proliferation as shown by MTT assay. Cultures were treated for 30 h with lipofectamine only or with lipofectamine plus either non-targeting siRNA or siRNA-2 targeting BIRC6 and then incubated for 24, 48 and 72 h in fresh media. The relative cell numbers in the siRNA-2 cultures were considerably lower than those in the non-targeting siRNA treated cultures by 2.85%, 10.78% and 25.88% at 54, 78 and 102 h, respectively. Error bars depict standard deviations.</p
Pathological Parameters of Study Patients and PSA Levels.
<p>Pathological Parameters of Study Patients and PSA Levels.</p
Decreased BIRC6 expression induces apoptosis and inhibits autophagosome formation.
<p><b>A</b>, apoptosis of LNCap cells transfected with Lipofectamine (Lipo), non-targeting control siRNA (NT) or <i>BIRC6</i> siRNA2 and subsequently incubated for 48 h, as assessed by flow cytometric analysis of Annexin V/PI stained cells. On the basis of Q2+Q3 values (apoptotic cell populations), <i>BIRC6</i> siRNA markedly increased apoptosis of LNCaP cells compared with Lipo (pβ=β0.0104) or NT siRNA treated cells (pβ=β0.0123). (Data are representative of 3 independent experiments); <b>B</b>, treatment of LNCaP cells with siRNA-2 targeting BIRC6 leads to activation of caspase-3 (as shown by appearance of cleaved caspase-3) and degradation of its substrate, PARP (as shown by loss of full length PARP and appearance of a cleaved PARP product). Untreated LNCaP cells (Lane 1); LNCaP cells incubated with lipofectamine only (Lane 2), siRNA-2 targeting BIRC6 (Lane 3) or non-targeting siRNA (Lane 4) for 96 h following transfection. <b>C</b>, treatment of LNCaP cells with siRNA-2 targeting BIRC6 leads to decreases in Beclin-1 expression and reduction of LC3B-II. Untreated LNCaP cells (Lane 1); LNCaP cells incubated with lipofectamine only (Lane 2), siRNA-2 targeting BIRC6 (Lane 3) or non-targeting siRNA (Lane 4) for 113 h following transfection. <b>D</b>, autophagosome formation in BIRC6-silenced cells was significantly lower compared to cells treated with non-targeting control siRNA. Cells transfected with non-targeting siRNA control (left) showed more LC3-GFP puncta than cells transfected with siRNA-2 targeting BIRC6 (right), which showed diffused expression of LC3-GFP. <b>E</b>, Reduced BIRC6-expressing LNCaP cells shows significantly fewer autophagic cells (33.6%) compared to control cells (51%). Autophagic cells were quantified by counting cells with more than 15 LC3-GFP puncta under a confocal microscope.</p
BIRC6 protein expression is elevated in prostate cancer cells in comparison with benign prostate cells.
<p>BIRC6 protein expression in malignant prostate cells (Lanes 1β6), positive control malignant cervical and ovarian cells (Lanes 7β8) and benign prostate cells (Lanes 9β10).</p
Doxorubicin-induced apoptosis in LNCaP cells is associated with a reduction in BIRC6 protein expression.
<p><b>A</b>, incubation (24 h) of LNCaP cells with doxorubicin (1 Β΅g/mL) leads to reduction of BIRC6 protein expression and apoptosis (loss of full length PARP and appearance of cleaved PARP) as indicated by Western blot analysis. <b>B</b>, LNCaP cells incubated for 24 h (1) without doxorubicin and (2) with doxorubicin (1 Β΅g/mL) at 100Γ magnification. <b>C</b>, Expression of BIRC6 and PARP cleavage were studied in LNCaP cells incubated with doxorubicin (500 ng/mL) for 4, 8, 12, 24 h. BIRC6 expression was decreased upon treatment in a time-dependent manner and the reduction precedes apoptosis induction as indicated by PARP cleavage at 24 h.</p
BIRC6 protein expression in TMA sections of prostate tissue samples.
<p><b>A</b>, arrows indicating weak cytoplasmic BIRC6 staining in benign prostatic luminal cells (score β1β²). <b>B</b>, arrows indicating moderate cytoplasmic BIRC6 staining in Gleason grade 3 prostate cancer cells (score β2β²). <b>C</b>, arrows indicating strong cytoplasmic BIRC6 staining in Gleason grade 4 prostate cancer cells (score β3β²). <b>D</b>, arrows indicating weak cytoplasmic BIRC6 staining in Gleason grade 5 prostate cancer cells (score β1β²). <b>E</b>, Black bars representing mean BIRC6 staining intensity in benign prostate epithelium (nβ=β35) and Gleason score 6 (nβ=β74), 7 (nβ=β23), 8 (nβ=β43) and 9β10 (nβ=β17) patient prostate cancer tissues. Grey bars representing frequency of BIRC6 expression (score β₯1). Elevated staining intensities were observed in Gleason score 6 (*<i>P</i>β=β3.24Γ10β3), 7 (*<i>P</i>β=β4.45Γ10β6) and 8 (*<i>P</i>β=β1.63Γ10β3) prostate cancer tissues as compared to benign prostate epithelium. Error bars depict standard deviations. <b>F</b>, Black bars representing mean BIRC6 staining intensity in untreated high grade (Gleason score 8β10) prostate cancer tissues (nβ=β60) and castration-resistant prostate cancer (CRPC) tissues which had developed from Gleason score 8β10 cancers following neo-adjuvant hormone therapy (nβ=β10). Grey bars representing frequency of BIRC6 expression (score β₯1). Castration-resistant prostate cancer tissues showed a significantly higher mean BIRC6 staining intensity than untreated high grade prostate cancer tissues (*<i>P</i>β=β1.38Γ10β3). Error bars depict standard deviations.</p
PR negatively regulates prostate cancer migration through a paracrine pathway.
<p>Conditioned media (CM) were collected from parental hCAFs, WPMY-1 or their derived cell lines expressing mock, PRA or PRB in the presence of vehicle or 10 nM P4. PC-3 cells were seeded in 6 well plates and incubated with CM from hCAFs (<b>A</b>) or from WPMY-1 (<b>B</b>) cells for 24 hours in wound healing assays. Representative images after 24 hour CM treatment were captured by an inverted microscope. WPMY-1 and its derived cell lines expressing mock, PRA or PRB were treated with 0, 10 nM and 100 nM of P4 for 24 hours (<b>C and E</b>) or with vehicle, 10 nM of P4 and/or 10 uM of RU486 for 24 hours (<b>D and F</b>). CM were then collected and incubated with PC-3 cells (<b>CβD</b>) and C4-2B (<b>EβF</b>) in cell migration assays as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0092714#s2" target="_blank">material and methods</a> section. One-way ANOVA and paired student's t-test calculate the statistical significance set at P<0.05 as * and P<0.001 as ***.</p