Soybean Date of Planting and Maturity in Northern Iowa

Inevitably, every year soybean planting gets delayed or needs to be replanted because of weather somewhere in Iowa. Even if soybean planting starts and progresses in a timely manner, there always is the question of what maturity group should be planted. This trial was setup to determine what maturities are well suited for a given geographic location, but also how maturity selection should be adjusted as planting dates get pushed into late spring

Inhibition of MMP7 by siRNA or addition of rhMMP7 modulates cell response to oxaliplatin-induced apoptosis.

<p>A) MMP7 expression was inhibited by siRNA and then treated with different doses of oxaliplatin in order to analyze the effects of MMP7 inhibition in the response to oxaliplatin treatment. Cell viability was then assessed by MTS assay. To analyze the protective effects of MMP7 cells were treated with rhMMP7 (5 ng/ml) and with different doses of oxaliplatin. The effects on cell viability were determined by B) MTS or C) by annexin V/IP staining. D) The effects of MMPs inhibition on cell survival were determined in HT29 and RHT29 cells treated with 1,10-phenantroline monohydrate and then with different doses of oxaliplatin. It can be observed that cells response equally to oxaliplatin when MMPs are inhibited. Results are the mean±SEM for 3 samples per group. Values that are significantly different between control and treated group by ANOVA analysis are indicated by ***p<0.0001. <i>OXA</i>: oxaliplatin.</p

Modulation of cell surface Fas expression in HT29 and RHT29 cell lines.

<p>To analyze the expression specifically in the plasma membrane, Fas receptor was determined by A) FACS analysis or B) by immunohistochemistry in a cell line microarray, as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0004728#s4" target="_blank">Materials and Methods</a>. Fas mRNA expression under oxaliplatin treatment was also determined C) <i>in vitro</i> and <i>in vivo</i> for the HT29 and RHT29 cell lines, D) and also <i>in vitro</i> for the HCT116 p53^+/+ and HCT116p53^−/− cell lines. Results are represented as mean±SEM for 5 samples per group in the <i>in vitro</i> experiments (C and D) and for 10 animals per group in the <i>in vivo</i> determinations (C). Values that are significantly different between control and treated group by ANOVA analysis are indicated by *p<0.05,**p<0.001,***p<0.0001. <i>INH</i>: inhibitor (1,10-phenantroline monohydrate), <i>OXA</i>: oxaliplatin.</p

Analysis of HT29 and RHT29 growth rate.

<p>A) HT29 and RHT29 cells were subcutaneously inoculated into nude mice. Two weeks later, animals were treated with oxaliplatin 10 mg/kg once a week for 27 days. Results illustrate tumor volume and are represented as mean±SEM for 10 animals per group. The lower panel is a representative image of tumors obtained from animals treated or not with oxaliplatin. B) Growth kinetics was studied by colony formation assay. This representative image shows that HT29 cells are not able to grow under oxaliplatin treatment compared to RHT29 cell line. However, HT29 cells have a higher clonogenic capacity than RHT29 after 21 days of growth. <i>OXA</i>: oxaliplatin.</p

MMP7 shows a higher expression in the oxaliplatin resistant cell lines.

<p>A) Immunofluorescence (IF) staining of MMP7 in the non-resistant (HT29, HCT116 p53^+/+, HCT116 p53^−/−) and resistant cells (RHT29, RHCT116 p53^+/+, RHCT116 p53^−/−). MMP7 is strongly upregulated in the RHT29 and RHCT116 p53^−/− cell lines and its expression is also increased by oxaliplatin treatment. Levels of MMP7 were also analyzed by qPCR both <i>in vitro</i> (B) and <i>in vivo</i> (C) obtaining a similar pattern than in the IF assay. Results represent the mean±SEM for 6 samples in each group. Values that are significantly different between control and treated group by ANOVA analysis are indicated by *p<0.05,**p<0.001,***p<0.0001, and between different cell lines †††p<0.0001. <i>OXA</i>: oxaliplatin.</p

Modulation of FasL expression in HT29 and RHT29 cells lines.

<p>FasL expression was detected in HT29 and RHT29 cell lines treated or not with oxaliplatin 10 µM by A) FACS analysis of its surface expression or B) qPCR of its mRNA levels. C) NF-κB p65 was detected by Western Blot by the use of an antibody that recognizes the nuclear localization signal (NLS) epitope. Tubulin expression was used as endogenous control. D) Sensitivity of both cell lines to the inhibition of the NF-κB pathway was measured by an MTS assay in which cells were treated with different doses of BAY117085, a known inhibitor of IKK activation. Values that are significantly different between control and treated group by ANOVA analysis are indicated by ***p<0.0001. <i>OXA</i>: oxaliplatin.</p

Mechanism of acquisition of resistance to oxaliplatin-induced cell death.

<p>A) In a tumor with <i>low levels of MMP7</i> oxaliplatin is able to induce an upregulation of Fas receptor at the plasma membrane (1), allowing its activation by cells expressing FasL, such as cytotoxic cells or the surrender cells of the tumor (2). B) On the other hand, in cells with <i>higher levels of MMP7</i>, such as cells that have acquired resistance to oxaliplatin, the activation of Fas receptor is different. Oxaliplatin in these cells induce a stronger upregulation of FasL (1) that is shaded by MMP7 to generate sFasL (2). This sFasL binds to Fas probably inducing its activation (3). Fas receptor could, in turn, be shaded by MMP7, decreasing the availability of Fas at the cell surface (4), although the remaining Fas receptor could be activated by its ligand inducing pathways related to cell survival.</p

Analysis of the different pathways activated by Fas in HT29 and RHT29 cells.

<p>To determine the ability of Fas to induce cell death, cells were treated or not with A) oxaliplatin and B) MMP inhibitor for 24 hours, and then apoptosis was induced by the addition of the agonistic antibody CH11 at a final concentration of 150 ng/ml. Apoptosis induction by Fas activation was determined by annexin V/IP staining as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0004728#s4" target="_blank">Materials and Methods</a>. Results are represented as arbitrary units assuming that control value is 1 and are mean±SEM for 3 samples per group. C) Activation of MAPK pathways in cells treated or not with 150 ng/ml of CH11 for 24 hours was detected by Western blot analysis. Tubulin expression was used as an endogenous control. Values that are significantly different between control and treated group by ANOVA analysis are indicated by *p<0.05,**p<0.001, ***p<0.0001. <i>OXA</i>: oxaliplatin.</p

Flowchart of the study.

<p>Positron emission tomography-Computed tomography images show pathological uptake (SUVmax = 9.44) in a 4-cm pancreatic mass at baseline (A). There is no evidence of metabolic activity in the follow-up study (B), being considered as a complete response by EORTC criteria (<u>PERCIST criteria).</u></p

Correlation between positron emission tomography-computed tomography and computed tomography.

<p>Abbreviations:</p><p>PTV: Planning target volumes.</p><p>Post-tt: Post treatment.</p><p>PET: Positron emission tomography.</p><p>CT: Computed tomography.</p><p>PFS: Progression-free survival.</p