16 research outputs found
Displayed correlation between gene expression profiles and submicroscopic alterations in response to cetuximab, gefitinib and EGF in human colon cancer cell lines
Background: EGFR is frequently overexpressed in colon cancer. We characterized HT-29 and
Caco-2, human colon cancer cell lines, untreated and treated with cetuximab or gefitinib alone and
in combination with EGF.
Methods: Cell growth was determined using a variation on the MTT assay. Cell-cycle analysis was
conducted by flow cytometry. Immunohistochemistry was performed to evaluate EGFR expression
and scanning electron microscopy (SEM) evidenced the ultrastructural morphology. Gene
expression profiling was performed using hybridization of the microarray Ocimum Pan Human 40
K array A.
Results: Caco-2 and HT-29 were respectively 66.25 and 59.24 % in G0/G1. They maintained this
level of cell cycle distribution after treatment, suggesting a predominantly differentiated state.
Treatment of Caco-2 with EGF or the two EGFR inhibitors produced a significant reduction in their
viability. SEM clearly showed morphological cellular transformations in the direction of cellular death in both cell lines treated with EGFR inhibitors. HT-29 and Caco-2 displayed an important
reduction of the microvilli (which also lose their erect position in Caco-2), possibly invalidating
microvilli absorption function. HT-29 treated with cetuximab lost their boundary contacts and
showed filipodi; when treated with gefitinib, they showed some vesicles: generally membrane
reshaping is evident. Both cell lines showed a similar behavior in terms of on/off switched genes
upon treatment with cetuximab. The gefitinib global gene expression pattern was different for the
2 cell lines; gefitinib treatment induced more changes, but directly correlated with EGF treatment.
In cetuximab or gefitinib plus EGF treatments there was possible summation of the morphological
effects: cells seemed more weakly affected by the transformation towards apoptosis. The genes
appeared to be less stimulated than for single drug cases.
Conclusion: This is the first study to have systematically investigated the effect of cetuximab or
gefitinib, alone and in combination with EGF, on human colon cancer cell lines. The EGFR inhibitors
have a weaker effect in the presence of EGF that binds EGFR. Cetuximab treatment showed an
expression pattern that inversely correlates with EGF treatment. We found interesting cytomorphological
features closely relating to gene expression profile. Both drugs have an effect on
differentiation towards cellular death
Inhibition of Effector Function but Not T Cell Activation and Increase in FoxP3 Expression in T Cells Differentiated in the Presence of PP14
Background: T-helper polarization of naïve T cells is determined by a complex mechanism that involves many factors, eventually leading to activation of Th1, Th2, or Th17 responses or alternatively the generation of regulatory T cells. Placental Protein 14 (PP14) is a 28 kDa glycoprotein highly secreted in early pregnancy that is able to desensitize T cell receptor (TCR) signaling and modulate T cell activation. Methodology/Principal Findings: Prolonged antigen-specific stimulation of T cells in the presence of PP14 resulted in an impaired secretion of IFN-c, IL-5 and IL-17 upon restimulation, although the cells proliferated and expressed activation markers. Furthermore, the generation of regulatory CD4 + CD25 high Foxp3 + T cells was induced in the presence of PP14, in both antigen-specific as well as polyclonal stimulation. In accordance with previous reports, we found that the induction of FoxP3 expression by PP14 is accompanied by down regulation of the PI3K-mTOR signaling pathway. Conclusions/Significance: These data suggest that PP14 arrests T cells in a unique activated state that is not accompanied with the acquisition of effector function, together with promoting the generation of regulatory T cells. Taken together, our results may elucidate the role of PP14 in supporting immune tolerance in pregnancy by reducing T cell effector function