Figure 1.
Time course analysis of induction of apoptosis in neuroblastoma cell cultures by Retinoic Acid.
Percentage of dead cells in IMR-32, SH-SY5Y, SK-N-DZ, SK-N-Be(2) and SK-N-AS cell cultures either untreated (control cells) or treated with (A) 9-cis RA or (B) ATRA for 9 consecutive days (RA(9)) or for 5 days followed by 4 days in the absence of RA (RA(5)) was determined by TUNEL after 1, 3, 5, 7 or 9 days of treatment. Data are represented as mean ± SEM (n = 3). Asterisks represent significant differences to the control (*, p<0.05; **, p<0.01).
Figure 2.
Morphological alterations induced in neuroblastoma cell lines treated with retinoic acid.
Control cells (A, D, G, J and M) and cells treated with 9-cis RA (B, E, H, K and N) or ATRA (C, F, I, L and O) are shown. The images shown were taken at different time points: A: Day 1; B: Day 9; C: Day 5; D: Day 5; E: Day 3; F: Day 9; G: Day 1; H: Day 3; I: Day 5; J: Day 9; K: Day 9; L: Day 9; M: Day 1; N: Day 7; O: Day 5; All images were taken at 100X.
Figure 3.
Time course analysis of the differentiation process induced in neuroblastoma cell cultures by Retinoic Acid.
Expression levels of the oncogenes RET and MYCN were measured in IMR-32, SH-SY5Y, SK-N-DZ, SK-N-Be(2) and SK-N-AS cell cultures either untreated (control cells) or treated with 9-cis RA or ATRA for 9 consecutive days (RA(9)), or treated for 5 days followed by 4 days in the absence of RA (RA(5)). Expression levels were measured by qPCR as described in Materials and Methods and are shown relative to GAPDH. RET levels in cells either untreated (control) or treated with (A) 9-cis RA or (B) ATRA were determined at days 1, 3, 5, 7 and 9. MYCN levels in cells either untreated (control) or treated with (C) 9-cis RA or (D) ATRA were measured at day 9. Data are represented as mean ± SEM (n = 3). Asterisks represent significant differences to the control (*, p<0.05; **, p<0.01).
Table 1.
Genes of the apoptotic pathway overexpressed at least two times, or downregulated by 50%, after treatment of neuroblastoma cell lines with 9-cis RA.
Table 2.
Genes of the apoptotic pathway overexpressed at least two times, or downregulated by 50%, after treatment of neuroblastoma cell lines with ATRA.
Figure 4.
Edwards-Venn diagrams of genes of the apoptotic pathway whose expression was altered in neuroblastoma cell lines after RA treatment.
Genes overexpresed at least 2 times (up arrows) or downregulated by 50% (down arrows) after 5 days of treatment either with 9-cis RA or ATRA. (A) IMR-32, (B) SH-SY5Y, (C) SK-N-DZ and (D) SK-N-Be(2). The line represents the division among the modified genes that would induce and inhibit apoptosis.
Figure 5.
Effect of RA treatment over apoptotic and cell survival pathways in SH-SY5Y RA sensitive cell line.
In green colour are represented those genes whose expression has been augmented as a result of 9-cis RA (A) or ATRA (B) treatment, while in red are shown those whose expression has been decreased. Green arrows indicate the pathways that will be activated while red arrows indicate the pathways that will be inhibited as a consequence of the modifications of the genes that take part in these pathways. CM: Cell Membrane; Cyt: Cytoplasm; Mit: Mitochondrion; Nuc: Nucleus; P: Phosphorous.
Figure 6.
Effect of RA treatment over apoptotic and cell survival pathways in SK-N-Be(2) RA sensitive cell line.
In green colour are represented those genes whose expression has been augmented as a result of 9-cis RA (A) or ATRA (B) treatment, while in red are shown those whose expression has been decreased. Green arrows indicate the pathways that will be activated while red arrows indicate the pathways that will be inhibited as a consequence of the modifications of the genes that take part in these pathways. CM: Cell Membrane; Cyt: Cytoplasm; Mit: Mitochondrion; Nuc: Nucleus; P: Phosphorous.
Figure 7.
Effect of RA treatment over apoptotic and cell survival pathways in IMR-32 RA sensitive cell line.
In green colour are represented those genes whose expression has been augmented as a result of 9-cis RA (A) or ATRA (B) treatment, while in red are shown those whose expression has been decreased. Green arrows indicate the pathways that will be activated while red arrows indicate the pathways that will be inhibited as a consequence of the modifications of the genes that take part in these pathways. CM: Cell Membrane; Cyt: Cytoplasm; Mit: Mitochondrion; Nuc: Nucleus; P: Phosphorous.
Figure 8.
Effect of RA treatment over apoptotic and cell survival pathways in SK-N-DZ RA sensitive cell line.
In green colour are represented those genes whose expression has been augmented as a result of 9-cis RA (A) or ATRA (B) treatment, while in red are shown those whose expression has been decreased. Green arrows indicate the pathways that will be activated while red arrows indicate the pathways that will be inhibited as a consequence of the modifications of the genes that take part in these pathways. CM: Cell Membrane; Cyt: Cytoplasm; Mit: Mitochondrion; Nuc: Nucleus; P: Phosphorous.
Figure 9.
Overlapping pattern of genes modified by RA treatment of neuroblastoma cell lines.
Edwards-Venn diagrams showing the genes overexpressed at least 2 times (up arrows) or downregulated by 50% (down arrows) after 5 days of RA treatment of the corresponding cell line. (A) Genes modified by 9-cis RA. (B) Genes modified by ATRA.