Angel Santos - Academia.edu (original) (raw)

Papers by Angel Santos

Research paper thumbnail of 15-Deoxy-D-12,14-prostaglandin J2 induces programmed cell death of breast cancer cells by a pleiotropic mechanism

Activation of peroxisome proliferator-activated receptor g (PPARg) has been found to induce cell ... more Activation of peroxisome proliferator-activated receptor g (PPARg) has been found to induce cell death in a variety of cells. In this regard, we reported recently that 15-deoxy-D-12,14 -prostaglandin J 2 (15dPG-J 2 ), a specific ligand of the nuclear receptor PPARg, inhibits proliferation and induces cellular differentiation and apoptosis in the breast cancer cell line MCF-7. In addition to PPARg activation other proteins, such as NF-kB and AP1, have been shown to be targets of 15dPG-J 2 . However, the mechanism by which 15dPG-J 2 triggers cell death is still elusive. Our results demonstrate that 15dPG-J 2 initiates breast cancer cell death via a very rapid and severe impairment of mitochondrial function, as revealed by a drop in mitochondrial membrane potential (DC m ), generation of reactive oxygen species (ROS) and a decrease in oxygen consumption. In addition, 15dPG-J 2 can also activate an intrinsic apoptotic pathway involving phosphatidyl serine externalization, caspase activation and cytochrome c release. Bcl-2 over-expression and zVADfmk, albeit preventing caspase activation, have no effect on 15dPG-J 2 -mediated mytochondrial dysfunction and loss of cell viability. In contrast, the addition of radical scavengers or rotenone, which prevent 15dPG-J 2 -induced ROS production, block the loss of cell viability induced by this prostaglandin. Finally, 15dPG-J 2 -induced cell death appears to involve disruption of the microtubule cytoskeletal network. Together, these results suggest that PG-J2-induced mitochondrial dysfunction and ROS production inevitably leads to death, with or without caspases.

Research paper thumbnail of The transcription factor early growth response factor-1 (EGR1) promotes apoptosis of neuroblastoma cells

Biochemical Journal, 2003

Early growth response factor-1 (EGR-1) is an immediate early gene, which is rapidly activated in ... more Early growth response factor-1 (EGR-1) is an immediate early gene, which is rapidly activated in quiescent cells by mitogens or in postmitotic neurons after depolarization. EGR-1 has been involved in diverse biological functions such as cell growth, differentiation and apoptosis. Here we report that enforced expression of the EGR-1 gene induces apoptosis, as determined by flow cytometry and terminal deoxynucleotidyl transferase-mediated dUTP-fluorescein nick-end labelling (TUNEL) analysis, in murine Neuro2A cells. In accordance with this role of EGR-1 in cell death, antisense oligonucleotides increase cell viability in cells cultured in the absence of serum. This apoptotic activity of the EGR-1 appears to be mediated by p73, a member of the p53 family of proteins, since an increase in the amount of p73 is observed in clones stably expressing the EGR-1 protein. We also observed an increase in the transcriptional activity of the mdm2 promoter in cells overexpressing EGR-1, which is paralleled by a marked decrease in the levels of p53 protein, therefore excluding a role of this protein in mediating EGR-1-induced apoptosis. Our results suggest that EGR-1 is an important factor involved in neuronal apoptosis.

Research paper thumbnail of Enhancement of BRCA1 gene expression by the peroxisome proliferator-activated receptor γ in the MCF7 breast cancer cell line

Oncogene, 2003

BRCA1 has been linked to the genetic susceptibility of a majority of familial breast and ovarian ... more BRCA1 has been linked to the genetic susceptibility of a majority of familial breast and ovarian cancers. Several lines of evidence indicate that BRCA1 is a tumor suppressor and its expression is downregulated in sporadic breast and ovarian cancer cases. Therefore, the identification of genes involved in the regulation of BRCA1 gene expression might lead to new insights into the pathogenesis and treatment of these tumors. Peroxisome proliferator-activated receptor gamma (PPARc) is a member of the nuclear receptor superfamily that has well-established roles in the regulation of adipocyte development and glucose homeostasis. More recently, it has been shown that ligands of PPARc have a potent antitumorigenic activity in breast cancer cells. In the present study we have found that two distinct ligands of PPARc; 15-deoxy-D-12,14 -prostaglandin J 2 (15dPG-J 2 ) and rosiglitazone, increase the levels of BRCA1 protein in human MCF-7 breast cancer cells. Immunofluorescence microscopy analysis showed that, after treatment with 15dPG-J 2 , the BRCA1 protein is mainly localized in the nucleus. Functional analysis by transient transfection of different 5 0 -flanking region fragments, as well as gel mobility shift assays and mutagenic analysis, suggests that the effects of 15dPG-J 2 and rosiglitazone are mediated through a functional DR1 located between the nucleotides À241 and À229, which is a canonical PPARc type response element. Our data suggest that PPARc is a crucial gene regulating BRCA1 gene expression and might therefore be important for the BRCA1 regulatory pathway involved in the pathogenesis of sporadic breast and ovarian cancer.

Research paper thumbnail of The spot 14 protein inhibits growth and induces differentiation and cell death of human MCF7 breast cancer cells

Biochemical Journal, 2005

The S14 (spot 14) gene encodes a protein that is predominantly expressed in lipogenic tissues, su... more The S14 (spot 14) gene encodes a protein that is predominantly expressed in lipogenic tissues, such as the liver, white and brown adipose tissues and the lactating mammary glands. Accumulated evidence suggests that S14 could play an important role in the induction of lipogenic enzymes. In humans, the S14 locus resides in the chromosome region 11q13, which is frequently amplified in breast tumours, and as a result, it has been suggested that this protein could play a role in the metabolism and growth of these kinds of tumours. In the present study, we have examined the effects of S14 overexpression in MCF-7 human breast cancer cells. We found that S14 causes (i) an inhibition of cell proliferation and of anchorage-independent growth, (ii) a marked reduction in the number of viable cells and (iii) the induction of differentiation and cell death of these cells. The inhibition of cell growth was associated with a decrease in the expression of cyclin D1 and a reduction of cyclin D1 promoter activity. Increased expression of S14 also caused the accumulation of cytochrome c in the cytosol and loss of mitochondrial membrane potential. These findings suggest that S14 may function as an important modulator of tumorigenesis in human breast by decreasing cell growth and inducing cell death and differentiation.

Research paper thumbnail of Direct Evidence of a Glucagon-Dependent Regulation of the Concentration of Glucagon Receptors in the Liver

European Journal of Biochemistry, 1982

Chronic treatment with exogenous glucagon in the rat results in a reduced concentration of glucag... more Chronic treatment with exogenous glucagon in the rat results in a reduced concentration of glucagon receptors in the liver. To determine if this is a direct effect of the hormone on its own receptors, the following experimental model in vilro was designed. Cultured rat hepatocytes were exposed to glucagon at 37 "C, washed and incubated with mono[1251]iodoglucagon (1251-glucagon), resulting in reduced binding of '251-glucagon. The magnitude of the diminution in binding was dependent on the concentration of glucagon present as well as on the duration of the exposure. Analysis of the data indicated that the decrease in binding of '251-glucagon was due to a loss of receptor numbers per cell rather than any change in receptor affinity for the hormone. Degradation of '251-glucagon during the incubation of cells in the presence or absence of glucagon was studied and did not account for the differences observed in binding. The binding of mono['2sI]iodoinsulin of these cells was not affected by glucagon, suggesting specificity of the phenomenon. Although the cells incubated with glucagon incorporate less ~-['~C]valine into hepatocyte proteins, this effect was smaller than the reduction in glucagon receptors. Exposure of the cells to cycloheximide (1 pg/ml) produced a progressive loss of glucagon receptors, and this effect was additive to the glucagon-induced receptor loss. This suggests that cycloheximide inhibited receptor synthesis while glucagon accelerated receptor loss. Glucagon-dependent reduction of glucagon receptors closely correlated with the decreased glucagon-stimulated production of CAMP. Furthermore, epinephrine (0.5 pM) stimulation of cyclic AMP production was similar in hepatocytes previously incubated with or without glucagon, indicating a specific role for glucagon in modifying target-cell sensitivity.

Research paper thumbnail of 15-Deoxy-D-12,14-prostaglandin J2 induces programmed cell death of breast cancer cells by a pleiotropic mechanism

Activation of peroxisome proliferator-activated receptor g (PPARg) has been found to induce cell ... more Activation of peroxisome proliferator-activated receptor g (PPARg) has been found to induce cell death in a variety of cells. In this regard, we reported recently that 15-deoxy-D-12,14 -prostaglandin J 2 (15dPG-J 2 ), a specific ligand of the nuclear receptor PPARg, inhibits proliferation and induces cellular differentiation and apoptosis in the breast cancer cell line MCF-7. In addition to PPARg activation other proteins, such as NF-kB and AP1, have been shown to be targets of 15dPG-J 2 . However, the mechanism by which 15dPG-J 2 triggers cell death is still elusive. Our results demonstrate that 15dPG-J 2 initiates breast cancer cell death via a very rapid and severe impairment of mitochondrial function, as revealed by a drop in mitochondrial membrane potential (DC m ), generation of reactive oxygen species (ROS) and a decrease in oxygen consumption. In addition, 15dPG-J 2 can also activate an intrinsic apoptotic pathway involving phosphatidyl serine externalization, caspase activation and cytochrome c release. Bcl-2 over-expression and zVADfmk, albeit preventing caspase activation, have no effect on 15dPG-J 2 -mediated mytochondrial dysfunction and loss of cell viability. In contrast, the addition of radical scavengers or rotenone, which prevent 15dPG-J 2 -induced ROS production, block the loss of cell viability induced by this prostaglandin. Finally, 15dPG-J 2 -induced cell death appears to involve disruption of the microtubule cytoskeletal network. Together, these results suggest that PG-J2-induced mitochondrial dysfunction and ROS production inevitably leads to death, with or without caspases.

Research paper thumbnail of The transcription factor early growth response factor-1 (EGR1) promotes apoptosis of neuroblastoma cells

Biochemical Journal, 2003

Early growth response factor-1 (EGR-1) is an immediate early gene, which is rapidly activated in ... more Early growth response factor-1 (EGR-1) is an immediate early gene, which is rapidly activated in quiescent cells by mitogens or in postmitotic neurons after depolarization. EGR-1 has been involved in diverse biological functions such as cell growth, differentiation and apoptosis. Here we report that enforced expression of the EGR-1 gene induces apoptosis, as determined by flow cytometry and terminal deoxynucleotidyl transferase-mediated dUTP-fluorescein nick-end labelling (TUNEL) analysis, in murine Neuro2A cells. In accordance with this role of EGR-1 in cell death, antisense oligonucleotides increase cell viability in cells cultured in the absence of serum. This apoptotic activity of the EGR-1 appears to be mediated by p73, a member of the p53 family of proteins, since an increase in the amount of p73 is observed in clones stably expressing the EGR-1 protein. We also observed an increase in the transcriptional activity of the mdm2 promoter in cells overexpressing EGR-1, which is paralleled by a marked decrease in the levels of p53 protein, therefore excluding a role of this protein in mediating EGR-1-induced apoptosis. Our results suggest that EGR-1 is an important factor involved in neuronal apoptosis.

Research paper thumbnail of Enhancement of BRCA1 gene expression by the peroxisome proliferator-activated receptor γ in the MCF7 breast cancer cell line

Oncogene, 2003

BRCA1 has been linked to the genetic susceptibility of a majority of familial breast and ovarian ... more BRCA1 has been linked to the genetic susceptibility of a majority of familial breast and ovarian cancers. Several lines of evidence indicate that BRCA1 is a tumor suppressor and its expression is downregulated in sporadic breast and ovarian cancer cases. Therefore, the identification of genes involved in the regulation of BRCA1 gene expression might lead to new insights into the pathogenesis and treatment of these tumors. Peroxisome proliferator-activated receptor gamma (PPARc) is a member of the nuclear receptor superfamily that has well-established roles in the regulation of adipocyte development and glucose homeostasis. More recently, it has been shown that ligands of PPARc have a potent antitumorigenic activity in breast cancer cells. In the present study we have found that two distinct ligands of PPARc; 15-deoxy-D-12,14 -prostaglandin J 2 (15dPG-J 2 ) and rosiglitazone, increase the levels of BRCA1 protein in human MCF-7 breast cancer cells. Immunofluorescence microscopy analysis showed that, after treatment with 15dPG-J 2 , the BRCA1 protein is mainly localized in the nucleus. Functional analysis by transient transfection of different 5 0 -flanking region fragments, as well as gel mobility shift assays and mutagenic analysis, suggests that the effects of 15dPG-J 2 and rosiglitazone are mediated through a functional DR1 located between the nucleotides À241 and À229, which is a canonical PPARc type response element. Our data suggest that PPARc is a crucial gene regulating BRCA1 gene expression and might therefore be important for the BRCA1 regulatory pathway involved in the pathogenesis of sporadic breast and ovarian cancer.

Research paper thumbnail of The spot 14 protein inhibits growth and induces differentiation and cell death of human MCF7 breast cancer cells

Biochemical Journal, 2005

The S14 (spot 14) gene encodes a protein that is predominantly expressed in lipogenic tissues, su... more The S14 (spot 14) gene encodes a protein that is predominantly expressed in lipogenic tissues, such as the liver, white and brown adipose tissues and the lactating mammary glands. Accumulated evidence suggests that S14 could play an important role in the induction of lipogenic enzymes. In humans, the S14 locus resides in the chromosome region 11q13, which is frequently amplified in breast tumours, and as a result, it has been suggested that this protein could play a role in the metabolism and growth of these kinds of tumours. In the present study, we have examined the effects of S14 overexpression in MCF-7 human breast cancer cells. We found that S14 causes (i) an inhibition of cell proliferation and of anchorage-independent growth, (ii) a marked reduction in the number of viable cells and (iii) the induction of differentiation and cell death of these cells. The inhibition of cell growth was associated with a decrease in the expression of cyclin D1 and a reduction of cyclin D1 promoter activity. Increased expression of S14 also caused the accumulation of cytochrome c in the cytosol and loss of mitochondrial membrane potential. These findings suggest that S14 may function as an important modulator of tumorigenesis in human breast by decreasing cell growth and inducing cell death and differentiation.

Research paper thumbnail of Direct Evidence of a Glucagon-Dependent Regulation of the Concentration of Glucagon Receptors in the Liver

European Journal of Biochemistry, 1982

Chronic treatment with exogenous glucagon in the rat results in a reduced concentration of glucag... more Chronic treatment with exogenous glucagon in the rat results in a reduced concentration of glucagon receptors in the liver. To determine if this is a direct effect of the hormone on its own receptors, the following experimental model in vilro was designed. Cultured rat hepatocytes were exposed to glucagon at 37 "C, washed and incubated with mono[1251]iodoglucagon (1251-glucagon), resulting in reduced binding of '251-glucagon. The magnitude of the diminution in binding was dependent on the concentration of glucagon present as well as on the duration of the exposure. Analysis of the data indicated that the decrease in binding of '251-glucagon was due to a loss of receptor numbers per cell rather than any change in receptor affinity for the hormone. Degradation of '251-glucagon during the incubation of cells in the presence or absence of glucagon was studied and did not account for the differences observed in binding. The binding of mono['2sI]iodoinsulin of these cells was not affected by glucagon, suggesting specificity of the phenomenon. Although the cells incubated with glucagon incorporate less ~-['~C]valine into hepatocyte proteins, this effect was smaller than the reduction in glucagon receptors. Exposure of the cells to cycloheximide (1 pg/ml) produced a progressive loss of glucagon receptors, and this effect was additive to the glucagon-induced receptor loss. This suggests that cycloheximide inhibited receptor synthesis while glucagon accelerated receptor loss. Glucagon-dependent reduction of glucagon receptors closely correlated with the decreased glucagon-stimulated production of CAMP. Furthermore, epinephrine (0.5 pM) stimulation of cyclic AMP production was similar in hepatocytes previously incubated with or without glucagon, indicating a specific role for glucagon in modifying target-cell sensitivity.