irene pino - Academia.edu (original) (raw)

Papers by irene pino

Research paper thumbnail of The dynamic DNA methylomes of double-stranded DNA viruses associated with human cancer

Research paper thumbnail of Different phospholipase-C-coupled receptors differentially regulate capacitative and non-capacitative Ca2+ entry in A7r5 cells

Biochemical Journal, 2005

Several receptors, including those for AVP (Arg 8 -vasopressin) and 5-HT (5-hydroxytryptamine), s... more Several receptors, including those for AVP (Arg 8 -vasopressin) and 5-HT (5-hydroxytryptamine), share an ability to stimulate PLC (phospholipase C) and so production of IP 3 (inositol 1,4,5trisphosphate) and DAG (diacylglycerol) in A7r5 vascular smooth muscle cells. Our previous analysis of the effects of AVP on Ca 2+ entry Biochem. J. 370, [439][440][441][442][443][444][445][446][447][448] showed that arachidonic acid released from DAG stimulated NO synthase. NO then stimulated an NCCE (non-capacitative Ca 2+ entry) pathway, and, via cGMP and protein kinase G, it inhibited CCE (capacitative Ca 2+ entry). This reciprocal regulation ensured that, in the presence of AVP, all Ca 2+ entry occurred via NCCE to be followed by a transient activation of CCE only when AVP was removed [Moneer and Taylor Biochem. J. 362, 13-21]. We confirm that, in the presence of AVP, all Ca 2+ entry occurs via NCCE, but 5-HT, despite activating PLC and evoking release of Ca 2+ from intracellular stores, stimulates Ca 2+ entry only via CCE. We conclude that two PLC-coupled receptors differentially regulate CCE and NCCE. We also address evidence that, in some A7r5 cells lines, AVP fails either to stimulate NCCE or inhibit CCE Biochem. J. 388, [237][238][239][240][241][242][243][244]. Quantitative PCR analysis suggests that these cells predominantly express TRPC1 (transient receptor potential canonical 1), whereas cells in which AVP reciprocally regulates CCE and NCCE express a greater variety of TRPC subtypes (TRPC1 = 6 > 2 > 3).

Research paper thumbnail of Different phospholipase-C-coupled receptors differentially regulate capacitative and non-capacitative Ca2+ entry in A7r5 cells

Biochemical Journal, 2005

Several receptors, including those for AVP (Arg 8 -vasopressin) and 5-HT (5-hydroxytryptamine), s... more Several receptors, including those for AVP (Arg 8 -vasopressin) and 5-HT (5-hydroxytryptamine), share an ability to stimulate PLC (phospholipase C) and so production of IP 3 (inositol 1,4,5trisphosphate) and DAG (diacylglycerol) in A7r5 vascular smooth muscle cells. Our previous analysis of the effects of AVP on Ca 2+ entry Biochem. J. 370, [439][440][441][442][443][444][445][446][447][448] showed that arachidonic acid released from DAG stimulated NO synthase. NO then stimulated an NCCE (non-capacitative Ca 2+ entry) pathway, and, via cGMP and protein kinase G, it inhibited CCE (capacitative Ca 2+ entry). This reciprocal regulation ensured that, in the presence of AVP, all Ca 2+ entry occurred via NCCE to be followed by a transient activation of CCE only when AVP was removed [Moneer and Taylor Biochem. J. 362, 13-21]. We confirm that, in the presence of AVP, all Ca 2+ entry occurs via NCCE, but 5-HT, despite activating PLC and evoking release of Ca 2+ from intracellular stores, stimulates Ca 2+ entry only via CCE. We conclude that two PLC-coupled receptors differentially regulate CCE and NCCE. We also address evidence that, in some A7r5 cells lines, AVP fails either to stimulate NCCE or inhibit CCE Biochem. J. 388, [237][238][239][240][241][242][243][244]. Quantitative PCR analysis suggests that these cells predominantly express TRPC1 (transient receptor potential canonical 1), whereas cells in which AVP reciprocally regulates CCE and NCCE express a greater variety of TRPC subtypes (TRPC1 = 6 > 2 > 3).

Research paper thumbnail of O-303 Altered expression of alphaCP-4, a putative tumor suppressor gene at 3p21, and its alternatively spliced variant alphaCP-4a in lung cancer

Lung Cancer, 2003

Lung Cancer is the leading cause of cancer death in men and women in the U.S. The cyclooxygenase ... more Lung Cancer is the leading cause of cancer death in men and women in the U.S. The cyclooxygenase (COX) pathway has a role in colon cancer chemoprevention and a potential role in lung cancer. COX inhibition reduces lung cancer multiplicity in a murine model and we have shown that prostacyclin synthase (PGIPS) overexpression protects against tumor development in a transgenic mouse model. We hypothesize that transcriptional regulation plays a pivotal role in lung tumorigenesis and that switching of the eicosanoid profile from predominantly PG12 to predominantly PGE2 may contribute to angiogenesis or tumor growth.PGIPS expression in human non-small cell lung cancer (NSCLC) samples and normal lung was evaluated by: 1) immunohistochemistry 2) quantitative PCR 3) Western analysis and 4) eicosanoid levels of 6-keto PGFI-alpha (the stable metabolite of prostacyclin) and PGEP. To determine transcriptional regulation, a reporter construct encoding the promoter region of PGl2S (3014 base pairs) ligated to a luciferase reporter was used for transfection. The promoter activity was evaluated following exposure to increasing levels of serum or platelet derived growth factor (PDGF) and in the setting of gain of function Ras. In addition, truncated constructs were made in order to identify minimal regions responsible for basal activity and regulation by these factors. Tumors produced very low levels of 6-Keto PGFI -alpha and had decreased PGIPS mRNA and protein expression by immunohistochemistry when compared to normal surrounding lung tissue. Cell lines transfected with PGl2S promoter exhibited augmented promoter activity when exposed to serum. In contrast, cotransfection with platelet derived growth factor (PDGF) or gain of function Ras resulted in significant promoter suppression. Promoter activity results were validated by evidence of suppression of PGIS protein as well as levels of the PG12 metabolite 6-Keto 1 alpha. These studies suggest an association between decreased expression of PGl2S and the development of NSCLC. In further studies, tumors will be evaluated by tissue microarray to determine whether 1) relative absence of PGl2S is associated with lung tumors of various histologies and 2) whether PGl2S may serve as a marker of tumor aggressiveness or survival. Suppression of PGl2S, through inhibition of transfection suggests that PG12 has an antimitogen/antiproliferative effect in lung cancers.

Research paper thumbnail of Hyperplasia of Alveolar Neuroendocrine Cells in Rat Lung Carcinogenesis by Silica with Selective Expression of Proadrenomedullin-Derived Peptides and Amidating Enzymes

Laboratory Investigation, 2001

Pulmonary neuroendocrine (NE) cells are found as clusters called neuroepithelial bodies (NEBs) or... more Pulmonary neuroendocrine (NE) cells are found as clusters called neuroepithelial bodies (NEBs) or as single cells scattered in the respiratory epithelium. They express a variety of bioactive peptides, and they are thought to be the origin of NE lung tumors. Proadrenomedullin N-terminal 20 peptide (PAMP) is a peptide derived from the same precursor as adrenomedullin (AM). AM and PAMP are C-terminally amidated during their processing by a well-characterized amidating enzyme, peptidylglycine alpha-amidating monooxygenase (PAM). We explored AM, PAMP, and PAM expression as markers for NE hyperplasia in three rodent species (Fischer 344 rats, Syrian golden hamsters, and A/J mice) after a single intratracheal instillation of crystalline silica (quartz), which was previously found to induce different reactions in the three species. Rats developed a marked silicosis, with alveolar and bronchiolar hyperplasia and formation of peripheral lung epithelial tumors. Mice developed a moderate degree of silicosis, but not epithelial hyperplasia or tumors. Hamsters showed dust-storage lesions, but not silicosis or tumors. NE cells were immunolabeled for calcitonin gene-related peptide (CGRP), AM, PAMP, and PAM in serial sections of each lung. The numbers of positive NEBs per lung area and positive cells per NEB were quantified. A marked hyperplastic reaction in the NEBs of silica treated rats occurred only in alveolar NEBs, but not in bronchiolar NEBs. From Month 11 onwards, there were marked differences in the number of alveolar NEBs per section and in the number of cells per alveolar NEB immunoreactive for CGRP. No hyperplastic NE cell reaction was observed in silica-treated mice and hamsters. Significant PAMP and PAM expression was seen only in rat hyperplastic alveolar and in bronchiolar NEBs from Month 11 onwards. In E18, rat fetal lung NEBs were found to be strongly positive for PAMP and PAM.

Research paper thumbnail of Relative amounts of antagonistic splicing factors, hnRNP A1 and ASF/SF2, change during neoplastic lung growth: Implications for pre-mRNA processing

Molecular Carcinogenesis, 2004

Pre-mRNA processing is an important mechanism for globally modifying cellular protein composition... more Pre-mRNA processing is an important mechanism for globally modifying cellular protein composition during tumorigenesis. To understand this process during lung cancer, expression of two key pre-mRNA alternative splicing factors was compared in a mouse model of early lung carcinogenesis and during regenerative growth following reversible lung injury. Heterogeneous nuclear ribonucleoprotein (hnRNP) A1 and alternative splicing factor/splicing factor 2 (ASF/SF2) act antagonistically to modulate splice site selection. Both hnRNP A1 and ASF/SF2 contents rose in adenomas and during injury-induced hyperplasia compared to control lungs, as measured by immunoblotting. While both proteins increased similarly during compensatory hyperplasia, hnRNP A1 increased to a much greater extent than ASF/SF2 in tumors, resulting in a 6-fold increase of the hnRNP A1 to ASF/SF2 ratio. Immunohistochemical analysis showed that hnRNP A1 localized exclusively within tumor nuclei, while ASF/SF2 appeared in cytoplasm and/or nuclei, depending on the growth pattern of the tumor cells. We also demonstrated cancer-associated changes in the pre-mRNA alternative splicing of CD44, a membrane glycoprotein involved in cell-cell and cell-extracellular matrix interactions. hnRNP A1 and ASF/SF2 expression is thus differentially altered in neoplastic lung cells by mechanisms that do not strictly arise from increased cell division. These changes are influenced by tumor histology and may be associated with production of variant CD44 mRNA isoforms. © 2004 Wiley-Liss, Inc.

Research paper thumbnail of Histone H3 and H4 Modification Profiles in a Rett Syndrome Mouse Model

Epigenetics, 2007

Rett syndrome (RTT) is a complex neurodevelopmental disorder that has been associated with mutati... more Rett syndrome (RTT) is a complex neurodevelopmental disorder that has been associated with mutations of methyl-CpG binding protein 2 (MeCP2). MeCP2 acts as a transcriptional repressor and binds to histone modifier proteins, which prompted us to wonder whether MeCP2 disruption affects global histone modification patterns. Taking a two-fold approach of using high-performance capillary electrophoresis (HPCE) and western blot, we analyzed the acetylation and methylation status of histones H3 and H4 in a mouse model of RTT where the MeCP2 locus is genetically disrupted. The comparison of cortex, midbrain and cerebellum in wild-type and MeCP2-knock out mice did not reveal any significant difference in the global H3 and H4 histone modification patterns. Our results suggest that MeCP2 deficiency involves local and gene-specific chromatin changes rather than massive histone modification changes.

Research paper thumbnail of Altered expression of adhesion molecules and epithelial–mesenchymal transition in silica-induced rat lung carcinogenesis

Laboratory Investigation, 2004

Loss of the epithelial phenotype and disruption of adhesion molecules is a hallmark in the epithe... more Loss of the epithelial phenotype and disruption of adhesion molecules is a hallmark in the epithelialmesenchymal transition (EMT) reported in several types of cancer. Most of the studies about the relevance of adhesion and junction molecules in lung cancer have been performed using established tumors or in vitro models. The sequential molecular events leading to EMT during lung cancer progression are still not well understood. We have used a rat model for multistep lung carcinogenesis to study the status of adherens and tight junction proteins and mesenchymal markers during EMT. After silica-induced chronic inflammation, rats sequentially develop epithelial hyperplasia, preneoplastic lesions, and tumors such as adenocarcinomas and squamous cell carcinomas. In comparison with normal and hyperplastic bronchiolar epithelium and with hyperplastic alveolar type II cells, the expression levels of E-cadherin, a-catenin and b-catenin were significantly reduced in adenomatoid preneoplastic lesions and in late tumors. The loss of E-cadherin in tumors was associated with its promoter hypermethylation. a-and b-catenin dysregulation lead to cytoplasmic accumulation in some carcinomas. No nuclear b-catenin localization was found at any stage of any preneoplastic or neoplastic lesion. Zonula occludens protein-1 was markedly decreased in 66% of adenocarcinomas and in 100% squamous cell carcinomas. The mesenchymal-associated proteins N-cadherin and vimentin were analyzed as markers for EMT. N-cadherin was de novo expressed in 32% of adenocarcinomas and 33% of squamous cell carcinomas. Vimentin-positive tumor cells were found in 35% of adenocarcinomas and 88% of squamous cell carcinomas. Mesenchymal markers were absent in precursor lesions, both hyperplastic and adenomatoid. The present results show that silica-induced rat lung carcinogenesis is a good model to study EMT in vivo, and also provide in vivo evidence suggesting that the changes in cell-cell adhesion molecules are an early event in lung carcinogenesis, while EMT occurs at a later stage.

Research paper thumbnail of Altered patterns of expression of members of the heterogeneous nuclear ribonucleoprotein (hnRNP) family in lung cancer

Lung Cancer, 2003

hnRNP A2/B1 has been suggested as a useful early detection marker for lung carcinoma. hnRNP A2/B1... more hnRNP A2/B1 has been suggested as a useful early detection marker for lung carcinoma. hnRNP A2/B1 is a member of a large family of heterogeneous nuclear ribonucleoproteins (hnRNP proteins) involved in a variety of functions, including regulation of transcription, mRNA metabolism, and translation. In lung cancer, we have evaluated the expression and cellular localization of several members of the hnRNP family, hnRNP A1, A2, B1, C1, C2 and K. 16 cell lines (SCLC and NSCLC) and biopsies from 32 lung cancer patients were analyzed. Our results suggest that, besides hnRNP A2/B1, the expression of other members of the hnRNP family is altered both in SCLC and NSCLC. In the biopsies, negative or low expression of the hnRNP proteins analyzed was observed in normal epithelial cells whereas lung cancer cells showed highly intense nuclear or cytoplasmic immunolocalization. In all the lung cancer cell lines, the mRNA for all the hnRNP proteins was detected. In general, higher levels of hnRNP mRNAs were found in SCLC as compared with NSCLC. Our results also suggest that the expression and processing of each hnRNP protein in lung cancer is independently regulated and is not exclusively related to proliferation status. In SCLC cell lines, hnRNP A1 protein expression correlated with that of Bcl-x L . In the lung cancer cell lines, hnRNP K protein localization varied with the cellular confluence.

Research paper thumbnail of The dynamic DNA methylomes of double-stranded DNA viruses associated with human cancer

Research paper thumbnail of Different phospholipase-C-coupled receptors differentially regulate capacitative and non-capacitative Ca2+ entry in A7r5 cells

Biochemical Journal, 2005

Several receptors, including those for AVP (Arg 8 -vasopressin) and 5-HT (5-hydroxytryptamine), s... more Several receptors, including those for AVP (Arg 8 -vasopressin) and 5-HT (5-hydroxytryptamine), share an ability to stimulate PLC (phospholipase C) and so production of IP 3 (inositol 1,4,5trisphosphate) and DAG (diacylglycerol) in A7r5 vascular smooth muscle cells. Our previous analysis of the effects of AVP on Ca 2+ entry Biochem. J. 370, [439][440][441][442][443][444][445][446][447][448] showed that arachidonic acid released from DAG stimulated NO synthase. NO then stimulated an NCCE (non-capacitative Ca 2+ entry) pathway, and, via cGMP and protein kinase G, it inhibited CCE (capacitative Ca 2+ entry). This reciprocal regulation ensured that, in the presence of AVP, all Ca 2+ entry occurred via NCCE to be followed by a transient activation of CCE only when AVP was removed [Moneer and Taylor Biochem. J. 362, 13-21]. We confirm that, in the presence of AVP, all Ca 2+ entry occurs via NCCE, but 5-HT, despite activating PLC and evoking release of Ca 2+ from intracellular stores, stimulates Ca 2+ entry only via CCE. We conclude that two PLC-coupled receptors differentially regulate CCE and NCCE. We also address evidence that, in some A7r5 cells lines, AVP fails either to stimulate NCCE or inhibit CCE Biochem. J. 388, [237][238][239][240][241][242][243][244]. Quantitative PCR analysis suggests that these cells predominantly express TRPC1 (transient receptor potential canonical 1), whereas cells in which AVP reciprocally regulates CCE and NCCE express a greater variety of TRPC subtypes (TRPC1 = 6 > 2 > 3).

Research paper thumbnail of Different phospholipase-C-coupled receptors differentially regulate capacitative and non-capacitative Ca2+ entry in A7r5 cells

Biochemical Journal, 2005

Several receptors, including those for AVP (Arg 8 -vasopressin) and 5-HT (5-hydroxytryptamine), s... more Several receptors, including those for AVP (Arg 8 -vasopressin) and 5-HT (5-hydroxytryptamine), share an ability to stimulate PLC (phospholipase C) and so production of IP 3 (inositol 1,4,5trisphosphate) and DAG (diacylglycerol) in A7r5 vascular smooth muscle cells. Our previous analysis of the effects of AVP on Ca 2+ entry Biochem. J. 370, [439][440][441][442][443][444][445][446][447][448] showed that arachidonic acid released from DAG stimulated NO synthase. NO then stimulated an NCCE (non-capacitative Ca 2+ entry) pathway, and, via cGMP and protein kinase G, it inhibited CCE (capacitative Ca 2+ entry). This reciprocal regulation ensured that, in the presence of AVP, all Ca 2+ entry occurred via NCCE to be followed by a transient activation of CCE only when AVP was removed [Moneer and Taylor Biochem. J. 362, 13-21]. We confirm that, in the presence of AVP, all Ca 2+ entry occurs via NCCE, but 5-HT, despite activating PLC and evoking release of Ca 2+ from intracellular stores, stimulates Ca 2+ entry only via CCE. We conclude that two PLC-coupled receptors differentially regulate CCE and NCCE. We also address evidence that, in some A7r5 cells lines, AVP fails either to stimulate NCCE or inhibit CCE Biochem. J. 388, [237][238][239][240][241][242][243][244]. Quantitative PCR analysis suggests that these cells predominantly express TRPC1 (transient receptor potential canonical 1), whereas cells in which AVP reciprocally regulates CCE and NCCE express a greater variety of TRPC subtypes (TRPC1 = 6 > 2 > 3).

Research paper thumbnail of O-303 Altered expression of alphaCP-4, a putative tumor suppressor gene at 3p21, and its alternatively spliced variant alphaCP-4a in lung cancer

Lung Cancer, 2003

Lung Cancer is the leading cause of cancer death in men and women in the U.S. The cyclooxygenase ... more Lung Cancer is the leading cause of cancer death in men and women in the U.S. The cyclooxygenase (COX) pathway has a role in colon cancer chemoprevention and a potential role in lung cancer. COX inhibition reduces lung cancer multiplicity in a murine model and we have shown that prostacyclin synthase (PGIPS) overexpression protects against tumor development in a transgenic mouse model. We hypothesize that transcriptional regulation plays a pivotal role in lung tumorigenesis and that switching of the eicosanoid profile from predominantly PG12 to predominantly PGE2 may contribute to angiogenesis or tumor growth.PGIPS expression in human non-small cell lung cancer (NSCLC) samples and normal lung was evaluated by: 1) immunohistochemistry 2) quantitative PCR 3) Western analysis and 4) eicosanoid levels of 6-keto PGFI-alpha (the stable metabolite of prostacyclin) and PGEP. To determine transcriptional regulation, a reporter construct encoding the promoter region of PGl2S (3014 base pairs) ligated to a luciferase reporter was used for transfection. The promoter activity was evaluated following exposure to increasing levels of serum or platelet derived growth factor (PDGF) and in the setting of gain of function Ras. In addition, truncated constructs were made in order to identify minimal regions responsible for basal activity and regulation by these factors. Tumors produced very low levels of 6-Keto PGFI -alpha and had decreased PGIPS mRNA and protein expression by immunohistochemistry when compared to normal surrounding lung tissue. Cell lines transfected with PGl2S promoter exhibited augmented promoter activity when exposed to serum. In contrast, cotransfection with platelet derived growth factor (PDGF) or gain of function Ras resulted in significant promoter suppression. Promoter activity results were validated by evidence of suppression of PGIS protein as well as levels of the PG12 metabolite 6-Keto 1 alpha. These studies suggest an association between decreased expression of PGl2S and the development of NSCLC. In further studies, tumors will be evaluated by tissue microarray to determine whether 1) relative absence of PGl2S is associated with lung tumors of various histologies and 2) whether PGl2S may serve as a marker of tumor aggressiveness or survival. Suppression of PGl2S, through inhibition of transfection suggests that PG12 has an antimitogen/antiproliferative effect in lung cancers.

Research paper thumbnail of Hyperplasia of Alveolar Neuroendocrine Cells in Rat Lung Carcinogenesis by Silica with Selective Expression of Proadrenomedullin-Derived Peptides and Amidating Enzymes

Laboratory Investigation, 2001

Pulmonary neuroendocrine (NE) cells are found as clusters called neuroepithelial bodies (NEBs) or... more Pulmonary neuroendocrine (NE) cells are found as clusters called neuroepithelial bodies (NEBs) or as single cells scattered in the respiratory epithelium. They express a variety of bioactive peptides, and they are thought to be the origin of NE lung tumors. Proadrenomedullin N-terminal 20 peptide (PAMP) is a peptide derived from the same precursor as adrenomedullin (AM). AM and PAMP are C-terminally amidated during their processing by a well-characterized amidating enzyme, peptidylglycine alpha-amidating monooxygenase (PAM). We explored AM, PAMP, and PAM expression as markers for NE hyperplasia in three rodent species (Fischer 344 rats, Syrian golden hamsters, and A/J mice) after a single intratracheal instillation of crystalline silica (quartz), which was previously found to induce different reactions in the three species. Rats developed a marked silicosis, with alveolar and bronchiolar hyperplasia and formation of peripheral lung epithelial tumors. Mice developed a moderate degree of silicosis, but not epithelial hyperplasia or tumors. Hamsters showed dust-storage lesions, but not silicosis or tumors. NE cells were immunolabeled for calcitonin gene-related peptide (CGRP), AM, PAMP, and PAM in serial sections of each lung. The numbers of positive NEBs per lung area and positive cells per NEB were quantified. A marked hyperplastic reaction in the NEBs of silica treated rats occurred only in alveolar NEBs, but not in bronchiolar NEBs. From Month 11 onwards, there were marked differences in the number of alveolar NEBs per section and in the number of cells per alveolar NEB immunoreactive for CGRP. No hyperplastic NE cell reaction was observed in silica-treated mice and hamsters. Significant PAMP and PAM expression was seen only in rat hyperplastic alveolar and in bronchiolar NEBs from Month 11 onwards. In E18, rat fetal lung NEBs were found to be strongly positive for PAMP and PAM.

Research paper thumbnail of Relative amounts of antagonistic splicing factors, hnRNP A1 and ASF/SF2, change during neoplastic lung growth: Implications for pre-mRNA processing

Molecular Carcinogenesis, 2004

Pre-mRNA processing is an important mechanism for globally modifying cellular protein composition... more Pre-mRNA processing is an important mechanism for globally modifying cellular protein composition during tumorigenesis. To understand this process during lung cancer, expression of two key pre-mRNA alternative splicing factors was compared in a mouse model of early lung carcinogenesis and during regenerative growth following reversible lung injury. Heterogeneous nuclear ribonucleoprotein (hnRNP) A1 and alternative splicing factor/splicing factor 2 (ASF/SF2) act antagonistically to modulate splice site selection. Both hnRNP A1 and ASF/SF2 contents rose in adenomas and during injury-induced hyperplasia compared to control lungs, as measured by immunoblotting. While both proteins increased similarly during compensatory hyperplasia, hnRNP A1 increased to a much greater extent than ASF/SF2 in tumors, resulting in a 6-fold increase of the hnRNP A1 to ASF/SF2 ratio. Immunohistochemical analysis showed that hnRNP A1 localized exclusively within tumor nuclei, while ASF/SF2 appeared in cytoplasm and/or nuclei, depending on the growth pattern of the tumor cells. We also demonstrated cancer-associated changes in the pre-mRNA alternative splicing of CD44, a membrane glycoprotein involved in cell-cell and cell-extracellular matrix interactions. hnRNP A1 and ASF/SF2 expression is thus differentially altered in neoplastic lung cells by mechanisms that do not strictly arise from increased cell division. These changes are influenced by tumor histology and may be associated with production of variant CD44 mRNA isoforms. © 2004 Wiley-Liss, Inc.

Research paper thumbnail of Histone H3 and H4 Modification Profiles in a Rett Syndrome Mouse Model

Epigenetics, 2007

Rett syndrome (RTT) is a complex neurodevelopmental disorder that has been associated with mutati... more Rett syndrome (RTT) is a complex neurodevelopmental disorder that has been associated with mutations of methyl-CpG binding protein 2 (MeCP2). MeCP2 acts as a transcriptional repressor and binds to histone modifier proteins, which prompted us to wonder whether MeCP2 disruption affects global histone modification patterns. Taking a two-fold approach of using high-performance capillary electrophoresis (HPCE) and western blot, we analyzed the acetylation and methylation status of histones H3 and H4 in a mouse model of RTT where the MeCP2 locus is genetically disrupted. The comparison of cortex, midbrain and cerebellum in wild-type and MeCP2-knock out mice did not reveal any significant difference in the global H3 and H4 histone modification patterns. Our results suggest that MeCP2 deficiency involves local and gene-specific chromatin changes rather than massive histone modification changes.

Research paper thumbnail of Altered expression of adhesion molecules and epithelial–mesenchymal transition in silica-induced rat lung carcinogenesis

Laboratory Investigation, 2004

Loss of the epithelial phenotype and disruption of adhesion molecules is a hallmark in the epithe... more Loss of the epithelial phenotype and disruption of adhesion molecules is a hallmark in the epithelialmesenchymal transition (EMT) reported in several types of cancer. Most of the studies about the relevance of adhesion and junction molecules in lung cancer have been performed using established tumors or in vitro models. The sequential molecular events leading to EMT during lung cancer progression are still not well understood. We have used a rat model for multistep lung carcinogenesis to study the status of adherens and tight junction proteins and mesenchymal markers during EMT. After silica-induced chronic inflammation, rats sequentially develop epithelial hyperplasia, preneoplastic lesions, and tumors such as adenocarcinomas and squamous cell carcinomas. In comparison with normal and hyperplastic bronchiolar epithelium and with hyperplastic alveolar type II cells, the expression levels of E-cadherin, a-catenin and b-catenin were significantly reduced in adenomatoid preneoplastic lesions and in late tumors. The loss of E-cadherin in tumors was associated with its promoter hypermethylation. a-and b-catenin dysregulation lead to cytoplasmic accumulation in some carcinomas. No nuclear b-catenin localization was found at any stage of any preneoplastic or neoplastic lesion. Zonula occludens protein-1 was markedly decreased in 66% of adenocarcinomas and in 100% squamous cell carcinomas. The mesenchymal-associated proteins N-cadherin and vimentin were analyzed as markers for EMT. N-cadherin was de novo expressed in 32% of adenocarcinomas and 33% of squamous cell carcinomas. Vimentin-positive tumor cells were found in 35% of adenocarcinomas and 88% of squamous cell carcinomas. Mesenchymal markers were absent in precursor lesions, both hyperplastic and adenomatoid. The present results show that silica-induced rat lung carcinogenesis is a good model to study EMT in vivo, and also provide in vivo evidence suggesting that the changes in cell-cell adhesion molecules are an early event in lung carcinogenesis, while EMT occurs at a later stage.

Research paper thumbnail of Altered patterns of expression of members of the heterogeneous nuclear ribonucleoprotein (hnRNP) family in lung cancer

Lung Cancer, 2003

hnRNP A2/B1 has been suggested as a useful early detection marker for lung carcinoma. hnRNP A2/B1... more hnRNP A2/B1 has been suggested as a useful early detection marker for lung carcinoma. hnRNP A2/B1 is a member of a large family of heterogeneous nuclear ribonucleoproteins (hnRNP proteins) involved in a variety of functions, including regulation of transcription, mRNA metabolism, and translation. In lung cancer, we have evaluated the expression and cellular localization of several members of the hnRNP family, hnRNP A1, A2, B1, C1, C2 and K. 16 cell lines (SCLC and NSCLC) and biopsies from 32 lung cancer patients were analyzed. Our results suggest that, besides hnRNP A2/B1, the expression of other members of the hnRNP family is altered both in SCLC and NSCLC. In the biopsies, negative or low expression of the hnRNP proteins analyzed was observed in normal epithelial cells whereas lung cancer cells showed highly intense nuclear or cytoplasmic immunolocalization. In all the lung cancer cell lines, the mRNA for all the hnRNP proteins was detected. In general, higher levels of hnRNP mRNAs were found in SCLC as compared with NSCLC. Our results also suggest that the expression and processing of each hnRNP protein in lung cancer is independently regulated and is not exclusively related to proliferation status. In SCLC cell lines, hnRNP A1 protein expression correlated with that of Bcl-x L . In the lung cancer cell lines, hnRNP K protein localization varied with the cellular confluence.