lisa gangi - Academia.edu (original) (raw)
Papers by lisa gangi
Human Reproduction, 2007
BACKGROUND: Gene expression profiling of normal receptive endometrium has been characterized, but... more BACKGROUND: Gene expression profiling of normal receptive endometrium has been characterized, but intrinsic defects in endometrial gene expression associated with implantation failure have not been reported. METHODS: Women who had previously participated as recipients in oocyte donation cycles and repeatedly exhibited implantation failure (Group A, study group) or had at least one successful cycle (Group B, control group) and spontaneously fertile women (Group C, normal fertility group) were recruited. All were treated with exogenous estradiol and progesterone to induce an endometrial cycle, and an endometrial biopsy was taken on the seventh day of progesterone administration. RNA from each sample was analysed by cDNA microarrays to identify differentially expressed genes between groups. RESULTS: 63 transcripts were differentially expressed (!2-fold) between Groups A and B, of which 16 were subjected to real time RT -PCR. Eleven of these were significantly decreased in Group A with regard to Groups B and C. Among the dysregulated genes were MMP-7, CXCR4, PAEP and C4BPA. CONCLUSIONS: Repeated implantation failure in some oocyte recipients is associated with an intrinsic defect in the expression of multiple genes in their endometrium. Significantly decreased levels of several transcripts in endometria without manifest abnormalities is demonstrated for the first time and shown to be associated with implantation failure.
BACKGROUND: Gene expression profiling of normal receptive endometrium has been characterized, but... more BACKGROUND: Gene expression profiling of normal receptive endometrium has been characterized, but intrinsic defects in endometrial gene expression associated with implantation failure have not been reported. METHODS: Women who had previously participated as recipients in oocyte donation cycles and repeatedly exhibited implantation failure (Group A, study group) or had at least one successful cycle (Group B, control group) and spontaneously fertile women (Group C, normal fertility group) were recruited. All were treated with exogenous estradiol and progesterone to induce an endometrial cycle, and an endometrial biopsy was taken on the seventh day of progesterone administration. RNA from each sample was analysed by cDNA microarrays to identify differentially expressed genes between groups. RESULTS: 63 transcripts were differentially expressed (!2-fold) between Groups A and B, of which 16 were subjected to real time RT -PCR. Eleven of these were significantly decreased in Group A with regard to Groups B and C. Among the dysregulated genes were MMP-7, CXCR4, PAEP and C4BPA. CONCLUSIONS: Repeated implantation failure in some oocyte recipients is associated with an intrinsic defect in the expression of multiple genes in their endometrium. Significantly decreased levels of several transcripts in endometria without manifest abnormalities is demonstrated for the first time and shown to be associated with implantation failure.
BMC Genomics, 2005
Background Microarrays for the analysis of gene expression are of three different types: short ol... more Background Microarrays for the analysis of gene expression are of three different types: short oligonucleotide (25–30 base), long oligonucleotide (50–80 base), and cDNA (highly variable in length). The short oligonucleotide and cDNA arrays have been the mainstay of expression analysis to date, but long oligonucleotide platforms are gaining in popularity and will probably replace cDNA arrays. As part of a validation study for the long oligonucleotide arrays, we compared and contrasted expression profiles from the three formats, testing RNA from six different cell lines against a universal reference standard. Results The three platforms had 6430 genes in common. In general, correlation of gene expression levels across the platforms was good when defined by concordance in the direction of expression difference (upregulation or downregulation), scatter plot analysis, principal component analysis, cell line correlation or quantitative RT-PCR. The overall correlations (r values) between platforms were in the range 0.7 to 0.8, as determined by analysis of scatter plots. When concordance was measured for expression ratios significant at p-values of <0.05 and at expression threshold levels of 1.5 and 2-fold, the agreement among the platforms was very high, ranging from 93% to 100%. Conclusion Our results indicate that the long oligonucleotide platform is highly suitable for expression analysis and compares favorably with the cDNA and short oligonucleotide varieties. All three platforms can give similar and reproducible results if the criterion is the direction of change in gene expression and minimal emphasis is placed on the magnitude of change.
Human Reproduction, 2007
BACKGROUND: Gene expression profiling of normal receptive endometrium has been characterized, but... more BACKGROUND: Gene expression profiling of normal receptive endometrium has been characterized, but intrinsic defects in endometrial gene expression associated with implantation failure have not been reported. METHODS: Women who had previously participated as recipients in oocyte donation cycles and repeatedly exhibited implantation failure (Group A, study group) or had at least one successful cycle (Group B, control group) and spontaneously fertile women (Group C, normal fertility group) were recruited. All were treated with exogenous estradiol and progesterone to induce an endometrial cycle, and an endometrial biopsy was taken on the seventh day of progesterone administration. RNA from each sample was analysed by cDNA microarrays to identify differentially expressed genes between groups. RESULTS: 63 transcripts were differentially expressed (!2-fold) between Groups A and B, of which 16 were subjected to real time RT -PCR. Eleven of these were significantly decreased in Group A with regard to Groups B and C. Among the dysregulated genes were MMP-7, CXCR4, PAEP and C4BPA. CONCLUSIONS: Repeated implantation failure in some oocyte recipients is associated with an intrinsic defect in the expression of multiple genes in their endometrium. Significantly decreased levels of several transcripts in endometria without manifest abnormalities is demonstrated for the first time and shown to be associated with implantation failure.
Nature Genetics, 1999
Mammary gland involution follows the cessation of lactation. Similar to lactogenesis, the involut... more Mammary gland involution follows the cessation of lactation. Similar to lactogenesis, the involution process requires an ordered sequence of events including the termination of milk production, increased secretion of lactoferrin, apoptosis of the mammary epithelium and remodelling of the extracellular matrix of the gland. A completed cycle of involution requires apoptosis of the epithelial mammary structures involved in milk production, resulting in the loss of the milk-secreting alveoli as the gland returns to a pre-pregnant state. The complex cellular remodelling and cell-specific targeted apoptosis make mammary involution an excellent model system to apply to microarray platforms. The development of a 3K mammaryenriched mouse cDNA microarray has enabled us to perform high-throughput hybridization analysis to establish the gene expression profiles associated with the distinct cellular stages of the involution process. We find that a fall in anti-apoptosis genes appears to initiate the process, followed by induction of stressresponse genes. In addition to uncovering many genes involved in cell-cycle regulation, apoptosis, DNA repair and cytoskeletal remodelling, these microarrays have proven to be a powerful tool for gene discovery in epithelial involution.
Nucleic Acids Research, 2004
The genome is burdened with repetitive sequences that are generally embedded in silenced chromati... more The genome is burdened with repetitive sequences that are generally embedded in silenced chromatin. We have previously demonstrated that Lsh (lymphoid-specific helicase) is crucial for the control of heterochromatin at pericentromeric regions consisting of satellite repeats. In this study, we searched for additional genomic targets of Lsh by examining the effects of Lsh deletion on repeat regions and single copy gene sequences. We found that the absence of Lsh resulted in an increased association of acetylated histones with repeat sequences and transcriptional reactivation of their silenced state. In contrast, selected single copy genes displayed no change in histone acetylation levels, and their transcriptional rate was indistinguishable compared to Lsh-deficient cells and wildtype controls. Microarray analysis of total RNA derived from brain and liver tissues revealed that ,0.4% of the 15 247 examined loci were abnormally expressed in Lsh/-/embryos and almost two-thirds of these deregulated sequences contained repeats, mainly retroviral LTR (long terminal repeat) elements. Chromatin immunoprecipitation analysis demonstrated a direct interaction of Lsh with repetitive sites in the genome. These data suggest that the repetitive sites are direct targets of Lsh action and that Lsh plays an important role as 'epigenetic guardian' of the genome to protect against deregulation of parasitic retroviral elements.
Tumor Biology, 2005
G-band chromosomal karyotyping of fetal cells obtained by invasive prenatal testing has been used... more G-band chromosomal karyotyping of fetal cells obtained by invasive prenatal testing has been used since the 1960s to identify structural chromosomal anomalies. Prenatal testing is usually performed in response to parental request, increased risk of fetal chromosomal abnormality associated with advanced maternal age, a high-risk screening test and/or the presence of a congenital malformation identified by ultrasonography. The results of karyotyping may inform the long-term prognosis (e.g. aneuploidy being associated with a poor outcome or microscopic chromosomal anomalies predicting global neurodevelopmental morbidity). Relatively recent advances in microarray technology are now enabling high-resolution genomewide evaluation for DNA copy number abnormalities (e.g. deletions or duplications). While such technological advances promise increased sensitivity and specificity they can also pose difficult challenges of interpretation and clinical management. This review aims to give interested clinicians without an extensive prior knowledge of microarray technology, an overview of its use in prenatal diagnosis, the literature to date, advantages, potential pitfalls and experience from our own tertiary center.
Journal of Cellular Physiology, 2006
Ovarian cancer is an aggressive disease of poor prognostic when detected at advanced stage. It is... more Ovarian cancer is an aggressive disease of poor prognostic when detected at advanced stage. It is widely accepted that the ovarian surface epithelium plays a central role in disease etiology, but little is known about disease progression at the molecular level. To identify genes involved in ovarian tumorigenesis, we carried out a genome-wide transcriptomic analysis of six spontaneously transformed mouse ovarian surface epithelial (MOSE) cell lines, an in vitro model for human ovarian carcinoma. Loess normalization followed by statistical analysis with control of multiple testing resulted in 509 differentially expressed genes using an adjusted P-value ≤0.05 as cut-off. The top 20 differentially expressed genes included 10 genes (Spp1, Cyp1b1, Btg1, Cfh, Mt1, Mt2, Igfbp5, Gstm1, Gstm2, and Esr1) implicated in various aspects of ovarian carcinomas, and other 3 genes (Gsto1, Lcn7, and Alcam) associated to breast cancer. Upon functional analysis, the majority of alterations affected genes involved in glutathione metabolism and MAPK signaling pathways. Interestingly, over 20% of the aberrantly expressed genes were related to extracellular components, suggestive of potential markers of disease progression. In addition, we identified the genes Pura, Cnn3, Arpc1b, Map4k4, Tgfb1i4, and Crsp2 correlated to in vivo tumorigenic parameters previously reported for these cells. Taken together, our findings support the utility of MOSE cells in studying ovarian cancer biology and as a source of novel diagnostic and therapeutic targets. J.Cell.Physiol. © 2005 Wiley-Liss, Inc.
Applied Bioinformatics, 2006
developmental diseases. Array comparative genomic hybridisation (aCGH) is an emerging technology ... more developmental diseases. Array comparative genomic hybridisation (aCGH) is an emerging technology that allows detection of chromosomal gains and losses in a high-resolution format. When aCGH is performed on cDNA and oligonucleotide microarrays, the impact of DNA copy number on gene transcription profiles may be directly compared. We have created an online software tool, WebaCGH, that functions to (i) upload aCGH and gene transcription results from multiple experiments; (ii) identify significant aberrant regions using a local Z-score threshold in user-selected chromosomal segments subjected to smoothing with moving averages; and (iii) display results in a graphical format with full genome and individual chromosome views. In the individual chromosome display, data can be zoomed in/out in both dimensions (i.e. ratio and physical location) and plotted features can have 'mouse over' linking to outside databases to identify loci of interest. Uploaded data can be stored indefinitely for subsequent retrieval and analysis. WebaCGH was created as a Java-based web application using the open-source database MySQL . Availability: WebaCGH is freely accessible at
To identify the molecular basis underlying the functions of tumor-associated macrophages (TAMs), ... more To identify the molecular basis underlying the functions of tumor-associated macrophages (TAMs), we characterized the gene expression profile of TAMs isolated from a murine fibrosarcoma in comparison with peritoneal macrophages (PECs) and myeloid suppressor cells (MSCs), using a cDNA microarray technology. Among the differentially expressed genes, 15 genes relevant to inflammation and immunity were validated by real-time polymerase chain reaction (PCR) and protein production. Resting TAMs showed a characteristic gene expression pattern with higher expression of genes coding for the immunosuppressive cytokine IL-10, phagocytosis-related receptors/molecules (Msr2 and C1q), and inflammatory chemokines as expected, as well as, unexpectedly, IFNinducible chemokines (CXCL9, CXCL10, CXCL16). Immunohistology confirmed and extended the in vitro analysis by showing that TAMs express M2-associated molecules (eg, IL-10 and MGL1), as well as CCL2, CCL5, CXCL9, CXCL10, and CXCL16, but no appreciable NOS2. Lipopolysaccharide (LPS)-mediated activation of TAMs resulted in defective expression of several proinflammatory cytokines (eg, IL-1, IL-6, TNF-␣) and chemokines (eg, CCL3), as opposed to a strong up-regulation of immunosuppressive cytokines (IL-10, TGF) and IFN-inducible chemokines (CCL5, CXCL9, CXCL10, CXCL16). Thus, profiling of TAMs from a murine sarcoma revealed unexpected expression of IFN-inducible chemokines, associated with an M2 phenotype (IL-10 high , IL-12 low ), and divergent regulation of the NF-B versus the IRF-3/STAT1 pathway. (Blood. 2006;107:2112-2122)
To identify the molecular basis underlying the functions of Tumor-Associated Macrophages (TAM), w... more To identify the molecular basis underlying the functions of Tumor-Associated Macrophages (TAM), we characterized the gene expression profile of TAM isolated from a murine fibrosarcoma in comparison with peritoneal macrophages (PEC) and immature suppressor cells (MSC), using a cDNA microarray technology. Among the differentially expressed genes, 15 genes relevant to inflammation and immunity were validated by realtime PCR and protein production. Resting TAM showed a characteristic gene expression pattern, with higher expression of genes coding for the immunosuppressive cytokine IL-10, phagocytosis-related receptors/molecules (Msr2 and C1q) and inflammatory chemokines (CCL2 and CCL5) as expected, as well as, unexpectedly, IFN-inducible chemokines (CXCL9, CXCL10, CXCL16). Immunohistology confirmed and extended the in vitro analysis by showing that TAM express M2-associated molecules (e.g. IL-10 and MGL1), as well as CCL2, CCL5, CXCL9, CXCL10 and CXCL16, but no appreciable NOS2. Lipopolysaccharide (LPS)mediated activation of TAM resulted in defective expression of several proinflammatory cytokines (e.g. IL-1β, IL-6, TNF-α) and chemokines (eg. CCL3), as opposed to a strong upregulation of immunosuppressive cytokines (IL-10, TGFβ) and IFN-inducible chemokines (CCL5, CXCL9, CXCL10, CXCL16). Thus, profiling of TAM from a murine sarcoma revealed unexpected expression of IFN-inducible chemokines, associated with an M2
Journal of Clinical Investigation, 2002
Cells. SW1 and B16F10 mouse melanoma cells were maintained in DMEM supplemented with 10% FBS, L-g... more Cells. SW1 and B16F10 mouse melanoma cells were maintained in DMEM supplemented with 10% FBS, L-glutamine, and antibiotics.
The resistance of melanoma to apoptosis, as well as its growth and metastasis capabilities, can b... more The resistance of melanoma to apoptosis, as well as its growth and metastasis capabilities, can be overcome by expression of a peptide derived from amino acid (aa) 51 to 100 of ATF2. Here we show that expression of ATF2 (51-100) in human melanoma cells reduced their growth in nude mice, which was additionally inhibited upon treatment with protein kinase inhibitors UCN-01 or SB203580. Injection of a fusion protein consisting of HIV-TAT and aa 51 to 100 of ATF2 into SW1 melanomas efficiently inhibits their growth and their metastasis up to complete regression. Additionally, expression of a 10aa peptide that corresponds to aa 51 to 60 of ATF2 sensitizes melanoma cells to spontaneous apoptosis, which coincides with activation of caspase 9 and poly(ADPribose) polymerase cleavage, and inhibit their growth in vivo. The 10aa peptide increases the association of c-Jun NH 2 -terminal kinase with c-Jun but not with ATF2, resulting in concomitant increase in TRE-mediated transcription. Our study points to mechanisms underlying the activities of the ATF2 peptide while highlighting its possible use in drug design.
To identify changes in gene expression with transformation and metastasis, we investigated differ... more To identify changes in gene expression with transformation and metastasis, we investigated differential gene expression in a squamous carcinoma model established in syngeneic mice. We used mRNA differential display (DD) to detect global differences and cDNA arrays enriched for cancer-associated genes using mRNA from primary keratinocytes, transformed Pam 212 squamous carcinoma cells, and metastases of Pam 212. After DD, 72 candidate cDNAs expressed primarily in transformed and metastatic cells were selected and cloned. Fifty-seven were detected, and 32 were confirmed to be differentially expressed by Northern blot analysis. mRNA expression profiles were also generated using a mouse cDNA array composed of 4000 elements representing known genes and expressed sequence tags plus the 57 DD candidate cDNAs detected by Northern analysis to facilitate data validation. cDNA array detected 76.9% of the differentially expressed mRNAs selected from DD and confirmed by Northern blot, whereas low-abundance mRNAs did not reach the threshold for detection by the lower-sensitivity array method. . Strikingly, 10 of 22 genes in the cluster expressed in metastases have been associated with activation of the nuclear factor (NF)-B signal pathway. The NF-B-inducible cytokine Gro-1 was recently shown to promote tumor growth, metastasis, and angiogenesis of squamous cell carcinomas in vivo (Loukinova et al., Oncogene, 19: 3477-3486, 2000). The results demonstrate that early response genes related to NF-B contribute to metastatic tumor progression. Comparison of cell lines and tumor tissue revealed a concordance of ϳ50% by array, and 70% for Northern-confirmed, metastasis-related genes. Functional genomic approaches comparing expression among cell lines and tumor tissue may promote a better understanding of the genes expressed by malignant and host cells during tumor progression and metastasis. growth factor/scatter factor-induced activation of MEK and PI3K pathways contributes to expression of proangiogenic cytokines IL-8, and VEGF in head, and neck squamous cell carcinoma,
Human Reproduction, 2007
BACKGROUND: Gene expression profiling of normal receptive endometrium has been characterized, but... more BACKGROUND: Gene expression profiling of normal receptive endometrium has been characterized, but intrinsic defects in endometrial gene expression associated with implantation failure have not been reported. METHODS: Women who had previously participated as recipients in oocyte donation cycles and repeatedly exhibited implantation failure (Group A, study group) or had at least one successful cycle (Group B, control group) and spontaneously fertile women (Group C, normal fertility group) were recruited. All were treated with exogenous estradiol and progesterone to induce an endometrial cycle, and an endometrial biopsy was taken on the seventh day of progesterone administration. RNA from each sample was analysed by cDNA microarrays to identify differentially expressed genes between groups. RESULTS: 63 transcripts were differentially expressed (!2-fold) between Groups A and B, of which 16 were subjected to real time RT -PCR. Eleven of these were significantly decreased in Group A with regard to Groups B and C. Among the dysregulated genes were MMP-7, CXCR4, PAEP and C4BPA. CONCLUSIONS: Repeated implantation failure in some oocyte recipients is associated with an intrinsic defect in the expression of multiple genes in their endometrium. Significantly decreased levels of several transcripts in endometria without manifest abnormalities is demonstrated for the first time and shown to be associated with implantation failure.
BACKGROUND: Gene expression profiling of normal receptive endometrium has been characterized, but... more BACKGROUND: Gene expression profiling of normal receptive endometrium has been characterized, but intrinsic defects in endometrial gene expression associated with implantation failure have not been reported. METHODS: Women who had previously participated as recipients in oocyte donation cycles and repeatedly exhibited implantation failure (Group A, study group) or had at least one successful cycle (Group B, control group) and spontaneously fertile women (Group C, normal fertility group) were recruited. All were treated with exogenous estradiol and progesterone to induce an endometrial cycle, and an endometrial biopsy was taken on the seventh day of progesterone administration. RNA from each sample was analysed by cDNA microarrays to identify differentially expressed genes between groups. RESULTS: 63 transcripts were differentially expressed (!2-fold) between Groups A and B, of which 16 were subjected to real time RT -PCR. Eleven of these were significantly decreased in Group A with regard to Groups B and C. Among the dysregulated genes were MMP-7, CXCR4, PAEP and C4BPA. CONCLUSIONS: Repeated implantation failure in some oocyte recipients is associated with an intrinsic defect in the expression of multiple genes in their endometrium. Significantly decreased levels of several transcripts in endometria without manifest abnormalities is demonstrated for the first time and shown to be associated with implantation failure.
BMC Genomics, 2005
Background Microarrays for the analysis of gene expression are of three different types: short ol... more Background Microarrays for the analysis of gene expression are of three different types: short oligonucleotide (25–30 base), long oligonucleotide (50–80 base), and cDNA (highly variable in length). The short oligonucleotide and cDNA arrays have been the mainstay of expression analysis to date, but long oligonucleotide platforms are gaining in popularity and will probably replace cDNA arrays. As part of a validation study for the long oligonucleotide arrays, we compared and contrasted expression profiles from the three formats, testing RNA from six different cell lines against a universal reference standard. Results The three platforms had 6430 genes in common. In general, correlation of gene expression levels across the platforms was good when defined by concordance in the direction of expression difference (upregulation or downregulation), scatter plot analysis, principal component analysis, cell line correlation or quantitative RT-PCR. The overall correlations (r values) between platforms were in the range 0.7 to 0.8, as determined by analysis of scatter plots. When concordance was measured for expression ratios significant at p-values of <0.05 and at expression threshold levels of 1.5 and 2-fold, the agreement among the platforms was very high, ranging from 93% to 100%. Conclusion Our results indicate that the long oligonucleotide platform is highly suitable for expression analysis and compares favorably with the cDNA and short oligonucleotide varieties. All three platforms can give similar and reproducible results if the criterion is the direction of change in gene expression and minimal emphasis is placed on the magnitude of change.
Human Reproduction, 2007
BACKGROUND: Gene expression profiling of normal receptive endometrium has been characterized, but... more BACKGROUND: Gene expression profiling of normal receptive endometrium has been characterized, but intrinsic defects in endometrial gene expression associated with implantation failure have not been reported. METHODS: Women who had previously participated as recipients in oocyte donation cycles and repeatedly exhibited implantation failure (Group A, study group) or had at least one successful cycle (Group B, control group) and spontaneously fertile women (Group C, normal fertility group) were recruited. All were treated with exogenous estradiol and progesterone to induce an endometrial cycle, and an endometrial biopsy was taken on the seventh day of progesterone administration. RNA from each sample was analysed by cDNA microarrays to identify differentially expressed genes between groups. RESULTS: 63 transcripts were differentially expressed (!2-fold) between Groups A and B, of which 16 were subjected to real time RT -PCR. Eleven of these were significantly decreased in Group A with regard to Groups B and C. Among the dysregulated genes were MMP-7, CXCR4, PAEP and C4BPA. CONCLUSIONS: Repeated implantation failure in some oocyte recipients is associated with an intrinsic defect in the expression of multiple genes in their endometrium. Significantly decreased levels of several transcripts in endometria without manifest abnormalities is demonstrated for the first time and shown to be associated with implantation failure.
Nature Genetics, 1999
Mammary gland involution follows the cessation of lactation. Similar to lactogenesis, the involut... more Mammary gland involution follows the cessation of lactation. Similar to lactogenesis, the involution process requires an ordered sequence of events including the termination of milk production, increased secretion of lactoferrin, apoptosis of the mammary epithelium and remodelling of the extracellular matrix of the gland. A completed cycle of involution requires apoptosis of the epithelial mammary structures involved in milk production, resulting in the loss of the milk-secreting alveoli as the gland returns to a pre-pregnant state. The complex cellular remodelling and cell-specific targeted apoptosis make mammary involution an excellent model system to apply to microarray platforms. The development of a 3K mammaryenriched mouse cDNA microarray has enabled us to perform high-throughput hybridization analysis to establish the gene expression profiles associated with the distinct cellular stages of the involution process. We find that a fall in anti-apoptosis genes appears to initiate the process, followed by induction of stressresponse genes. In addition to uncovering many genes involved in cell-cycle regulation, apoptosis, DNA repair and cytoskeletal remodelling, these microarrays have proven to be a powerful tool for gene discovery in epithelial involution.
Nucleic Acids Research, 2004
The genome is burdened with repetitive sequences that are generally embedded in silenced chromati... more The genome is burdened with repetitive sequences that are generally embedded in silenced chromatin. We have previously demonstrated that Lsh (lymphoid-specific helicase) is crucial for the control of heterochromatin at pericentromeric regions consisting of satellite repeats. In this study, we searched for additional genomic targets of Lsh by examining the effects of Lsh deletion on repeat regions and single copy gene sequences. We found that the absence of Lsh resulted in an increased association of acetylated histones with repeat sequences and transcriptional reactivation of their silenced state. In contrast, selected single copy genes displayed no change in histone acetylation levels, and their transcriptional rate was indistinguishable compared to Lsh-deficient cells and wildtype controls. Microarray analysis of total RNA derived from brain and liver tissues revealed that ,0.4% of the 15 247 examined loci were abnormally expressed in Lsh/-/embryos and almost two-thirds of these deregulated sequences contained repeats, mainly retroviral LTR (long terminal repeat) elements. Chromatin immunoprecipitation analysis demonstrated a direct interaction of Lsh with repetitive sites in the genome. These data suggest that the repetitive sites are direct targets of Lsh action and that Lsh plays an important role as 'epigenetic guardian' of the genome to protect against deregulation of parasitic retroviral elements.
Tumor Biology, 2005
G-band chromosomal karyotyping of fetal cells obtained by invasive prenatal testing has been used... more G-band chromosomal karyotyping of fetal cells obtained by invasive prenatal testing has been used since the 1960s to identify structural chromosomal anomalies. Prenatal testing is usually performed in response to parental request, increased risk of fetal chromosomal abnormality associated with advanced maternal age, a high-risk screening test and/or the presence of a congenital malformation identified by ultrasonography. The results of karyotyping may inform the long-term prognosis (e.g. aneuploidy being associated with a poor outcome or microscopic chromosomal anomalies predicting global neurodevelopmental morbidity). Relatively recent advances in microarray technology are now enabling high-resolution genomewide evaluation for DNA copy number abnormalities (e.g. deletions or duplications). While such technological advances promise increased sensitivity and specificity they can also pose difficult challenges of interpretation and clinical management. This review aims to give interested clinicians without an extensive prior knowledge of microarray technology, an overview of its use in prenatal diagnosis, the literature to date, advantages, potential pitfalls and experience from our own tertiary center.
Journal of Cellular Physiology, 2006
Ovarian cancer is an aggressive disease of poor prognostic when detected at advanced stage. It is... more Ovarian cancer is an aggressive disease of poor prognostic when detected at advanced stage. It is widely accepted that the ovarian surface epithelium plays a central role in disease etiology, but little is known about disease progression at the molecular level. To identify genes involved in ovarian tumorigenesis, we carried out a genome-wide transcriptomic analysis of six spontaneously transformed mouse ovarian surface epithelial (MOSE) cell lines, an in vitro model for human ovarian carcinoma. Loess normalization followed by statistical analysis with control of multiple testing resulted in 509 differentially expressed genes using an adjusted P-value ≤0.05 as cut-off. The top 20 differentially expressed genes included 10 genes (Spp1, Cyp1b1, Btg1, Cfh, Mt1, Mt2, Igfbp5, Gstm1, Gstm2, and Esr1) implicated in various aspects of ovarian carcinomas, and other 3 genes (Gsto1, Lcn7, and Alcam) associated to breast cancer. Upon functional analysis, the majority of alterations affected genes involved in glutathione metabolism and MAPK signaling pathways. Interestingly, over 20% of the aberrantly expressed genes were related to extracellular components, suggestive of potential markers of disease progression. In addition, we identified the genes Pura, Cnn3, Arpc1b, Map4k4, Tgfb1i4, and Crsp2 correlated to in vivo tumorigenic parameters previously reported for these cells. Taken together, our findings support the utility of MOSE cells in studying ovarian cancer biology and as a source of novel diagnostic and therapeutic targets. J.Cell.Physiol. © 2005 Wiley-Liss, Inc.
Applied Bioinformatics, 2006
developmental diseases. Array comparative genomic hybridisation (aCGH) is an emerging technology ... more developmental diseases. Array comparative genomic hybridisation (aCGH) is an emerging technology that allows detection of chromosomal gains and losses in a high-resolution format. When aCGH is performed on cDNA and oligonucleotide microarrays, the impact of DNA copy number on gene transcription profiles may be directly compared. We have created an online software tool, WebaCGH, that functions to (i) upload aCGH and gene transcription results from multiple experiments; (ii) identify significant aberrant regions using a local Z-score threshold in user-selected chromosomal segments subjected to smoothing with moving averages; and (iii) display results in a graphical format with full genome and individual chromosome views. In the individual chromosome display, data can be zoomed in/out in both dimensions (i.e. ratio and physical location) and plotted features can have 'mouse over' linking to outside databases to identify loci of interest. Uploaded data can be stored indefinitely for subsequent retrieval and analysis. WebaCGH was created as a Java-based web application using the open-source database MySQL . Availability: WebaCGH is freely accessible at
To identify the molecular basis underlying the functions of tumor-associated macrophages (TAMs), ... more To identify the molecular basis underlying the functions of tumor-associated macrophages (TAMs), we characterized the gene expression profile of TAMs isolated from a murine fibrosarcoma in comparison with peritoneal macrophages (PECs) and myeloid suppressor cells (MSCs), using a cDNA microarray technology. Among the differentially expressed genes, 15 genes relevant to inflammation and immunity were validated by real-time polymerase chain reaction (PCR) and protein production. Resting TAMs showed a characteristic gene expression pattern with higher expression of genes coding for the immunosuppressive cytokine IL-10, phagocytosis-related receptors/molecules (Msr2 and C1q), and inflammatory chemokines as expected, as well as, unexpectedly, IFNinducible chemokines (CXCL9, CXCL10, CXCL16). Immunohistology confirmed and extended the in vitro analysis by showing that TAMs express M2-associated molecules (eg, IL-10 and MGL1), as well as CCL2, CCL5, CXCL9, CXCL10, and CXCL16, but no appreciable NOS2. Lipopolysaccharide (LPS)-mediated activation of TAMs resulted in defective expression of several proinflammatory cytokines (eg, IL-1, IL-6, TNF-␣) and chemokines (eg, CCL3), as opposed to a strong up-regulation of immunosuppressive cytokines (IL-10, TGF) and IFN-inducible chemokines (CCL5, CXCL9, CXCL10, CXCL16). Thus, profiling of TAMs from a murine sarcoma revealed unexpected expression of IFN-inducible chemokines, associated with an M2 phenotype (IL-10 high , IL-12 low ), and divergent regulation of the NF-B versus the IRF-3/STAT1 pathway. (Blood. 2006;107:2112-2122)
To identify the molecular basis underlying the functions of Tumor-Associated Macrophages (TAM), w... more To identify the molecular basis underlying the functions of Tumor-Associated Macrophages (TAM), we characterized the gene expression profile of TAM isolated from a murine fibrosarcoma in comparison with peritoneal macrophages (PEC) and immature suppressor cells (MSC), using a cDNA microarray technology. Among the differentially expressed genes, 15 genes relevant to inflammation and immunity were validated by realtime PCR and protein production. Resting TAM showed a characteristic gene expression pattern, with higher expression of genes coding for the immunosuppressive cytokine IL-10, phagocytosis-related receptors/molecules (Msr2 and C1q) and inflammatory chemokines (CCL2 and CCL5) as expected, as well as, unexpectedly, IFN-inducible chemokines (CXCL9, CXCL10, CXCL16). Immunohistology confirmed and extended the in vitro analysis by showing that TAM express M2-associated molecules (e.g. IL-10 and MGL1), as well as CCL2, CCL5, CXCL9, CXCL10 and CXCL16, but no appreciable NOS2. Lipopolysaccharide (LPS)mediated activation of TAM resulted in defective expression of several proinflammatory cytokines (e.g. IL-1β, IL-6, TNF-α) and chemokines (eg. CCL3), as opposed to a strong upregulation of immunosuppressive cytokines (IL-10, TGFβ) and IFN-inducible chemokines (CCL5, CXCL9, CXCL10, CXCL16). Thus, profiling of TAM from a murine sarcoma revealed unexpected expression of IFN-inducible chemokines, associated with an M2
Journal of Clinical Investigation, 2002
Cells. SW1 and B16F10 mouse melanoma cells were maintained in DMEM supplemented with 10% FBS, L-g... more Cells. SW1 and B16F10 mouse melanoma cells were maintained in DMEM supplemented with 10% FBS, L-glutamine, and antibiotics.
The resistance of melanoma to apoptosis, as well as its growth and metastasis capabilities, can b... more The resistance of melanoma to apoptosis, as well as its growth and metastasis capabilities, can be overcome by expression of a peptide derived from amino acid (aa) 51 to 100 of ATF2. Here we show that expression of ATF2 (51-100) in human melanoma cells reduced their growth in nude mice, which was additionally inhibited upon treatment with protein kinase inhibitors UCN-01 or SB203580. Injection of a fusion protein consisting of HIV-TAT and aa 51 to 100 of ATF2 into SW1 melanomas efficiently inhibits their growth and their metastasis up to complete regression. Additionally, expression of a 10aa peptide that corresponds to aa 51 to 60 of ATF2 sensitizes melanoma cells to spontaneous apoptosis, which coincides with activation of caspase 9 and poly(ADPribose) polymerase cleavage, and inhibit their growth in vivo. The 10aa peptide increases the association of c-Jun NH 2 -terminal kinase with c-Jun but not with ATF2, resulting in concomitant increase in TRE-mediated transcription. Our study points to mechanisms underlying the activities of the ATF2 peptide while highlighting its possible use in drug design.
To identify changes in gene expression with transformation and metastasis, we investigated differ... more To identify changes in gene expression with transformation and metastasis, we investigated differential gene expression in a squamous carcinoma model established in syngeneic mice. We used mRNA differential display (DD) to detect global differences and cDNA arrays enriched for cancer-associated genes using mRNA from primary keratinocytes, transformed Pam 212 squamous carcinoma cells, and metastases of Pam 212. After DD, 72 candidate cDNAs expressed primarily in transformed and metastatic cells were selected and cloned. Fifty-seven were detected, and 32 were confirmed to be differentially expressed by Northern blot analysis. mRNA expression profiles were also generated using a mouse cDNA array composed of 4000 elements representing known genes and expressed sequence tags plus the 57 DD candidate cDNAs detected by Northern analysis to facilitate data validation. cDNA array detected 76.9% of the differentially expressed mRNAs selected from DD and confirmed by Northern blot, whereas low-abundance mRNAs did not reach the threshold for detection by the lower-sensitivity array method. . Strikingly, 10 of 22 genes in the cluster expressed in metastases have been associated with activation of the nuclear factor (NF)-B signal pathway. The NF-B-inducible cytokine Gro-1 was recently shown to promote tumor growth, metastasis, and angiogenesis of squamous cell carcinomas in vivo (Loukinova et al., Oncogene, 19: 3477-3486, 2000). The results demonstrate that early response genes related to NF-B contribute to metastatic tumor progression. Comparison of cell lines and tumor tissue revealed a concordance of ϳ50% by array, and 70% for Northern-confirmed, metastasis-related genes. Functional genomic approaches comparing expression among cell lines and tumor tissue may promote a better understanding of the genes expressed by malignant and host cells during tumor progression and metastasis. growth factor/scatter factor-induced activation of MEK and PI3K pathways contributes to expression of proangiogenic cytokines IL-8, and VEGF in head, and neck squamous cell carcinoma,