Biomedical subjects

Dominique Lorang

Publications and source records attributed to Dominique Lorang.

11 recordsLinked to original sources

PubMed2005

Gland size is associated with changes in gene expression profiles in sporadic parathyroid adenomas.

BACKGROUND: Sporadic parathyroid adenomas (SPAs) are benign neoplasms responsible for most cases of primary hyperparathyroidism (pHPT). The molecular pathways responsible for the variations in clinical severity of pHPT are unknown. We studied gene expression profiles in patients with SPAs and pHPT to determine associations between these changes and clinical parameters. METHODS: We selected 10 patients with solitary SPAs and nonfamilial, non-MEN1 pHPT treated with surgery from 2001 to 2003. Pathologic and clinical data were reviewed. At operation, tissues from SPAs were frozen in liquid nitrogen; total RNA was obtained from sections, and the diagnosis was confirmed with hematoxylin and eosin staining. Control normal parathyroid RNA was age- and sex-matched. RNA was amplified, labeled, and hybridized to a microarray of 22,272 human oligonucleotides. Cluster analysis of gene expression, analysis of expression ratios, and comparison of clinical parameters were performed. RESULTS: All patients were cured; all specimens were consistent with SPAs. K means clustering divided the 10 patients into 2 distinct 5-patient gene expression groups by using uncentered correlation based on gene subgrouping. Of the clinical parameters, only the mean gland volume was significantly different between group 1 (390 +/- 160 mm(3)) and group 2 (1080 +/- 615 mm(3); P = .032 by Mann-Whitney test). Seventy-five genes were significantly upregulated or downregulated (with a ratio of <.33 or >3) compared with controls. These genes included the v-fos viral oncogene homolog and six calcium ion-binding signaling proteins. CONCLUSIONS: Differential expression of a few critical genes may contribute to differences in gland volume in SPAs. A better understanding of these pathways may help to define the pathophysiology of pHPT.

Adenoma↗

PubMed2004

Modulation of tumor-host interactions, angiogenesis, and tumor growth by tissue inhibitor of metalloproteinase 2 via a novel mechanism.

Solid tumors depend on angiogenesis for sustained growth. Tissue inhibitor of metalloproteinase 2 (TIMP-2) is an angiogenesis inhibitor initially characterized for its ability to block matrix metalloproteinases; however, recent data suggest that the antiangiogenic action of TIMP-2 may rely on matrix metalloproteinase-independent mechanisms. The aim of this study was to identify molecular pathways involved in the effects of TIMP-2 on processes dependent on tumor-host interactions such as angiogenesis. Using in vitro cell culture and a syngeneic murine tumor model, we compared the effects of TIMP-2 overexpression on gene expression profiles in vitro to those observed in vivo. Validating these findings by real-time quantitative PCR and layered protein scanning, we identified up-regulation of mitogen-activated protein kinase phosphatase 1 as an effector of the antiangiogenic function of TIMP-2. Up-regulation of mitogen-activated protein kinase phosphatase 1 in tumors overexpressing TIMP-2 leads to dephosphorylation of p38 mitogen-activated protein kinase and inhibition of tumor growth and angiogenesis. Phosphatase activity appears important in regulating tumor angiogenesis, offering a promising direction for the identification of novel molecular targets and antiangiogenic compounds for the treatment of cancer.

Animals↗

PubMed2004

Gene array analysis of macronodular adrenal hyperplasia confirms clinical heterogeneity and identifies several candidate genes as molecular mediators.

Corticotropin (ACTH)-independent macronodular adrenal hyperplasia (AIMAH) is a heterogeneous condition in which cortisol secretion may be mediated by gastrointestinal peptide (GIP), vasopressin, catecholamines and other hormones. We studied the expression profile of AIMAH by genomic cDNA microarray analysis. Total RNA was extracted from eight tissues (three GIP-dependent) and compared to total RNA obtained from adrenal glands from 62 normal subjects. Genes had to be altered in 75% of the patients, and be up- or downregulated at a cutoff ratio of at least 2.0; 82 and 31 genes were found to be consistently up- and downregulated, respectively. Among the former were regulators of transcription, chromatin remodeling, and cell cycle and adhesion. Downregulated sequences included genes involved in immune responses and insulin signaling. Hierarchical clustering correlated with the two main AIMAH diagnostic groups: GIP-dependent and non-GIP-dependent. The genes encoding the 7B2 protein (SGNE1) and WNT1-inducible signaling pathway protein 2 (WISP2) were specifically overexpressed in the GIP-dependent AIMAH. For these, and six more genes, the data were validated by semiquantitative amplification in samples from a total of 32 patients (the original eight, six more cases of AIMAH, and 18 other adrenocortical hyperplasias and tumors) and the H295R adrenocortical cancer cell line. In conclusion, our data confirmed AIMAH's clinical heterogeneity by identifying molecularly distinct diagnostic subgroups. Several candidate genes that may be responsible for AIMAH formation and/or progression were also identified, suggesting pathways that affect the cell cycle, adhesion and transcription as possible mediators of adrenocortical hyperplasia.

Adrenal Cortex Neoplasms↗

PubMed2004

Insulinomas and expression of an insulin splice variant.

BACKGROUND: Insulinomas are beta-cell tumours characterised by uncontrolled insulin secretion even in the presence of hypoglycaemia. However, the mechanisms allowing such excessive insulin secretion are not known. Insulin secretion can occur only when the beta-cell insulin stores have been replenished by insulin biosynthesis, which is mainly controlled by translation. Such specific translational regulation often involves the 5' untranslated region. We have identified an insulin splice variant in isolated human pancreatic islets of non-diabetic donors that retains 26 bp of intron 1 and thereby changes the 5' untranslated region, but leaves the coding region unchanged. This splice variant has increased translation efficiency in vitro and in vivo compared with native insulin mRNA. However, splice variant expression is less than 1% of native insulin mRNA in normal islets. METHODS: To test whether this splice variant is involved in insulin production by human insulinomas, we extracted RNA from nine laser-captured surgical insulinoma samples and from isolated islets of nine donors who did not have diabetes. We then determined the ratio of splice variant to native insulin mRNA by quantitative real-time RT-PCR. FINDINGS: The mean ratio of the splice variant to native insulin mRNA was increased more than 50-fold in insulinomas compared with normal islets, and this difference was present in all nine human insulinomas. Overexpression of the splice variant therefore seems to be a general characteristic of insulinomas and is estimated to contribute about 90% to insulin synthesis by these tumours. INTERPRETATION: Overexpression of the insulin splice variant with increased translation efficiency in insulinomas might explain how these tumours maintain high levels of insulin synthesis and secretion leading to hyperinsulinaemia-the hallmark of this disease.

Alternative Splicing↗

PubMed2004

A novel technique for quantifying changes in vascular density, endothelial cell proliferation and protein expression in response to modulators of angiogenesis using the chick chorioallantoic membrane (CAM) assay.

Reliable quantitative evaluation of molecular pathways is critical for both drug discovery and treatment monitoring. We have modified the CAM assay to quantitatively measure vascular density, endothelial proliferation, and changes in protein expression in response to anti-angiogenic and pro-angiogenic agents. This improved CAM assay can correlate changes in vascular density with changes seen on a molecular level. We expect that these described modifications will result in a single in vivo assay system, which will improve the ability to investigate molecular mechanisms underlying the angiogenic response.

Journal Article↗

PubMed2004

Early genetic mechanisms underlying the inhibitory effects of endostatin and fumagillin on human endothelial cells.

A tumor needs to initiate angiogenesis in order to develop its own blood supply, to grow, to invade, and to spread. Angiogenesis, under normal conditions, is a tightly regulated balance between endogenous pro- and antiangiogenic factors. In this study, we investigated, by microarray analysis, the effects of two known antiangiogenic agents (endostatin and fumagillin) on the gene expression profiles of human umbilical vein endothelial cells (HUVEC) in order to elucidate pathways common to the effects of these agents. We observed a majority of gene expression changes within 1 and 2 h of treatment. The genes demonstrating these early expression changes are involved in cell proliferation, gene transcription, and a number have unknown functions. We selected four genes (DOC1, KLF4, TC-1, ID1) from the microarray profile that showed a similar pattern of expression for both of the antiangiogenic agents we tested. We then used small interfering RNAs (siRNA) in an attempt to better understand the role of these selected genes in the inhibitory activity of these agents. Because the gene expression changes occurred within 1 and 2 h of treatment, these genes might be involved in the initial pathways of angiogenesis inhibition.

Angiogenesis Inhibitors↗

PubMed2003

Parathyroid gland-specific deletion of the mouse Men1 gene results in parathyroid neoplasia and hypercalcemic hyperparathyroidism.

The inactivation of the MEN1 tumor suppressor gene in patients leads to a constellation of changes in endocrine tissues, including parathyroid neoplasia, pituitary adenomas, pancreatic neuroendocrine tumors, and carcinoids. To study the pathophysiological consequences of the deletion of the MEN1 gene, we set out to create a mouse model of hyperparathyroidism resulting from the deletion of the Men1 gene in parathyroid tissue. We introduced a Men1 gene flanked by loxP sites into the mouse germ line and then used a parathyroid cell-specific promoter to drive the expression of Cre recombinase, resulting in the deletion of the Men1 gene. Here, we show that loss of Men1 gene function in the parathyroid glands of mice results in histological changes consistent with parathyroid neoplasia as well as systemic hypercalcemia. This model provides a means for dissecting the molecular basis of this familial cancer syndrome and may allow for the development of new strategies to treat related forms of hypercalcemia.

Animals↗

PubMed2003

Regulation of the pro-angiogenic microenvironment by carboxyamido-triazole.

Anti-angiogenic agents regulate tumor growth by inhibiting endothelial cell proliferation and invasion. Carboxyamido-triazole (CAI), an inhibitor of non-voltage-operated calcium entry and calcium influx-mediated pathways, has angiogenesis and invasion inhibitory activity. We hypothesized that CAI may express its anti-angiogenic effects through negative regulation of pro-angiogenic cytokine production and/or function. In vivo, orally administered CAI prevented A2058 human melanoma xenograft growth and concomitantly resulted in a marked reduction in circulating vascular endothelial growth factor (VEGF) and interleukin-8 (IL-8). In vitro, A2058 cell secretion of VEGF was inhibited by CAI treatment under limiting micronutrient conditions that approximate the tumor microenvironment, media restriction, and acidification to pH 6.8 (P=0.0003 and P=0.0006, respectively). VEGF and HIF-1alpha message and protein were also reduced by CAI treatment. Oral CAI treatment reduced vascular ingrowth in vivo into VEGF-containing Matrigel plugs. Commensurate with those findings, human umbilical vein endothelial cell (HUVEC) migration towards VEGF was reduced below background by exposure to CAI in the migration chamber (P<0.0001). An 88% reduction in circulating IL-8 concentration was measured in CAI-treated animals. However, IL-8 protein secretion and gene expression were increased by CAI treatment in culture (P< or =0.01), where CAI caused a dose-dependent acidification of the culture milieu (P< or =0.005). This paradox suggests that IL-8 production in vitro may be more sensitive to ambient pH than cytosolic calcium. These observations suggest that CAI inhibition of tumor cell VEGF production and endothelial cell response to VEGF results in disruption of signaling between the tumor and its microenvironment, causing a net anti-angiogenic effect.

Animals↗

PubMed2003

Peak stimulated insulin secretion is associated with specific changes in gene expression profiles in sporadic insulinomas.

BACKGROUND: The molecular pathways that are responsible for pathologic insulin secretion by insulinomas have not been characterized. We studied gene expression profiles from insulinomas and determined associations between these changes and preoperative peak serum insulin levels. METHODS: Ten patients with insulinomas underwent calcium-stimulated arteriography and surgical resection. Tumor RNA was isolated; corresponding complementary DNA was hybridized to 10K human complementary DNA arrays. Pooled human islet cell complementary DNA served as the control. Cluster analysis of gene expression and analysis of expression ratios was performed. RESULTS: Nineteen genes were up-regulated at least 3-fold in insulinomas compared with controls, which included the genes for islet amyloid polypeptide and proprotein convertase type 2. Cluster analysis revealed 2 groups of patients with insulinoma and with distinct patterns of gene expression. Mean peak serum insulin values between groups were 196 and 1100 (U/mL (P<.05), which demonstrates a significant difference in insulin response to calcium stimulation between these 2 groups. CONCLUSION: We show that genes that are relevant to the pathogenesis of hyperinsulinemia are expressed preferentially in insulinomas. In addition, patients with a distinct and common pattern of gene expression had significantly higher stimulated insulin secretion levels. The study of these genes may help to identify the biochemical pathways that are responsible for pathologic insulin secretion.

Adult↗

PubMed2002

Retroviral gene transfer of interferon-inducible protein 10 inhibits growth of human melanoma xenografts.

Interferon-inducible protein 10 (IP-10) is an immunomodulatory chemokine recently recognized to have potent antiangiogenic activity in vivo. Due to difficulties in the stability, manufacture and chronic administration of recombinant forms of endogenous antiangiogenic proteins, antiangiogenic gene therapy has emerged as a promising new form of cancer treatment. We retrovirally transduced A375 human melanoma cells with the human IP-10 gene and injected cells subcutaneously into nude mice. IP-10-transduced cells also were mixed with null-transduced cells in varying proportions before injection. In vivo growth of IP-10-transduced melanoma cells was markedly diminished compared to parental or null-transduced cells (p = 0.0002, Kruskal-Wallis test). This growth inhibition was associated with a marked reduction in microvessel density. The degree of growth inhibition of tumors following injection of a mixed population of null- and IP-10-transduced cells was directly associated with the fraction of IP-10-transduced cells present. We conclude that retroviral transduction of human melanoma cells with the IP-10 gene leads to sufficient protein secretion to inhibit angiogenesis and tumor growth. These findings suggest that IP-10 gene therapy might be an effective therapy in patients with cancer.

Animals↗

PubMed2002

Microarray gene expression analysis of murine tumor heterogeneity defined by dynamic contrast-enhanced MRI.

Current methods of studying angiogenesis are limited in their ability to serially evaluate in vivo function throughout a target tissue. Dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) and pharmacokinetic modeling provide a useful method for evaluating tissue vasculature based on contrast accumulation and washout. While it is often assumed that areas of high contrast enhancement and washout comprise areas of increased angiogenesis and tumor activity, the actual molecular pathways that are active in such areas are poorly understood. Using DCE-MRI in a murine subcutaneous tumor model, we were able to perform pharmacokinetic functional analysis of a tumor, coregistration of MRI images with histological cross-sections, immunohistochemistry, laser capture microdissection, and genetic profiling of tumor heterogeneity based on pharmacokinetic parameters. Using imaging as a template for biologic investigation, we have not found evidence of increased expression of proangiogenic modulators at the transcriptional level in either distinct pharmacokinetic region. Furthermore, these regions show no difference on histology and CD31 immunohistochemistry. However, the expression of ribosomal proteins was greatly increased in high enhancement and washout regions, implying increased protein translation and consequent increased cellular activity. Together, these findings point to the potential importance of posttranscriptional regulation in angiogenesis and the need for the development of angiogenesis-specific contrast agents to evaluate in vivo angiogenesis at a molecular level.

Animals↗