ABSTRACT: Human uterine fibroids are benign tumors derived from the smooth muscle layers of the uterus. Fibroid disease impacts up to 50% of premenopausal women in their daily life, and accounts for up to 50% of hysterectomies in the United States. Despite the health burden they impose, the development of these tumors is barely understood. To improve our understanding of this disease, we developed and characterized a patient-derived xenograft models by transplanting small pieces of human uterine fibroid tissue subcutaneously into immunodeficient mice. Growth of the fibroid grafts was dependent on 17beta-estradiol and progesterone supplementation and maintained for up to 60 days. A gene expression profiling study was performed in order to determine molecular characteristis of well-grown compared to not-grown uterine fibroid xenografts.
Project description:Normal myometrium and uterine fibroids (partially paired from the same donor) were profiled. FISH analysis was used to analyze the karyotype of the uterine fibroid samples. This study provides further insights in the development of uterine fibroids. Additional uterine fibroid samples from the same sample collection and cohort can be found at ArrayExpress under E-MTAB-340.
Project description:Plexiform leiomyomata are a histologically defined subgroup of benign uterine smooth muscle tumors based on their epitheliod cytology and abundant extracellular matrix. We used microarrays to compare plexiform leiomyomata to normal myometrium (smooth muscle of the uterine wall), typical leiomyomata, cellular or atypical leiomyomata and malignant leiomyosarcoma of the uterus. Experiment Overall Design: Samples analyzed on U133 Plus 2.0 and HuFL (GPL570 and GPL80). Experiment Overall Design: Data from the microarrays was merged by selection data for probe sets in which Entrez gene ID code on the GPL570 table equaled the Entrez gene ID on the GPL80 table. Experiment Overall Design: If more than one row on the GPL570 matched the value of the Entrez gene ID for a probe set on the GPL80 table, the values from the GPL570 were averaged. Thus, the GPL570 data was condensed to conform to the GPL80 format. Experiment Overall Design: The reformated raw data from both microarrays was then normalized such that the sum of the expression values was 3 million, and that values less than 20 were subsequently set to 20 (to permit log transformation in some statistical analysis).
Project description:Xenotransplantation holds the promise of providing an unlimited supply of donor organs for terminal patients with organ failure. The gal carbohydrate results in rejection of wild type pig grafts, however, chimerism established by expression of the GalT gene prior to transplantation in GalT knockout mice results in tolerance to Gal+ heart grafts. We used microarrays in order to further understand the early events that occur within grafts that demonstrate tolerance. Experiment Overall Design: The GalT BMT recipient is a GalT knockout mouse which recieved GalT gene transduced allo-bone marrow cells transplantation after sublethal irradiation. A heart of wild type C57BL/6 was heterotopically transplanted into the recipient after GalT BMT. Syngeneic Control recipient is a wild type C57BL/6 transplanted a heart of wild C57BL/6.
Project description:To elucidate mechanisms of cancer progression, we generated inducible human neoplasia in 3-dimensionally intact epithelial tissue. Gene expression profiling of both epithelia and stroma at specific time points during tumor progression revealed sequential enrichment of genes mediating discrete biologic functions in each tissue compartment. A core cancer progression signature was distilled using the increased signaling specificity of downstream oncogene effectors and subjected to network modeling. Network topology predicted that tumor development depends upon specific ECM-interacting network hubs. Blockade of one such hub, the b1 integrin subunit, disrupted network gene expression and attenuated tumorigenesis in vivo. Thus, integrating network modeling and temporal gene expression analysis of inducible human neoplasia provides an approach to prioritize and characterize genes functioning in cancer progression. There are 3 experiments in this study: (1a) a time course of human epidermal tissue transformed by oncogenic ER:H-Ras and IkBaM comprising days 0, 5, 20, and 35 post Ras activation using 4OHT; (1b) a matched time course of adjacent mouse stromal tissue during tumor progression; (2) Arrays comparing 4OHT-induced, Raf-1:ER/IkBaM transformed epidermal tissue and -4OHT controls; and (3) Arrays comparing Ras:ER/IkBaM grafts co-treated with 4OHT and either IgG control antibody or an anti-b1 integrin blocking antibody, P5D2 for 30 days. All arrays were done in biologic duplicate. Epidermal tissue co-expressing ER:H-RasG12V (ER:Ras) and IkBaM was regenerated on female scid/scid mouse recipients. Grafts were allowed to heal for at least 3 weeks before Ras activation via daily i.p. injections of 730ug of 4OHT (in 110ul of a corn oil/ethanol mixture). Duplicate grafts were harvested after 0, 5, 20, and 35 days of 4OHT treatment. Laser capture microdissection was utilized to independently isolate epithelial cells of the basal most layers and adajacent stromal tissue for each time point. RNA was subjected to one round of T7-based linear amplification (Ambion Message Amp II enhanced kit) and hybridized to either HG-U133A 2.0 or MG-430A affymetrix oligonucleotide arrays. Arrays from this experiment are labeled Ras and Stroma, respectively. The second experment was performed on Raf-1:ER/IkBaM expressing grafts with and without 4OHT treament for 30 days. The third experiment was done comparing ER:Ras/IkBaM grafts concomitantly treated with 4OHT and 1.5mg/week of either IgG control antibody or a mouse monoclonal blocking antibody against b1 integrin for 30 days. For experiments 2 and 3, HG-U133A 2.0 GeneChips were used; 4OHT treatment and RNA isolation/amplification was performed as above.
Project description:Uterine fibroid tissues are often compared to their matched myometrium in an effort to understand their pathophysiology, but it is not clear whether the myometria of uterine fibroid patients represent truly non-disease control tissues. We analyzed the transcriptomes of myometrial samples from non-fibroid patients (M) and from matched myometrial (MF) and fibroid (F) samples to determine whether there is a phenotypic difference between fibroid and non-fibroid myometria. Multidimensional scaling plots revealed that M samples clustered separately from both MF and F samples. A total of 1,169 differentially expressed genes (DEGs) (false discovery rate < 0.05) were observed in the MF comparison with M. Overrepresented Gene Ontology terms showed a high concordance of upregulated gene sets in MF compared to M, particularly extracellular matrix and structure organization. Gene set enrichment analyses showed that the leading-edge genes from the TGFβ signaling and inflammatory response gene sets were significantly enriched in MF. Overall comparison of the three tissues by three-dimensional principal component analyses showed that M, MF, and F samples clustered separately from each other and that a total of 732 DEGs from F vs M were not found in the F vs MF, which are likely understudied in the pathogenesis of uterine fibroids. These results suggest that the transcriptome of MF tissues are different from non-diseased myometrial tissues. Many dysregulated genes were not included in the F vs MF DEGs and may contain key genes for future investigations suggesting that fibroid studies should consider using not only matched myometrium but also non-diseased myometrium as the control.
Project description:We generated h-hepatocyte chimeric mice with livers that were predominantly repopulated with h-hepatocytes in a h-growth hormone (GH)-deficient state. Using microarray profiles, comparison between h-hepatocytes from h-GH-treated and untreated mice identified 14 GH-up-regulated and four GH-down-regulated genes, including IGF-1, SOCS2, NNMT, IGFLS, P4AH1, SLC16A1, and SRD5A1, and FADS1 and AKR1B10, respectively. The chimeric mice were treated or untreated with h-GH at 2.5 mg/kg b.w. /day for 2 weeks before sacrifice. Hepatocytes or liver tissue were isolated from the mouse livers and their cDNAs were used for microarray analysis.
Project description:Bone morphogenetic proteins (BMPs) are transforming growth factor β (TGFβ) family members that regulate the post-implantation and mid-gestation stages of pregnancy. In this study we discovered that signaling via activin-like kinase 3 (ALK3/BMPR1A), a BMP type 1 receptor, is necessary for blastocyst attachment. To understand the role of ALK3 in the luminal uterine epithelium, we obtained the gene expression profiles of isolated luminal uterine epithelium from 3.5dpc control and Alk3 cKO mice. Gene expression profiling of isolated luminal uterine epithelium from control and Alk3 cKO mice. two group comparison
Project description:The heavy metal Cadmium (Cd), a widespread environmental contaminant, poses serious hazards to human health, and is considered a metallohormone and carcinogen. In women with uterine fibroids, there is a significant association between blood Cd levels and fibroid tumor size. The aim of this study was to determine if benign uterine fibroid cells could be malignantly transformed in vitro by continuous Cd exposure, and if so, explore a molecular mechanism by which this could occur. We found when fibroid cells were exposed to 10 µM CdCl2 for 8 weeks, a robust and fast-growing Cd- Resistant Leiomyoma (CR-LM) culture was established. The CR-LM cells showed the ability to form viable colonies in soft agar, increased glycogen aggregates, enhanced cell motility, a higher percentage of cells in G2/M phase, and increased expression of the proliferation marker Ki-67. NanoString analysis showed downregulation of genes encoding for ECM components, such as collagens, fibronectins, laminins, and SLRP family proteins; whereas, genes involved in ECM degradation (MMP1, MMP3 and MMP10) were significantly upregulated. A volcano plot showed that the direction of expression changes of the top differentially genes favored cancer progression. Functional analysis by IPA predicted a significant inhibition of TGFβ signaling, leading to enhanced proliferation and attenuated fibrosis. Prolonged Cd exposure altered phenotypic characteristics and dysregulated genes in fibroid cells predicative of progression towards a cancer phenotype. Therefore, continuous Cd exposures changes the phenotype of benign fibroid cells in vitro and could possibly pose a health risk for women with uterine fibroids.