Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Transcription profiling of mouse cartilage tumours IGFBP-3 reveals is regulated by Gli signaling in cartilage tumors.

ABSTRACT: Mice that develop benign cartilage lesions due to overexpression of Gli2 in chondrocytes developed lesions similar to chondrosarcomas when also deficient in p53. Gli2 overexpression and p53 deficiency had opposing effects on chondrocyte differentiation, but had additive effects negatively regulating apoptosis. Regulation of Igfbp3 expression and IGF signaling by Gli and p53 integrated their effect on apoptosis. Treatment of human chondrosarcomas or fetal mouse limbs explants with IGFBP3 or by blocking IGF increased the apoptosis rate, and mice expressing Gli2 developed substantially fewer tumors when also deficient for Igf2. IGF signaling meditated apoptosis regulates the progression to malignant chondrosarcoma. Experiment Overall Design: The CH10T1/2 cell line was stably transfected with either the R150C variant PTHR1 present in enchondromas (Hopyan, S., Gokgoz, N., Poon, R., Gensure, R. C., Yu, C., Cole, W. G., Bell, R. S., Juppner, H., Andrulis, I. L., Wunder, J. S., and Alman, B. A. (2002). A mutant PTH/PTHrP type I receptor in enchondromatosis. Nat Genet 30, 306-310) or a wild type PTHR1, as previously reported (Mau, E., Whetstone, H., Yu, C., Hopyan, S., Wunder, J. S., and Alman, B. A. (2007). PTHrP regulates growth plate chondrocyte differentiation and proliferation in a Gli3 dependent manner utilizing hedgehog ligand dependent and independent mechanisms. Dev Biol 305, 28-39). Cells were grown overnight in serum free media and treated with either 10â?? 7 M PTHrP (Bachem, King of Prussia, Pennsylvania, USA), or carrier alone for one hour. RNA was isolated from the cells CH10T1/2 cells expressing either the R150C variant PTHR1 or wild type PTHR1. RNA was converted to double-stranded cDNA using Superscript (Gibco-Invitrogen) with a T7-(dT)24 primer, which was then transcribed to biotinylated complementary RNA (cRNA) by incorporating biotin-CTP and biotin-UTP using Enzo BioArray High Yield RNA labeling kit (Enzo Diagnostics, New York, NY). The cRNA labeling and hybridizations were then performed according to Affymetrix GeneChip Protocol (Affymetrix Inc., Santa Clara, CA). The chips were scanned for fluorescence signal detection. Affymetrix - GeneChipÂ® Mouse Genome 430A 2.0 Array was utilized.

ORGANISM(S): Mus musculus

SUBMITTER: benjamin alman

PROVIDER: E-GEOD-15118 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Prolonged skeletal unloading through bedrest results in bone loss similar to that observed in elderly osteoporotic patients, but with an accelerated timeframe. This rapid effect on weight-bearing bones is also observed in astronauts who lose up to 2% of their bone mass per month spent in Space. Despite important implications for Spaceflight travellers and bedridden patients on Earth, the exact mechanisms involved in disuse osteoporosis have not been elucidated. Parathyroid hormone-related protein (PTHrP) regulates many physiological processes including skeletal development, and has been proposed as a gravisensor. To investigate the role of PTHrP in microgravity-induced bone loss, trabecular osteoblasts (TOs) from Pthrp+/+ and -/- mice were exposed to simulated microgravity for 6 days. Viability of TOs decreased in inverse proportion to PTHrP expression levels. Microarray analysis of Pthrp+/+ TOs after 6 days at 0g revealed expression changes in genes encoding prolactins,apoptosis and survival molecules, bone metabolism and extra-cellular matrix composition proteins, chemokines, IGF family and Wnt-related signalling molecules. Importantly, 88% of 0g-induced expression changes in Pthrp+/+ cells overlap those observed in Pthrp-/- cells in normal gravity. Pulsatile treatment with PTHrP1-36 peptide during microgravity exposure reversed a large proportion of 0g-induced changes in Pthrp+/+ TOs. Our results confirm PTHrP efficacy as an anabolic agent to prevent microgravity-induced cell death in TOs. Total RNA samples extracted from Pthrp+/+and -/- trabecular osteoblasts (TOs) exposed for 6 days to simulated 0g in Synthecon rotating cell, or left 6 days in culture at 1g. Cells had either been treated with a pulsatile treatment (2 h/day) of PTHrP1-36 peptide (10-8M) or received a change in growth medium. In total: 8 different conditions with 2 replicates each, i.e. Pthrp+/+ TOs at 0g or 1g with or without PTHrP1-36 treatment, and Pthrp-/- TOs at 0g or 1 g,with or without PTHrP1-36 treatment.

Project description:Previously, we found low-carbohydrate diets slowed prostate cancer (PC) growth and increased survival vs. a Western diet in mice, by inhibiting the insulin/IGF-1 axis. Thus, we tested whether modifying carbohydrate quality to lower glycemic index (GI) without changing quantity results in similar benefits as with reduced quantity. Male SCID mice injected with LAPC-4 cells were single-housed and randomized when their tumors reached 200mm3 on average to a LoGI (48% carbohydrate kcal, from Hylon-VII) or HiGI Western diet (48% carbohydrate kcal, from sucrose). Body weight and tumor volume were measured weekly. Body composition was assessed 35 days after randomization. Blood glucose and serum insulin, IGF-1 and IGFBP3 were measured at study end when tumor volumes reached 800mm3. We analyzed gene expression of mice tumors by RNA-sequencing and human tumors using the Prostate Cancer Transcriptome Atlas. There were no significant differences in tumor volume (P>0.05), tumor proliferation (P=0.29), and overall survival (P=0.15) between groups. At 35 days after randomization, the LoGI group had 30% lower body fat (P=0.007) despite similar body weight (P=0.58). At sacrifice, LoGI mice had smaller livers (P<0.001) and lower glucose (P=0.15), insulin (P=0.11), IGF-1 (P=0.07) and IGF-1:IGFBP3 ratio (P=0.05), and higher IGFBP3 (P=0.09) vs. HiGI, although none of these metabolic differences reached statistical significance. We observed differential gene expression and pathway enrichment in mice tumors by diet. The most upregulated and downregulated gene in the LoGI group showed expression patterns more closely resembling expression in human benign prostate tissue vs. PC. In this single mouse xenograft model, consuming a low GI diet did not delay PC growth or survival vs. a high GI diet despite suggestions of decreased activation of the insulin/IGF-1 pathway. These data suggest that improving carbohydrate quality alone while consuming a high carbohydrate diet may not effectively slow PC growth.

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Transcription profiling of mouse cartilage tumours IGFBP-3 reveals is regulated by Gli signaling in cartilage tumors.

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