ABSTRACT: RNA fragments were isolated from mouse lung conditioned medium and analyzed by a microarray This analysis surveys RNAs, probably complexed with RNA-binding protein, secreted in the culture media
Project description:Primary human skeletal muscle cells (Lonza) were treated with LLC1 conditioned medium, LLC1 conditioned medium plus Calcitriol, LLC1 non-conditioned medium or LLC1 non-conditioned medium plus Calcitriol for a period of 24 hours prior to isolation of RNA.
Project description:We treated nine lines of breast cancer cells with conditioned medium derived from NAF and CAF to study the paracrine interaction beetween stroma and tumor. Gene expression profiles for breast cancer cell lines are reported after 72h of treatment with conditioned medium derived from NAF and CAF or with control media.
Project description:To identify cytokines secreted by pancreatic cancer cells, we compared the cytokine profiles of conditioned medium from pancreatic cancer cell Su.86.86 treated with or without JAG1 siRNA, using a Human Antibody Arrays 507 (L-507, Raybiotech)
Project description:Malignant melanoma is a complex genetic disease and the most aggressive form of skin cancer. Melanoma progression and metastatic dissemination fundamentally relies on the process of angiogenesis. Melanomas produce an array of angiogenic modulators that mediate pathological angiogenesis. Such tumor-associated modulators arbitrate the enhanced proliferative, survival and migratory responses exhibited by endothelial cells, in the hypoxic tumor environment. The current study focuses on melanoma-induced survival of endothelial cells under hypoxic conditions. Melanoma conditioned media were capable of enabling prolonged endothelial cell survival under hypoxia, in contrast with the conditioned media derived from melanocytes, breast and pancreatic tumors. To identify the global changes in gene expression and further characterize the pro-survival pathway induced in endothelial cells, we performed microarray analysis on endothelial cells treated with melanoma conditioned medium under normoxic and hypoxic conditions.
Project description:One challenging point in analysing cellular secretome collected as conditioned medium is cross contamination by cell culture media components, especially bovine serum proteins. A common approach for serum removal is to wash the cells, an alternative is to grow cells using serum-free conditions. Given that the sample processing may influence the phenotype of cells and thus the secretome, it is important to establish the optimal protocol for each cell type. In this study, we compared two methods for preparing conditioned medium from human adipocytes derived from mesenchymal stem cells. Cells were either washed twice with PBS or cultured the last four days of differentiation in serum free adipogenic medium. Gene expression of the cells was evaluated by using real time PCR and one dimensional LC-MS/MS was used to compare secreted proteins present in the culture supernatants. Surprisingly, results showed significant differences in gene expression patterns of the cells and in protein content of the conditioned media and suggested that PBS washes induced severe modifications of the phenotype of cells and thus changes in protein secretion profiles. These data emphasize the significant variations in protein species related to cell manipulations and underline the importance of procedure optimization prior to any proteomic investigation.
Project description:The endometrium contains a distinct population of immune cells consisting of 70% natural killer (NK) cells that undergo cyclic changes during the menstrual cycle. However, how these uterine NK (uNK) cells interact with uterine stromal cells (SC) remains unclear. We therefore investigated the paracrine effect of medium conditioned by uNK cells on the gene expression profile of endometrial SC in-vitro using a cDNA Microarray. Our results, verified by real-time PCR and ELISA, reveal that soluble factors from uNK cells substantially alter endometrial SC gene expression. The largest group of up-regulated genes found were chemokines and cytokines, including IL-15 and IL-15Rα. The latter could produce a niche for uNK cells allowing proliferation within and recruitment into the uterus, as seen in bone marrow. In addition, the most abundantly up-regulated genes, including IL-8, CCL8 and CXCL1 have also been shown to be stimulated by contact of SC with trophoblast, suggesting that uNK cells work synergistically to support the initial trophoblast migration during implantation. Overall this study demonstrates for the first time the paracrine communication between uNK cells and uterine SC, and adds to the understanding of how the uterine immune system contributes to the changes seen within the cycling endometrium. Keywords: Response of endometrial stromal cells to uNK conditioned medium This study was designed to identify the response of non-decidualised stromal cells from the endometrium, to soluble factors secreted by uterine NK cells. Endometrial stromal cells were isolated from 7 patients and treated with control medium or medium conditioned by uterine Nk cells. THE 'REF' COLUMN ON EACH ARRAY IS THE SIGNAL PRODUCED BY A COMMON REFERNCE RNA SAMPLE THAT WAS LABELLED AS A SINGLE BATCH SAMPLE AND HYBRIDISED TO ALL THE ARRAYS- IT IS A 'COMMON REFERENCE'. THE 'TEST' SAMPLE COMPRISES EACH INDIVIDUAL SAMPLE OF CELLS TREATED AS DESCRIBED IN THE SERIES SUBMISSION. FOR EXAMPLE; SAMPLE Y1 (gsm2435820) IS RNA FROM PATIENT 1 TREATED WITH CONTROL MEDIUM, SAMPLE G1 (GSM245371) IS RNA FROM PATIENT 1 TREATED WITH NK CONDITIONED MEDIUM THESE TWO SAMPLES THEREFORE FORM A PAIR- IE CELLS FROM THE SAME PATIENT TREATED WITH CONTROL OR NK-CONDITIONED MIDIUM. Y2,G2 ARE FROM PATIENT 2, ETC
Project description:Malignant melanoma is a complex genetic disease and the most aggressive form of skin cancer. Melanoma progression and metastatic dissemination fundamentally relies on the process of angiogenesis. Melanomas produce an array of angiogenic modulators that mediate pathological angiogenesis. Such tumor-associated modulators arbitrate the enhanced proliferative, survival and migratory responses exhibited by endothelial cells, in the hypoxic tumor environment. The current study focuses on melanoma-induced survival of endothelial cells under hypoxic conditions. Melanoma conditioned media were capable of enabling prolonged endothelial cell survival under hypoxia, in contrast with the conditioned media derived from melanocytes, breast and pancreatic tumors. To identify the global changes in gene expression and further characterize the pro-survival pathway induced in endothelial cells, we performed microarray analysis on endothelial cells treated with melanoma conditioned medium under normoxic and hypoxic conditions. Huvec cells were grown in melanoma conditioned medium or DMEM 10% FCS for 12 h under hypoxic or normoxic conditions. In order to identify the transcripts modulated by melanoma CM, samples treated with MCM were compared to those grown in DMEM alone.