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RNAseq for Nestin+ bone marrow stromal cells in normal and MLL-AF9 mice

ABSTRACT: Cumulative evidence points out to the importance of the bone microenvironment for leukaemic development. Our preliminary results show that bone marrow stem cells, identified by the expression of the intermediate filament protein nestin (Nestin+ cells), provide support and chemoresistance to leukaemic cells. The rationale for these experiments is that leukaemic/Nestin+ cells crosstalk might regulate Nestin+ cells genetic profile to become more supportive elements for leukaemia. Nestin expression level distinguish two different Nestin+ populations, which seem to possess distint characteristics. Therefore, we would like to separately study high-expressing (Nestin high) and low-expressing Nestin cells (Nestin low). From our in vitro results, Nestin+ cells seem to be providing detoxifying tools to leukaemic cells such as antioxidant aminoacids and enzymes. The aim of the project is to identify pathways and genes differentially regulated in Nestin+ high and/or low cells in a leukaemic background.

ORGANISM(S): Mus musculus

PROVIDER: GSE140135 | GEO | 2020/11/24

REPOSITORIES: GEO

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Project description:H1299 cells were stably transfected with the Oct4 promoter/GFP or Nestin promoter/GFP reporter vectors. By FACS, 106 cells expressing high levels of GFP were isolated and placed into cell culture for twenty-four hours. Total RNA was used. Bone morphogenetic proteins (BMP) are aberrantly expressed in most lung carcinomas. BMPs mediate cell fate decisions and self-renewal of stem cells. Inhibition of BMP signaling decreases the growth and induces cell death of lung cancer cells lines. It is not known whether the BMP signaling cascade is growth promoting in lung cancer cells expressing the stem cell markers Oct4 and/or nestin. Lung cancer cells expressing Oct4 or nestin were isolated from lung cancer cell lines by stably transfecting the Oct4 promoter or Nestin promoter expression vectors that activate the green fluorescent protein reporter. Our studies support that lung cancer cells activating the Oct4 or nestin promoter are different cell populations. Microarray and quantitative RT-PCR demonstrated that the expression levels of specific stem cell markers were different between the isolated Oct4 and nestin cells. Both the Oct4 and nestin populations were more tumorigenic that controls but histologically they were quite different. The isolated Oct4 and nestin cells also responded differently to inhibition of BMP signaling. Blockade of BMP signaling with the BMP receptor antagonist DMH2 caused significant growth inhibition in both the Oct4 and nestin cell populations but only increased cell death in the nestin population. DMH2 also induced the expression of nestin in the Oct4 population but not in the nestin cells. We also show that BMP signaling is an important regulator of the inhibitor differentiation proteins Id1 and Id3 in Oct4 and nestin cell populations Lung cancer cells expressing Oct4 or nestin were isolated and transfected with Oct4 or Nestin promoter expression vectors that activate the green fluorescent protein reporter. Microarray and quantitative RT-PCR were performed to study the differentially expressed genes. BMPs mediate cell fate decisions and self-renewal of stem cells were investigated.

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RNAseq for Nestin+ bone marrow stromal cells in normal and MLL-AF9 mice

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