Project description:High ploidy large cytoplasmic megakaryocytes (LCM) are critical negative regulators of hematopoietic stem cells (HSC) and are responsible for platelet formation. Using a mouse knockout model with normal megakaryocyte numbers but essentially devoid of LCM (MK-LCM KO), we demonstrated a pronounced increase in bone marrow HSC concurrent with endogenous mobilization and extramedullary hematopoiesis. When HSC isolated from a MK-LCM KO microenvironment were transplanted in lethally irradiated mice, the absence of LCM increased HSC in BM, blood and spleen. Severe thrombocytopenia was observed in animals with diminished LCM, although there was no change in megakaryocyte ploidy distribution. In contrast, WT HSC-generated LCM regulated a normal HSC pool and prevented thrombocytopenia. The present label-free quantitative LC-MSMS data was used to determine proteins that are differentially expressed in bone marrow cells of MK-LCM WT versus MK-LCM KO mice.
Project description:Analysis of the transcriptome of bone marrow-derived macrophages from Cd28 KO mice. Methods: C57BL/6J (WT) and Cd28 KO (B6.129S2-Cd28tm1Mak/J) mice were used. Bone marrow-derived macrophages were obtained by flushing the femurs of 6- to 10-wk-old C57BL/6, and Cd28 KO mice, and culturing cells during 7 days in DMEM supplemented with 10% FCS and 50 mM 2-ME, containing human M-CSF (25 ng/ml), with cytokine addition every 2 days.
Project description:To fully elucidate the effects of long non-coding RNA 5730403I07Rik (lnc57Rik) and lncGM1082 on myeloid-derived suppressor cell (MDSCs), we generated lnc57Rik knockout (KO) and lncGM1082 ko mice. Then bone marrow cells were obtained from the femurs of C57BL/6, lnc57Rik KO, or lncGM1082 ko mice and cultured in RPMI-1640 medium supplemented with GM-CSF plus IL6 for 4 days. We further performed a high-throughput sequencing analysis in cultured MDSCs obtained from bone marrow of lnc57Rik ko, lncGM1082 ko and wild type (WT) mice.
Project description:Mammalian telomeres are formed by tandem repeats of the TTAGGG sequence, and shorten with each round of cell division in the absence of telomerase. Telomere shortening and dysfunction has been implicated in the pathology of several age-related diseases and premature ageing syndromes. Telomerase is important for telomere length maintenance. Telomerase RNA component, also known as TERC, is a component of telomerase. Terc knockout leads to telomerase deficiency and telomere shortening. Heterozygous telomerase-deficient (Terc+/-) mice were housed and bred for homozygous generation. ESC lines were generated with high efficiency from wild-type (WT, Terc+/+), heterozygous (Het, Terc+/-) and early- to late-generation (G1, G3 and G4) Terc-/- mouse blastocysts. Telomeres were shorter in Terc+/- ES cells than in WT ES cells, and further shortened from G1 to G4 Terc-/- ES cells. We took advantage of ES cell lines with various telomere lengths to investigate roles of telomere length on differentiation capacity of ES cells. We found that telomere length, but not telomerase activity, is required for differentiation of ES cells into epidermis. We performed microarray analysis to investigate differential gene expression profile at genome-wide levels between WT and G3/G4 Terc-/- (KO) mouse ES cells and during differentiation in vitro of WT and G4 Terc-/- mouse ES cells.
Project description:Unstimulated murine bone marrow derived macrophages cultured in L92 media from WT (4 biological replicates), Nrf2 KO (3 biological replicates) and Keap1 KD BMDM (3 biological replicates) were processed and analysed utilising DIA (label free) proteomic analysis. The Nrf2 KO mouse (DOI: 10.1006/bbrc.1997.6943 ) and Keap1 KD mouse ( DOI: 10.1128/MCB.01591-09) were previously published as noted.
Project description:RATIONALE: Radiation therapy uses high-energy x-rays to damage cancer cells. Drugs used in chemotherapy use different ways to stop cancer cells from dividing so they stop growing or die. Combining chemotherapy with bone marrow transplantation may allow the doctor to give higher doses of chemotherapy drugs and kill more tumor cells.
PURPOSE: Phase II trial to study the effectiveness of bone marrow transplantation in treating patients who have hematologic cancer.
Project description:RNA transcriptome difference between WT and SIRPa knockout (KO) bone marrow derived macrophages (BMDMs). To understand how SIRPa inhibits the phagocytotic capacity of macrophages, a mouse lacking SIRPa was generated, and the transcriptional profiles of BMDMs from wild-type and SIRPa KO mice were compared, using RNA sequencing.
Project description:Dyskeratosis congenita (DC) is an inherited multi-system disorder, characterized by oral leukoplakia, nail dystrophy, and abnormal skin pigmentation, as well as high rates of bone marrow failure, solid tumors, and other medical problems such as osteopenia. DC and telomere biology disorders (collectively referred to as TBD here) are caused by germline mutations in telomere biology genes leading to very short telomeres and limited proliferative potential of hematopoietic stem cells. We found that skeletal stem cells (SSCs) within the bone marrow stromal cell population (BMSCs, also known as bone marrow-derived mesenchymal stem cells), may contribute to the hematological phenotype. TBD-BMSCs exhibited reduced clonogenicity, reduced telomerase activity, spontaneous differentiation into adipocytes and fibrotic cells, and increased senescence in vitro. Upon in vivo transplantation into mice, TBD-BMSCs failed to form bone or support hematopoiesis, unlike normal BMSCs. TERC reduction (a TBD-associated gene) in normal BMSCs by siTERC-RNA recapitulated the TBD-BMSC phenotype by reducing proliferation and secondary colony forming efficiency, and by accelerating senescence in vitro. Microarray profiles of control and siTERC-BMSCs showed decreased hematopoietic factors at the mRNA level, and decreased secretion of factors at the protein level. These findings are consistent with defects in SSCs/BMSCs contributing to bone marrow failure in TBD.