Project description:We took advantage of a dystrophic mouse model of transient macrophage-depletion, mdxITGAM-DTR mice, in order to analyze the role of macrophage in skeletal muscle regeneration. We generated the transcriptome of satellite cells (SCs) and alpha7Sca1 cells purified by cell sorting from mdxITGAM-DTR mice. The mice were treated, by intramuscular injection, with PBS, as vehicle, or with Diphtheria toxin (DT) in order to achieve the macrophage depletion form hind-limb muscle We described a shift in identity of muscle stem cells dependent on the crosstalk between macrophages and satellite cells. Indeed macrophage depletion determines an exacerbated dystrophic phenotype associated with adipogenic conversion of SCs and reduction of the SC pool.
Project description:KPR tumors were subcutaneously implanted in Lrrc15DTRGFPwt/wt (DTR–) or Lrrc15DTRGFPwt/ki (DTR+) mice and DT treatment was initiated in both DTR– and DTR+ mice when tumors reached a mean volume of 100-200mm3. scRNAseq of CD24-CD45- stromal cells from a total of 40 tumor-bearing animals across both DTR– and DTR+ genotypes was carried out at four different time points including 10 days post tumor implantation and immediately prior to initiation of DT treatment (IOT) (referred to as day 0) and on days 7, 14, and 21 post IOT.
Project description:CLEC4C-DTR exhibit exacerbated contact hypersensitivty to the FITC hapten. Here we assess the transcriptional differences in whole skin of FITC challenged CLEC4C-DTR+ mice and DTR- mice treated with diptheria toxin. Overall, CLEC4C-DTR upregualated 439 genes and downregulated 152 genes compared to transgene negative controls, filtering output on 2 fold expression level difference and FDR<0.05.
Project description:To identify the target genes of Dnmt1 in skeletal muscle satellite cells, we generated satellite cell specific Dnmt1 KO using Pax7-CreERT2 and Dnmt1 flox mice. After Tamoxifen (cKO) or vehicle (Control) treatment for 5 consecutive days, mice were euthanized and muscle satellite cells were collected by FACS sorting and total RNA were extracted.
Project description:Skin draining lymph node dendritic cell populations are known to be diverse and distinguishable via their migratory or resident properties as well as by lineage. We analyzed these populations at steady state and during the sensitization phase of the Th2-driven FITC-hapten contact hypersensitivity (CHS) model. We also analyzed the dendritic cells populations of CLEC4C-DTR+ mice treated with diptheria toxin, which demonstrate exacerbated pathology in the FITC-CHS model. We demonstrate the diversity of dendritic cell populations at steady state and during type 2 inflammation, and surprisingly, reveal a skewing of the DC populations in DT treated CLEC4C-DTR+ mice, with an enrichment of DC2 migratory populations and depletion of CD326+ migratory cells in addition to pDC.
Project description:We studied the impact of DT treatment (Treg depletion) on NK cells (CD3-, NKp46+, DX5+) in axillary TDLNs and lungs of tumor bearing Foxp3GFP-DTR mice . Here we generated RNAseq data from sorted NK cells from DT or PBS treated Foxp3GFP-DTR mice bearing 100mm2 mammary tumors from lungs, axillary TDLNs
Project description:Following skeletal muscle injury, muscle stem cells (satellite cells) are activated, proliferate, and differentiate to form myofibers. We show that mRNA decay protein AUF1 regulates satellite cell function through targeted degradation of specific mRNAs. AUF1 targets certain mRNAs containing 3 AU-rich elements (AREs) for rapid decay. Auf1-/- (KO) mice undergo accelerated skeletal muscle wasting with age and impaired muscle repair following injury. Satellite cell mRNA analysis and regeneration studies demonstrate that auf1-/- satellite cell self-renewal is impaired due to increased stability and overexpression of ARE-mRNAs. Control of ARE-mRNA decay by AUF1 and potentially other ARE-binding proteins represents a mechanism for adult stem cell regulation and is implicated in human muscle wasting diseases. We report the RNA transcript expression profiles from sorted satellite cells isolated from wild type (WT) and AUF1-null (KO) mice hindlimb muscles Examination of RNA transcript expression from satellite cells of two genotypes Please note that mice are bred through a C57BL/6 strain of 129 background.
Project description:Adult muscle stem cells (satellite cells) are required for adult skeletal muscle regeneration. A proper balance between quiescence, proliferation, and differentiation is essential for the maintenance of the satellite cell pool and their regenerative function. Although the ubiquitin-proteasome is required for most protein degradation in mammalian cells, how its dysfunction affects tissue stem cells remains unclear. Here, we investigated the function of the proteasome in satellite cells using mice lacking the crucial proteasomal component, Rpt3. Ablation of Rpt3 in satellite cells decreased proteasome activity. Proteasome dysfunction in Rpt3-deficient satellite cells impaired their ability to proliferate, survive and differentiate, resulting in defective muscle regeneration. We found that inactivation of proteasomal activity induced proliferation defects and apoptosis in satellite cells. Mechanistically, insufficient proteasomal activity upregulated the p53 pathway, which caused cell-cycle arrest. Our findings delineate a critical function of the proteasome system in maintaining satellite cells in adult muscle.