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Home > Archive>Volume 33, Issue 6, 2023 >35-45,61. DOI:10. 3969 / j.issn.1671-7856. 2023. 06. 005
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Effect of Marcksl1 gene knockout on adult hematopoiesis in mice
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1. Comparative Medicine Center, Peking Union Medical College, Institute of Laboratory Animal Science, Chinese Academy of Medical Sciences, Beijing Engineering Research Center for Experimental Animal Models of Human Diseases, Key Laboratory of Human Diseases Comparative Medicine, National Health Commission of China, Beijing 100021, China. 2. Comparative Medicine Center, Peking Union Medical College, Institute of Laboratory Animal Science, Chinese Academy of Medical Sciences, National Human Diseases Animal Model Resource Center, Beijing 100021

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R-33

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    Abstract:

    Objective To establish hematopoietic system-specific Marcksl1 knockout mice and explore the effect of Marcksl1 gene deletion on hematopoiesis. Methods Using fetal liver competitive transplantation of E15. 5 Marcskl1 gene knockout mice, we analyzed the proportions of various hematopoietic cell types in peripheral blood at 1, 2, 3, and 4 months after fetal liver transplantation. KSL cells were sorted by flow cytometry and analyzed by RNA-seq. We used CRISPR/ Cas9 technology to generate Marcksl1 conditional knockout mice and obtained hematopoietic system-specific Marcksl1 knockout mice by crossing with Vav1-Cre mice. The genotype of the produced mice was confirmed by PCR and Sanger sequencing. Real-time PCR was used to assess Marcskl1 mRNA expression in bone marrow and hematopoietic stem cells. Flow cytometry was used to analyze the proportions of various hematopoietic cell types in bone marrow, peripheral blood, spleen, and thymus. The effect of Marcksl1 gene knockout on hematopoietic reconstitution was analyzed by competitive bone marrow transplantation. Results Marcksl1 gene knockout decreased the proportion of B cells, increased the proportion of myeloid cells, decreased CLP and CMP, and increased GMP after fetal liver transplantation. RNA-seq showed that 252 genes were upregulated and 400 genes were downregulated. GO analysis showed that the differentially expressed genes were significantly enriched in the immune response, plasma membrane, and low-pressure gate calcium channel activity. KEGG analysis showed that the differentially expressed genes were significantly enriched in hematopoietic lineage differentiation and cytokine-cytokine receptor interaction-related signaling pathways. We generated hematopoietic system-specific Marcksl1 knockout mice successfully. Flow cytometry showed that Marcksl1 deletion did not affect the steady-state hematopoietic functions of adult mice, but it did affect the hematopoietic reconstitution ability after competitive bone marrow transplantation. Conclusions We successfully established hematopoietic system-specific Marcksl1 knockout mice and found that Marcksl1 gene knockout does not affect stable hematopoiesis, but affects hematopoietic reconstitution. The underlying mechanism requires further investigation. This study provides an animal model to examine the gene function of Marcksl1 in adult hematopoiesis and other aspects.

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History
  • Received:March 15,2023
  • Online: August 18,2023
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