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Identification of Cellular Compositions in Different Microenvironments and Their Potential Impacts on Hematopoietic Stem Cells HSCs Using Single-Cell RNA Sequencing with Systematical Confirmation
Hematopoietic stem cells (HSCs) are stem cells that can differentiate into various blood cells and have long-term self-renewal capacity. At present, HSC transplantation is an effective therapeutic means for many malignant hematological diseases, such as aplastic hematological diseases and autoimmune...
Autores principales: | , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10671877/ https://www.ncbi.nlm.nih.gov/pubmed/38004297 http://dx.doi.org/10.3390/life13112157 |
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author | Chi, Yanan Yang, Guanheng Guo, Chuanliang Zhang, Shaoqing Hong, Lei Tang, Huixiang Sang, Xiao Wang, Jie Ma, Ji Xue, Yan Zeng, Fanyi |
author_facet | Chi, Yanan Yang, Guanheng Guo, Chuanliang Zhang, Shaoqing Hong, Lei Tang, Huixiang Sang, Xiao Wang, Jie Ma, Ji Xue, Yan Zeng, Fanyi |
author_sort | Chi, Yanan |
collection | PubMed |
description | Hematopoietic stem cells (HSCs) are stem cells that can differentiate into various blood cells and have long-term self-renewal capacity. At present, HSC transplantation is an effective therapeutic means for many malignant hematological diseases, such as aplastic hematological diseases and autoimmune diseases. The hematopoietic microenvironment affects the proliferation, differentiation, and homeostasis of HSCs. The regulatory effect of the hematopoietic microenvironment on HSCs is complex and has not been thoroughly studied yet. In this study, we focused on mononuclear cells (MNCs), which provided an important microenvironment for HSCs and established a methodological system for identifying cellular composition by means of multiple technologies and methods. First, single-cell RNA sequencing (scRNA-seq) technology was used to investigate the cellular composition of cells originating from different microenvironments during different stages of hematopoiesis, including mouse fetal liver mononuclear cells (FL-MNCs), bone marrow mononuclear cells (BM-MNCs), and in vitro-cultured fetal liver stromal cells. Second, bioinformatics analysis showed a higher proportion and stronger proliferation of the HSCs in FL-MNCs than those in BM-MNCs. On the other hand, macrophages in in vitro-cultured fetal liver stromal cells were enriched to about 76%. Differential gene expression analysis and Gene Ontology (GO) functional enrichment analysis demonstrated that fetal liver macrophages have strong cell migration and actin skeleton formation capabilities, allowing them to participate in the hematopoietic homeostasis through endocytosis and exocytosis. Last, various validation experiments such as quantitative real-time PCR (qRT-PCR), ELISA, and confocal image assays were performed on randomly selected target genes or proteins secreted by fetal liver macrophages to further demonstrate the potential relationship between HSCs and the cells inhabiting their microenvironment. This system, which integrates multiple methods, could be used to better understand the fate of these specific cells by determining regulation mechanism of both HSCs and macrophages and could also be extended to studies in other cellular models. |
format | Online Article Text |
id | pubmed-10671877 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-106718772023-11-02 Identification of Cellular Compositions in Different Microenvironments and Their Potential Impacts on Hematopoietic Stem Cells HSCs Using Single-Cell RNA Sequencing with Systematical Confirmation Chi, Yanan Yang, Guanheng Guo, Chuanliang Zhang, Shaoqing Hong, Lei Tang, Huixiang Sang, Xiao Wang, Jie Ma, Ji Xue, Yan Zeng, Fanyi Life (Basel) Article Hematopoietic stem cells (HSCs) are stem cells that can differentiate into various blood cells and have long-term self-renewal capacity. At present, HSC transplantation is an effective therapeutic means for many malignant hematological diseases, such as aplastic hematological diseases and autoimmune diseases. The hematopoietic microenvironment affects the proliferation, differentiation, and homeostasis of HSCs. The regulatory effect of the hematopoietic microenvironment on HSCs is complex and has not been thoroughly studied yet. In this study, we focused on mononuclear cells (MNCs), which provided an important microenvironment for HSCs and established a methodological system for identifying cellular composition by means of multiple technologies and methods. First, single-cell RNA sequencing (scRNA-seq) technology was used to investigate the cellular composition of cells originating from different microenvironments during different stages of hematopoiesis, including mouse fetal liver mononuclear cells (FL-MNCs), bone marrow mononuclear cells (BM-MNCs), and in vitro-cultured fetal liver stromal cells. Second, bioinformatics analysis showed a higher proportion and stronger proliferation of the HSCs in FL-MNCs than those in BM-MNCs. On the other hand, macrophages in in vitro-cultured fetal liver stromal cells were enriched to about 76%. Differential gene expression analysis and Gene Ontology (GO) functional enrichment analysis demonstrated that fetal liver macrophages have strong cell migration and actin skeleton formation capabilities, allowing them to participate in the hematopoietic homeostasis through endocytosis and exocytosis. Last, various validation experiments such as quantitative real-time PCR (qRT-PCR), ELISA, and confocal image assays were performed on randomly selected target genes or proteins secreted by fetal liver macrophages to further demonstrate the potential relationship between HSCs and the cells inhabiting their microenvironment. This system, which integrates multiple methods, could be used to better understand the fate of these specific cells by determining regulation mechanism of both HSCs and macrophages and could also be extended to studies in other cellular models. MDPI 2023-11-02 /pmc/articles/PMC10671877/ /pubmed/38004297 http://dx.doi.org/10.3390/life13112157 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Chi, Yanan Yang, Guanheng Guo, Chuanliang Zhang, Shaoqing Hong, Lei Tang, Huixiang Sang, Xiao Wang, Jie Ma, Ji Xue, Yan Zeng, Fanyi Identification of Cellular Compositions in Different Microenvironments and Their Potential Impacts on Hematopoietic Stem Cells HSCs Using Single-Cell RNA Sequencing with Systematical Confirmation |
title | Identification of Cellular Compositions in Different Microenvironments and Their Potential Impacts on Hematopoietic Stem Cells HSCs Using Single-Cell RNA Sequencing with Systematical Confirmation |
title_full | Identification of Cellular Compositions in Different Microenvironments and Their Potential Impacts on Hematopoietic Stem Cells HSCs Using Single-Cell RNA Sequencing with Systematical Confirmation |
title_fullStr | Identification of Cellular Compositions in Different Microenvironments and Their Potential Impacts on Hematopoietic Stem Cells HSCs Using Single-Cell RNA Sequencing with Systematical Confirmation |
title_full_unstemmed | Identification of Cellular Compositions in Different Microenvironments and Their Potential Impacts on Hematopoietic Stem Cells HSCs Using Single-Cell RNA Sequencing with Systematical Confirmation |
title_short | Identification of Cellular Compositions in Different Microenvironments and Their Potential Impacts on Hematopoietic Stem Cells HSCs Using Single-Cell RNA Sequencing with Systematical Confirmation |
title_sort | identification of cellular compositions in different microenvironments and their potential impacts on hematopoietic stem cells hscs using single-cell rna sequencing with systematical confirmation |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10671877/ https://www.ncbi.nlm.nih.gov/pubmed/38004297 http://dx.doi.org/10.3390/life13112157 |
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