小鼠造血各系细胞中氧化还原酶差异表达的研究

Differential Expression of Oxidative Reductases in Different Subsets of Mouse Hematopoietic Cells

造血细胞来源于造血干细胞(HSC),而HSC分化呈明显等级关系。细胞中活性氧簇(ROS)对HSC维持至关重要,细胞内过多ROS会造成HSC衰老。细胞主要通过氧化还原酶以及抗氧化物调节胞内ROS水平达到稳态,从而避免过多的ROS对细胞损伤。细胞内的氧化还原酶有多种,包括过氧化氢酶(catalase)、锰-超氧化物歧化酶(MnSOD)、谷胱苷肽过氧化物酶(GPX1)、NQO1〔NAD(P)H dehydrogenenasequinone 1〕和硫氧还蛋白还原酶(TXNRD1),其在呈等级分化的各类血细胞中的活性及其在这些细胞中参与ROS水平的调节作用还不清楚。本研究应用流式细胞分选技术,分选出小鼠造血各系细胞,用半定量实时PCR法检测氧化还原酶基因(Catalase、MnSOD、GPX1、Txrnd1和Nqo1)在造血各系细胞中的表达,进而筛选出参与HSC中ROS调控的重要的氧化还原酶。结果表明,以长期HSC(LT-HSC)作为参照,T细胞中catalase基因表达是LT-HSC的0.14倍,显著减低(P<0.05)。CLP和髓系细胞中MnSOD基因的表达分别是LT-HSC的0.56和0.47倍,显著减低(P<0.05)。ST-HSC、GMP和髓系细胞中GPX1基因表达分别为LT-HSC的1.79、2.96和2.07倍,显著增加(P<0.05);MEP、T淋巴细胞和B淋巴细胞中GPX1基因表达分别为LT-HSC的0.58、0.10和0.6倍,显著减低(P<0.05)。ST-HSC、MPP、CMP、GMP和髓系细胞中Txrnd1的表达分别为LT-HSC的3.36、3.18、4.19、6.39和4.27,显著增加(P<0.05);T淋巴细胞和B淋巴细胞中Txrnd1的表达分别为LT-HSC的0.016和0.56倍,显著减低(P<0.05)。ST-HSC、MPP、CMP、GMP、CLP和B淋巴细胞中Nqo1的表达为LT-HSC的0.30、0.17、0.25、0.10、0.04和0.01倍,显著减低(P<0.05)。结论:氧化还原酶在造血各系细胞的表达差异较大,提示在不同细胞中发挥主要作用的氧化还原酶种类不同。更为重要的是,Nqo1在LT-HSC中表达明显高于其他各系,提示其可能通过调控Nqo1的表达调节HSC功能。

Hematopoietic stem cells（HSC） are the source of all blood cells,which can differentiate into various hematopoietic hierarchy cells.Physiological level of reactive oxygen species（ROS） plays an important role in regulating functions of HSC as excessive ROS is harmful to HSC.Oxidative reductases and antioxidants can eliminate cellular ROS to maintain ROS homeostasis and thus avoid excessive ROS-caused damages.There are several types of oxidative reductases in cells such as catalase,manganese superoxide dismutase（MnSOD）,glutathione peroxidase 1（GPX1）,thioredoxin reductase 1（Txrnd1） and Nqo1（NAD（P）H dehydrogenenase quinone 1）.However,the functional roles of various oxidative reductases in regulating ROS level in hematopoietic cells remain unclear.This study was to investigate the expression patterns of these oxidative reductases in mouse hematopoietic cells that were sorted out via flow cytometry and to find out important oxidative reductases involving in HSC ROS regulation.The expression of various oxidative reductases was detected by semi-quantitative r eal-time PCR.The results showed that the expression level of catalase in T cell population was 0.14 times that in LT-HSC population（P 〈 0.05）.The expression levels of MnSOD in CLP population and myeloid cells were 0.56 and 0.47 times that in LT-HSC population respectively（P 〈 0.05）.The expression levels of GPX1 in ST-HSC,GMP,Myeloid cells,MEP,T lymphocytes and B lymphocytes were 1.79,2.96,2.07,0.58,0.10,0.6 times that in LT-HSC population respectively（P 〈 0.05）.The expression levels of Txrnd1 in ST-HSC,MPP,CMP,GMP,Myeloid cells,T lymphocytes and B lymphocytes were 3.36,3.18,4.19,6.39,4.27,0.016,0.56 time that in LT-HSC population,respectively（P 〈 0.05）.The expression levels of Nqo1 in ST-HSC,MPP,CMP,GMP,CLP and B cell were 0.30,0.17,0.25,0.10,0.04,0.01 times that in LT-HSC population,respectively（P 〈 0.05）.It is concluded that the expression levels of oxidative reductases（catalase,MnSOD,GPX1,Txrnd1 and Nqo1） in hematopoietic hierarchy cells are cell-type specific.It suggests that reductases may play divergent roles in various hematopoietic cell populations.More importantly,the expression level of Nqo1 in LT-HSC population significantly increased as compared with other cell populations,thereby suggesting its unique regulatory role in HSC.