乙型肝炎病毒感染时的骨髓造血干细胞

The relationship between of hematopoietic stem cells and hepatitis B virus infection

目的研究乙型肝炎患者骨髓造血干细胞中HBV的感染、复制及对造血干细胞增殖的影响。方法收集乙型肝炎患者9例,健康者7例骨髓液,磁珠分离仪分离纯化骨髓液CD34+细胞,取部分干细胞进行免疫组化和原位杂交,以感染HBV的干细胞(2.2.15细胞)为阳性对照,并设健康人骨髓干细胞为阴性对照。余两组干细胞分别分为两组,一组在含有干细胞生长因子(SCF)、酪氨酸激酶受体家族Ⅲ的配体(FLT3)、促血小板生成素(TPO)、白介素-3(IL-3)和10%FBS的IMDM培养基中孵育,另一组在无细胞因子的同样培养基中孵育。第0、1、6、12d进行PCR病毒载量检测并做细胞计数。结果患者组骨髓干细胞经免疫组化染色后为变为棕黄色与阳性对照组一致,经原位杂交后细胞染色为蓝紫色与阳性对照组一致,而阴性对照组均未染色。对患者组骨髓干细胞加细胞因子和无细胞因子第1、6、12d细胞计数分别比较,均明显少于正常对照组(t=0.818,P<0.05;t=3.599,P<0.05;t=2.967,P<0.05)。加细胞因子患者组第6、12d细胞内病毒载量明显高于不加细胞因子患者组(t=3.36,P<0.05;t=5.71,P<0.01)。结论 HBV可以感染骨髓造血干细胞并且可随干细胞的增殖不断复制。感染了HBV的干细胞增殖能力减弱。

Objective To investigate the impact of HBV infection on the bone marrow haematopoietic stem cell（HSC）.Methods Nine hepatitis B patients and 7 healthy donors were included in this study.Bone marrow samples were collected and the CD34＋ cells were purified by mini-MACS.HSCs were studied by immunohistochemiststy and hybridization in situ.HepG2.21 and HSC of healthy donors were used as a positive and negative control,respectively.HSCs were incubated with or without cytokine markers（SCF,FLT3,TPO,IL-3）.HBVDNA was detected by PCR and cells were counted on day 0,1,6 and 12 day.Results The immunohistochemistry result of HSCs in HBV patients showed blue-purple color among positive controls compared to healthy control group.The number of HSCs in the patient group was less than healthy control group on day 1,6 and 12 after incubation with or without cytokines（t=0.818,P0.05;t=3.599,P0.05;t=2.967,P0.05）.HBVDNA was higher after incubation with cytokine in the patient group than healthy control group on day 6 and 12（t=3.36,P0.05;t=5.71,P0.01）.Conclusion HBV infects and replicate in bone marrow inhibiting the proliferation ability of HSCs.