人巨细胞病毒对脐血多向祖细胞体外生长的影响及干预研究

目的探讨人巨细胞病毒(HCMV)对脐血多向造血祖细胞(CFU-Mix)体外增殖的影响及黄芪对此的保护作用.方法采用甲基纤维素半固体培养技术,培养、观察、计数HCMV-AD169感染的CFU-Mix集落数、抑制率、集落峰值及集落维持时间;用聚合酶链反应(PCR)和荧光定量PCR技术检测集落细胞内HCMV-AD169DNA.在110感染组中加入黄芪,观察黄芪对HCMV-AD169感染的CFU-Mix集落增殖抑制的干预作用.结果 HCMV-AD169感染的CFU-Mix集落数较对照组明显减少(P＜0.05),集落数随病毒感染滴度的增高而减少,抑制率随病毒感染滴度的增高而逐渐增加;感染组集落维持时间较对照组明显缩短(P＜0.01),但各组集落峰值出现时间无明显差异(P＞0.05);用PCR技术在感染组集落细胞内检测到HCMV-AD169DNA;在110感染组中加入黄芪后,CFU-Mix集落数明显提高,集落增殖提高率为38.9%,集落维持时间较110感染组明显延长(P＜0.01),荧光定量PCR技术检测集落细胞内HCMV-DNA copies较110感染组明显减少.结论在体外HCMV-AD169对CFU-Mix生长和集落形成有明显的抑制作用,HCMV能直接感染多向造血祖细胞,此可能与临床HCMV感染引起贫血、血小板减少和粒细胞减少有关;黄芪在体外有明显的抗HCMV作用.

Effect of human cytomegalovirus on proliferation of multipotential hematopoietic progenitors and intervention study in vitro

Objective To investigate the effect of human cytomegalovirus(HCMV) on proliferation of multipotential hematopoietic progenitors(CFU-Mix) with the presence of membranous milkvetch root (MMR) in vitro. Methods Colony forming unit-assay was applied to observe the effect of HCMV-AD169 strain on CFU-Mix of cord blood(CB). The techniques of PCR and fluorescence quantification PCR were used to demonstrate the existence of HCMV-DNA in the cells of cultured CFU-Mix. The suppression effect of HCMV on CFU-Mix was intervened in by adding MMR in 10-1HCMV group. Results The numbers of CFU-Mix colonies in HCMV-infected groups decreased significantly compared with that of control groups, and showed a decrease tendency with the increase of HCMV consentration. The suppression effect showed a dose-dependent fashion: The higher the HCMV consentration was, the more the colonies decreased. The peak of CFU-Mix (d10~12) was not significantly different between control groups and infection groups, but the duration of CFU-Mix in infected groups was significantly shorter than that in control groups. The numbers of CFU-Mix colonies increased significantly in 10-1 HCMV group with the presence of MMR. (4) HCMV-DNA was positively detected in the colony cells of virally infected groups by PCR, while negative in the control groups. HCMV-AD169 DNA copies decreased obviously by fluorescence quantification PCR in 10-1 HCMV group with the presence of MMR. Conclusions HCMV-AD169 strain inhibites the differentiation and proliferation of multipotential hematopoietic progenitors. HCMV may cause the suppression of hematopoiesis by direct infection of hematopoietic progenitors, which may be the main reason of HCMV infection associating with anemia, neutropenia and thrombocytopenia. The growth of multipotential hematopoietic progenitors after HCMV-AD169 infection is promoted by MMR, which suggests that MMR has an effect of anti-HCMV in vitro.