Production of erythropoietic cells in vitro for continuous culture of Plasmodium vivax
Production of erythropoietic cells in vitro for continuous culture of Plasmodium vivax
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DOI：
10.1016/j.ijpara.2007.05.009
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发表时间：
2007-12-01
影响因子：
4
通讯作者：
Udomsangpetch, Rachanee
中科院分区：
文献类型：
--
作者：
Tasanee, Panichakul;Sattabongkot, Jetsumon;Udomsangpetch, Rachanee
间日疟原虫不能在连续培养中维持。为了克服间日疟原虫研究中的这一主要障碍，我们开发了一种体外方法，从新鲜分离的人脐血造血干细胞(HSCs)中分离出易感红细胞(RBC)前体细胞，并用促红细胞生成素激活HSCs，使其分化为类冷冻细胞。用细胞表面标志物(CD71、CD36、GPA和Fy6)监测红系细胞的分化和成熟。冷冻细胞培养10天后出现Duffy(+)网织红细胞，14-16天呈指数增长。从第10天开始，这些被称为生长性红细胞(GRBC)的红系细胞与直接从患者身上分离的感染间日疟原虫的血液进行共培养。用Giemsa染色和间日疟原虫特异性免疫荧光试验检测了14株间日疟原虫分离株中11株感染了寄生虫的gRBC。通过补充新鲜的gRBC，这些间日疟原虫培养物连续保持2周以上；其中一种在停止培养前保持85天。结果表明，HSCs来源的gRBC可为间日疟原虫的连续培养提供敏感的Duffy(+)网织红细胞。特别令人感兴趣的是，我们发现寄生虫能够入侵正常存在于骨髓中的有核红系细胞或红系细胞。间日疟原虫导致红细胞破坏，从而导致骨髓炎症的可能性需要解决。(C)2007澳大利亚寄生虫学会有限公司出版。保留所有权利。
Plasmodium vivax cannot be maintained in a continuous culture. To overcome this major obstacle to P. vivax research, we have developed an in vitro method to produce susceptible red blood cell (RBC) precursors from freshly isolated human cord hematopoietic stem cells (HSCs), which were activated with erythropoietin to differentiate into crythroid cells. Differentiation and maturation of erythroid cells were monitored using cell surface markers (CD71, CD36, GPA and Fy6). Duffy(+) reticulocytes appeared after 10 days of crythroid cell culture and exponentially increased to high numbers on days 14-16. Beginning on day 10 these erythroid cells, referred to as growing RBCs (gRBCs), were co-cultured with P. vivax-infected blood directly isolated from patients. Parasite-infected gRBCs were detected by Giemsa staining and a P. vivax-specific immunofluorescence assay in 11 out of 14 P. vivax isolates. These P. vivax cultures were continuously maintained for more than 2 weeks by supplying fresh gRBCs; one was maintained for 85 days before discontinuing the culture. Our results demonstrate that gRBCs derived in vitro from HSCs can provide susceptible Duffy(+) reticulocytes for continuous culture of P. vivax. Of particular interest, we discovered that parasites were able to invade nucleated erythroid cells or erythroblasts that are normally in the bone marrow. The possibility that P. vivax causes erythroblast destruction and hence inflammation in the bone marrow needs to be addressed. (C) 2007 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.