Schwab Ute E, Tallmadge Rebecca L, Matychak Mary Beth, Felippe M Julia B
Am J Vet Res. 2017 Oct;78(10):1215-1228. doi: 10.2460/ajvr.78.10.1215.
OBJECTIVE To develop an in vitro system for differentiation of equine B cells from bone marrow hematopoietic progenitor cells on the basis of protocols for other species. SAMPLE Bone marrow aspirates aseptically obtained from 12 research horses. PROCEDURES Equine bone marrow CD34 cells were sorted by use of magnetic beads and cultured in medium supplemented with cytokines (recombinant human interleukin-7, equine interleukin-7, stem cell factor, and Fms-like tyrosine kinase-3), murine OP9 stromal cell preconditioned medium, and equine fetal bone marrow mesenchymal stromal cell preconditioned medium. Cells in culture were characterized by use of flow cytometry, immunocytofluorescence microscopy, and quantitative reverse-transcriptase PCR assay. RESULTS For these culture conditions, bone marrow-derived equine CD34 cells differentiated into CD19IgM B cells that expressed the signature transcription factors early B-cell factor and transcription factor 3. These conditions also supported the concomitant development of autologous stromal cells, and their presence was supportive of B-cell development. CONCLUSIONS AND CLINICAL RELEVANCE Equine B cells were generated from bone marrow aspirates by use of supportive culture conditions. In vitro generation of equine autologous B cells should be of use in studies on regulation of cell differentiation and therapeutic transplantation.
目的 基于其他物种的方案，开发一种从马骨髓造血祖细胞分化出马B细胞的体外系统。样本 从12匹研究用马无菌采集的骨髓抽吸物。方法 使用磁珠分选马骨髓CD34细胞，并在补充了细胞因子（重组人白细胞介素-7、马白细胞介素-7、干细胞因子和Fms样酪氨酸激酶-3）、小鼠OP9基质细胞预处理培养基和马胎儿骨髓间充质基质细胞预处理培养基的培养基中培养。通过流式细胞术、免疫细胞荧光显微镜检查和定量逆转录聚合酶链反应分析对培养的细胞进行表征。结果 在这些培养条件下，源自骨髓的马CD34细胞分化为表达标志性转录因子早期B细胞因子和转录因子3的CD19IgM B细胞。这些条件也支持自体基质细胞的同时发育，并且它们的存在有助于B细胞发育。结论及临床意义 通过使用支持性培养条件从骨髓抽吸物中生成了马B细胞。马自体B细胞的体外生成在细胞分化调节和治疗性移植研究中应具有应用价值。
