Gene Therapy Research Center, Digestive Disease Research Institute, Tehran University of Medical Sciences, Tehran, Iran.
Department of Hematology, School of Allied Medical Sciences, Tehran University of Medical Sciences, Tehran, Iran.
BMC Immunol. 2021 Jul 3;22(1):43. doi: 10.1186/s12865-021-00435-7.
The successful ex vivo expansion of T-cells in great numbers is the cornerstone of adoptive cell therapy. We aimed to achieve the most optimal T-cell expansion condition by comparing the expansion of T-cells at various seeding densities, IL-2 concentrations, and bead-to-cell ratios. we first expanded the peripheral blood mononuclear cells (PBMCs) of a healthy donor at a range of 20 to 500 IU/mL IL-2 concentrations, 125 × 10 to 1.5 × 10 cell/mL, and 1:10 to 10:1 B:C (Bead-to-cell) ratios and compared the results. We then expanded the PBMC of three healthy donors using the optimized conditions and examined the growth kinetics. On day 28, CD3, CD4, and CD8 expression of the cell populations were analyzed by flow cytometry.
T-cells of the first donor showed greater expansion results in IL-2 concentrations higher than 50 IU/mL compared to 20 IU/mL (P = 0.02). A seeding density of 250 × 10 cell/mL was superior to higher or lower densities in expanding T-cells (P = 0.025). Also, we witnessed a direct correlation between the B:C ratio and T-cell expansion, in which, in 5:1 and 10:1 B:C ratios T-cell significantly expanded more than lower B:C ratios. The results of PBMC expansions of three healthy donors were similar in growth kinetics. In the optimized condition, 96-98% of the lymphocyte population expressed CD3. While the majority of these cells expressed CD8, the mean expression of CD4 in the donors was 19.3, 16.5, and 20.4%.
Our methodology demonstrates an optimized culture condition for the production of large quantities of polyclonal T-cells, which could be useful for future clinical and research studies.
成功地大量扩增 T 细胞是过继细胞疗法的基石。我们旨在通过比较不同接种密度、IL-2 浓度和珠细胞比的 T 细胞扩增情况，找到最佳的 T 细胞扩增条件。我们首先在 20 至 500 IU/mL 的 IL-2 浓度、125×10 至 1.5×10 细胞/mL 和 1:10 至 10:1 的 B:C（珠细胞比）范围内，对一名健康供体的外周血单核细胞（PBMC）进行扩增，并比较结果。然后，我们使用优化的条件对三名健康供体的 PBMC 进行扩增，并检查生长动力学。在第 28 天，通过流式细胞术分析细胞群体中 CD3、CD4 和 CD8 的表达。
与 20 IU/mL 相比，第一供体的 T 细胞在高于 50 IU/mL 的 IL-2 浓度下显示出更大的扩增效果（P=0.02）。250×10 细胞/mL 的接种密度优于较高或较低的细胞密度（P=0.025）。此外，我们还观察到 B:C 比与 T 细胞扩增之间存在直接相关性，其中在 5:1 和 10:1 的 B:C 比下，T 细胞的扩增明显高于较低的 B:C 比。三名健康供体的 PBMC 扩增结果在生长动力学上相似。在优化条件下，96-98%的淋巴细胞群体表达 CD3。虽然这些细胞中的大多数表达 CD8，但供体的 CD4 平均表达水平为 19.3、16.5 和 20.4%。
我们的方法为生产大量多克隆 T 细胞提供了优化的培养条件，这可能对未来的临床和研究研究有用。
