Department of Biomedical Sciences, Seoul National University Graduate School, Seoul, Republic of Korea; National Research Laboratory for Stem Cell Niche, Seoul National University College of Medicine, Seoul, Republic of Korea; Innovative Research Institute for Cell Therapy, Seoul National University Hospital, Seoul, Republic of Korea.
Biomaterials. 2013 Dec;34(38):9819-29. doi: 10.1016/j.biomaterials.2013.09.013. Epub 2013 Sep 24.
As the search for new cell types for cardiovascular regeneration continues, it has become increasingly important to optimize ex vivo cell processing. We aimed to develop an optimal processing strategy for human cardiac progenitor cells. We hypothesized that enhancing the stemness potential and promoting the secretory activity for paracrine effects are mutually exclusive routes. Therefore, we investigated the two divergent cell processing methods to enhance cellular potency and humoral activity, respectively. We obtained human right ventricular tissues and sequentially generated primary cardiosphere (CS), primary CS-derived cells (PCDC) and secondary CSs. During secondary CS formation, inhibiting the ERK pathway, using selective RTK1 and TGF-β inhibitors, Oct4 increased 20 fold and VEGF was decreased. When the ERK pathway was stimulated by addition of EGF and TGF-β, VEGF expression was upregulated and Oct4 was downregulated, indicating that the ERK pathway serves a directional role for cellular potency versus paracrine capacity. Transplantation of PCDCs or secondary CSs into the infarcted heart of immunocompromised mouse showed significant angiogenic effects compared with PBS injection. Interestingly, combined transplantation of the two differently-processed, dual-purpose secondary CSs resulted in an additional increase in neovascularization. Human VEGF was primarily produced from secondary CSs under ERK stimulating conditions. Cardiomyocyte-like cells were produced from secondary CSs under ERK inhibitory conditions. These findings indicate that combined transplantation of specifically-processed human secondary CSs enhances infarct repair through the complementary enhancement of cardiopoietic regenerative and paracrine protective effect. Furthermore, these results underscore the fact that optimal cell processing methods have the potential to maximize the therapeutic benefits.
随着对心血管再生新细胞类型的不断探索，优化细胞的离体处理方法变得越来越重要。本研究旨在开发优化的人心肌祖细胞处理策略。我们假设增强干细胞特性和促进旁分泌作用的分泌活性是相互排斥的途径。因此，我们分别采用两种不同的细胞处理方法来增强细胞活力和体液活性。我们获取了人心肌组织，依次生成原代心脏球（CS）、原代 CS 衍生细胞（PCDC）和第二代 CS。在第二代 CS 形成过程中，通过抑制 ERK 通路，使用选择性 RTK1 和 TGF-β抑制剂，Oct4 增加了 20 倍，而 VEGF 减少。当通过添加 EGF 和 TGF-β刺激 ERK 通路时，VEGF 表达上调，Oct4 下调，这表明 ERK 通路对细胞活力和旁分泌能力具有定向作用。将 PCDC 或第二代 CS 移植到免疫缺陷小鼠的梗死心脏中，与 PBS 注射相比，显示出明显的促血管生成作用。有趣的是，两种不同处理方法的、具有双重用途的第二代 CS 联合移植可进一步增加新生血管形成。在 ERK 刺激条件下，人 VEGF 主要由第二代 CS 产生。在 ERK 抑制条件下，心肌样细胞由第二代 CS 产生。这些发现表明，联合移植经过特定处理的人心肌第二代 CS 通过互补增强心脏生成再生和旁分泌保护作用，增强了梗死修复。此外，这些结果强调了优化细胞处理方法有可能最大限度地提高治疗效果。
