Gene Transfer and Immunogenicity Branch, Division of Cellular and Gene Therapies, Center for Biologics Evaluation and Research, Bethesda, MD, USA.
Xenotransplantation. 2010 Jul-Aug;17(4):288-99. doi: 10.1111/j.1399-3089.2010.00591.x.
Baboons are commonly used as models for transplantation and preclinical testing of various types of therapeutic agents. For proper assessment of information gathered from these models, differences between the baboon and human immune systems need to be characterized. Natural killer (NK) cells are the first line of defense against many infectious agents and cancer and are important mediators of transplantation rejection reactions, particularly during xenotransplantation. In this study, we examined baboon NK cell function and developed methods for purifying and expanding these cells.
Baboon NK cells were analyzed using a combination of extracellular and intracellular cell staining, cell sorting, interleukin (IL)-2 mediated stimulation and expansion, and 4 h cytotoxicity assays with human and pig target cell lines.
Baboon peripheral blood mononuclear cell (PBMC) exert very low but detectable cytolytic activity against both human (K562) and pig (PAEC, J2) target cells, and this activity is enhanced within 4 h of treatment with IL-2. Like human NK cells, many baboon PBMC express the lytic enzymes granzyme A, granzyme B, and perforin. Based on these markers, we identified a subpopulation of CD3(-) baboon lymphocytes that are CD8(dim) and CD16(bright) that likely represents the baboon NK cells. These cells also are characterized by expression of the natural cytotoxicity receptor NKp46. Baboon CD3(-)NKp46(+) cells purified by flow cytometric cell sorting have high cytolytic capacity that can be further enhanced by IL-2 stimulation. These baboon NK cells can be expanded in vitro and retain extremely high cytolytic capacity. While fresh baboon lymphocytes express very little CD56, the expanded baboon NK cells are predominantly CD56(+); approximately 10% of the expanded NK cells are CD56(dim), and the remainder are CD56(bright).
Baboon NK cells that are IL-2 responsive can be identified on the basis of a CD3(-)NKp46(+)CD8(dim)CD16(+/-) or CD3(-)CD8(dim)CD16(bright) phenotype and can be isolated and expanded in culture. These results may allow for a more accurate representation of the human innate immune system in baboon models and more accurate analyses of the role of the baboon innate immune system cells in preclinical models.
狒狒通常被用作各种治疗药物的移植和临床前测试的模型。为了正确评估从这些模型中收集到的信息，需要对狒狒和人类免疫系统之间的差异进行特征描述。自然杀伤 (NK) 细胞是抵御许多传染病和癌症的第一道防线，也是移植排斥反应的重要介质，特别是在异种移植中。在这项研究中，我们研究了狒狒 NK 细胞的功能，并开发了纯化和扩增这些细胞的方法。
我们使用细胞外和细胞内细胞染色、细胞分选、白细胞介素 (IL)-2 介导的刺激和扩增以及 4 小时细胞毒性测定，与人类和猪靶细胞系一起分析狒狒 NK 细胞。
狒狒外周血单核细胞 (PBMC) 对人类 (K562) 和猪 (PAEC、J2) 靶细胞表现出非常低但可检测到的细胞溶解活性，并且在 IL-2 处理后 4 小时内增强。与人类 NK 细胞一样，许多狒狒 PBMC 表达裂解酶颗粒酶 A、颗粒酶 B 和穿孔素。基于这些标志物，我们鉴定了一群 CD3(-)狒狒淋巴细胞，它们是 CD8(dim)和 CD16(bright)，可能代表狒狒 NK 细胞。这些细胞还表达自然细胞毒性受体 NKp46。通过流式细胞术细胞分选纯化的狒狒 CD3(-)NKp46(+)细胞具有高细胞溶解能力，可通过 IL-2 刺激进一步增强。这些狒狒 NK 细胞可以在体外扩增，并保持极高的细胞溶解能力。虽然新鲜的狒狒淋巴细胞表达很少的 CD56，但扩增的狒狒 NK 细胞主要是 CD56(+)；大约 10%的扩增 NK 细胞是 CD56(dim)，其余是 CD56(bright)。
基于 CD3(-)NKp46(+)CD8(dim)CD16(+/-)或 CD3(-)CD8(dim)CD16(bright)表型，可以识别对白细胞介素 (IL)-2 有反应的狒狒 NK 细胞，并可以在培养中分离和扩增。这些结果可能允许在狒狒模型中更准确地代表人类先天免疫系统，并更准确地分析狒狒先天免疫系统细胞在临床前模型中的作用。
