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首页> 美国卫生研究院文献>Bioengineering >An Approach towards a GMP Compliant In-Vitro Expansion of Human Adipose Stem Cells for Autologous Therapies
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An Approach towards a GMP Compliant In-Vitro Expansion of Human Adipose Stem Cells for Autologous Therapies

机译：一种符合人脂肪疗法的GMP符合体外扩增的方法用于自体疗法

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Human Adipose Tissue Stem Cells (hASCs) are a valuable source of cells for clinical applications (e.g., treatment of acute myocardial infarction and inflammatory diseases), especially in the field of regenerative medicine. However, for autologous (patient-specific) and allogeneic (off-the-shelf) hASC-based therapies, in-vitro expansion is necessary prior to the clinical application in order to achieve the required cell numbers. Safe, reproducible and economic in-vitro expansion of hASCs for autologous therapies is more problematic because the cell material changes for each treatment. Moreover, cell material is normally isolated from non-healthy or older patients, which further complicates successful in-vitro expansion. Hence, the goal of this study was to perform cell expansion studies with hASCs isolated from two different patients/donors (i.e., different ages and health statuses) under xeno- and serum-free conditions in static, planar (2D) and dynamically mixed (3D) cultivation systems. Our primary aim was I) to compare donor variability under in-vitro conditions and II) to develop and establish an unstructured, segregated growth model as a proof-of-concept study. Maximum cell densities of between 0.49 and 0.65 × 105 hASCs/cm2 were achieved for both donors in 2D and 3D cultivation systems. Cell growth under static and dynamically mixed conditions was comparable, which demonstrated that hydrodynamic stresses (P/V = 0.63 W/m3, τnt = 4.96 × 10−3 Pa) acting at Ns1u (49 rpm for 10 g/L) did not negatively affect cell growth, even under serum-free conditions. However, donor-dependent differences in the cell size were found, which resulted in significantly different maximum cell densities for each of the two donors. In both cases, stemness was well maintained under static 2D and dynamic 3D conditions, as long as the cells were not hyperconfluent. The optimal point for cell harvesting was identified as between cell densities of 0.41 and 0.56 × 105 hASCs/cm2 (end of exponential growth phase). The growth model delivered reliable predictions for cell growth, substrate consumption and metabolite production in both types of cultivation systems. Therefore, the model can be used as a basis for future investigations in order to develop a robust MC-based hASC production process for autologous therapies.
机译：人脂肪组织干细胞（HASC）是临床应用的有价值的细胞来源（例如，急性心肌梗死和炎性疾病的治疗），特别是在再生医学领域。然而，对于基于自体（特异性患者特异性的）和同种异体（搁板）的基于且基于架子的疗法，在临床应用之前需要在体外​​膨胀，以达到所需的细胞数。用于自体疗法的HASC的安全性，可重复和经济的体外膨胀是更有问题的，因为细胞材料对每种治疗变化。此外，细胞材料通常与非健康或老年患者分离，进一步使体外扩张的成功变得复杂。因此，本研究的目的是在静态，平面（2D）中的无血清和血清条件下与两种不同患者/供体（即，不同年龄和健康状况）分离的HASCs进行细胞扩张研究，并在静态，平面（2D）和动态混合（ 3D）栽培系统。我们的主要目的是i）在体外条件下比较捐赠者变异，II）以发展和建立一个非结构化的被隔离的生长模型作为概念验证研究。对于2D和3D栽培系统的供体，实现了0.49和0.65×105个HASCS / CM2之间的最大细胞密度。静态和动态混合条件下的细胞生长是可比的，其表明，在NS1U（49rpm 10g / L）上作用的流体动力应力（p / v = 0.63 w / m 3，τnt= 4.96×10-3 pa）不会产生负面影响即使在无血清条件下，也会影响细胞生长。然而，发现细胞大小的供体依赖性差异，这导致两个供体中的每一个的最大细胞密度显着不同。在这两种情况下，只要细胞不是超链型，茎不良就会保持良好的静态2D和动态3D条件。细胞收获的最佳点被鉴定为0.41和0.56×105碱基/ cm2（指数生长阶段的末端）之间的细胞密度。增长模型在两种类型的栽培系统中提供了对细胞生长，底物消耗和代谢物产生的可靠预测。因此，该模型可以用作未来调查的基础，以便开发用于自体疗法的鲁棒基于MC的HASC生产过程。

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