LUNAM Université, UMR S-1066, F-49933 Angers, France; INSERM U1066, MINT "Micro et Nanomédecines Biomimétiques", F-49933 Angers, France; National Institute for Cardiovascular Research (INRC), I-40126 Bologna, Italy; Department of Biomedical and Neuromotor Sciences (DIBINEM), University of Bologna, 40126 Bologna, Italy.
National Institute for Cardiovascular Research (INRC), I-40126 Bologna, Italy; Department of Biomedical and Neuromotor Sciences (DIBINEM), University of Bologna, 40126 Bologna, Italy.
J Control Release. 2014 Oct 28;192:82-94. doi: 10.1016/j.jconrel.2014.06.052. Epub 2014 Jul 3.
The challenge of tissue engineering of the infarcted heart is how to improve stem cell engraftment, survival, homing, and differentiation for myocardial repair. We here propose to integrate human adipose-derived stem cells (ADSCs) and pharmacologically active microcarriers (PAMs), a three-dimensional (3D) carrier of cells and growth factors, into an injectable hydrogel (HG), to obtain a system that stimulates the survival and/or differentiation of the grafted cells toward a cardiac phenotype. PAMs are biodegradable and non-cytotoxic poly(lactic-co-glycolic acid) (PLGA) microspheres conveying cells on their 3D surface that deliver continuously and in a controlled manner a growth factor (GF) acting on the transported cells and on the microenvironment to improve engraftment. The choice of the appropriate GF and its protection during the formulation process and delivery are essential. In this study two GFs, hepatocyte growth factor (HGF) and insulin-like growth factor (IGF-1), have been encapsulated under a solid state in order to limit their interaction with the polymer and conserve their integrity. GF precipitation conditions and release profile from PAMs have been first investigated before combining them to ADSCs. The released IGF-1 and HGF induced the protein synthesis of cardiac differentiation markers GATA4, Nkx2.5, cTnI and CX43 after 1week in vitro. Moreover, the GFs accelerated cell cycle progression, as suggested by the increased expression of Cyclin D1 mRNA and the widespread distribution of Ki67 protein. Integrating PAMs within the thermosensitive P407 hydrogel increased their elastic properties but decreased the transcription of most cardiac markers. In contrast, CX43 expression increased in ADSC-PAM-GF complexes embedded within the hydrogel compared to the ADSCs cultured alone in the absence of P407. These results suggest that particulate scaffolds releasing HGF and IGF-1 may be beneficial for applications in tissue-engineering strategies for myocardial repair and the association with a P407 hydrogel can increase substrate elasticity and junction connections in ADSCs.
梗死心脏的组织工程学面临的挑战在于如何提高干细胞的移植、存活、归巢和分化，以实现心肌修复。我们提出将人脂肪来源干细胞（ADSCs）和具有药理活性的微载体（PAMs）——一种细胞和生长因子的三维（3D）载体——整合到可注射水凝胶（HG）中，获得一种刺激移植细胞存活和/或向心脏表型分化的系统。PAMs 是可生物降解和非细胞毒性的聚乳酸-共-羟基乙酸（PLGA）微球，在其 3D 表面携带细胞，以受控方式持续输送作用于运输细胞和微环境的生长因子（GF），以提高移植细胞的植入。选择合适的 GF 及其在配方过程和输送过程中的保护至关重要。在这项研究中，两种 GF——肝细胞生长因子（HGF）和胰岛素样生长因子（IGF-1）——已被封装在固态下，以限制其与聚合物的相互作用并保持其完整性。首先研究了 GF 的沉淀条件和从 PAMs 中的释放曲线，然后将其与 ADSCs 结合。在体外培养 1 周后，释放的 IGF-1 和 HGF 诱导了心脏分化标志物 GATA4、Nkx2.5、cTnI 和 CX43 的蛋白合成。此外，GFs 加速了细胞周期进程，这一点可以从 Cyclin D1 mRNA 的表达增加和 Ki67 蛋白的广泛分布中得到证明。将 PAMs 整合到热敏性 P407 水凝胶中增加了其弹性特性，但降低了大多数心脏标志物的转录。相比之下，与单独在不存在 P407 的情况下培养的 ADSCs 相比，嵌入水凝胶中的 ADSC-PAM-GF 复合物中 CX43 的表达增加。这些结果表明，释放 HGF 和 IGF-1 的颗粒状支架可能有益于心肌修复的组织工程策略的应用，并且与 P407 水凝胶的结合可以增加 ADSCs 中的基质弹性和连接。
