Institute of Macromolecular Chemistry, Czech Academy of Sciences, Heyrovsky sq.2, 162 06 Prague, Czech Republic.
Department of Chemical Engineering, University of Chemistry and Technology, Technicka 5, 166 28 Prague, Czech Republic.
Int J Mol Sci. 2021 Oct 28;22(21):11666. doi: 10.3390/ijms222111666.
Long-term delivery of growth factors and immunomodulatory agents is highly required to support the integrity of tissue in engineering constructs, e.g., formation of vasculature, and to minimize immune response in a recipient. However, for proteins with a net positive charge at the physiological pH, controlled delivery from negatively charged alginate (Alg) platforms is challenging due to electrostatic interactions that can hamper the protein release. In order to regulate such interactions between proteins and the Alg matrix, we propose to complex proteins of interest in this study - CXCL12, FGF-2, VEGF - with polyanionic heparin prior to their encapsulation into Alg microbeads of high content of α-L-guluronic acid units (high-G). This strategy effectively reduced protein interactions with Alg (as shown by model ITC and SPR experiments) and, depending on the protein type, afforded control over the protein release for at least one month. The released proteins retained their in vitro bioactivity: CXCL12 stimulated the migration of Jurkat cells, and FGF-2 and VEGF induced proliferation and maturation of HUVECs. The presence of heparin also intensified protein biological efficiency. The proposed approach for encapsulation of proteins with a positive net charge into high-G Alg hydrogels is promising for controlled long-term protein delivery under in vivo conditions.
长期输送生长因子和免疫调节剂对于支持工程构建物中组织的完整性非常重要，例如血管的形成，并最大程度地减少受体中的免疫反应。然而，对于在生理 pH 值下带正净电荷的蛋白质，由于静电相互作用会阻碍蛋白质释放，因此从带负电荷的藻酸盐（Alg）平台进行控制释放具有挑战性。为了调节蛋白质和 Alg 基质之间的这种相互作用，我们建议在将本研究中的感兴趣的蛋白质（CXCL12、FGF-2、VEGF）包封到具有高α-L-古洛糖醛酸单元含量（高-G）的 Alg 微球之前，先将其与聚阴离子肝素复合。该策略有效地减少了蛋白质与 Alg 的相互作用（如模型 ITC 和 SPR 实验所示），并且根据蛋白质类型，至少可以控制蛋白质释放一个月。释放的蛋白质保留了其体外生物活性：CXCL12 刺激 Jurkat 细胞的迁移，FGF-2 和 VEGF 诱导 HUVECs 的增殖和成熟。肝素的存在也增强了蛋白质的生物学效率。将带正净电荷的蛋白质封装到高-G Alg 水凝胶中的这种方法有望在体内条件下实现蛋白质的控制长效输送。
