Stable and mature vascular formation is a current challenge in engineering functional tissues. Transient, non-viral gene delivery presents a unique platform for delivering genetic information to cells for tissue engineering purposes and to restore blood flow to ischemic tissue. The formation of new blood vessels can be induced by upregulation of hypoxia-inducible factor-1α (HIF-1), among other factors. We hypothesized that biodegradable polymers could be used to efficiently deliver the HIF-1α gene to human adipose-derived stromal/stem cells (hASCs) and that this treatment could recruit an existing endogenous endothelial cell population to induce angiogenesis in a 3D cell construct in vitro. In this study, end-modified poly(β-amino ester) (PBAE) nanocomplexes were first optimized for transfection of hASCs and a new biodegradable polymer with increased hydrophobicity and secondary amine structures, N'-(3-aminopropyl)-N,N-dimethylpropane-1,3-diamine end-modified poly(1,4-butanediol diacrylate-co-4-amino-1-butanol), was found to be most effective. Optimal PBAE nanocomplexes had a hydrodynamic diameter of approximately 140 nm and had a zeta potential of 30 mV. The PBAE polymer self-assembled with HIF-1α plasmid DNA and treatment of hASCs with these nanocomplexes induced 3D vascularization. Cells transfected with this polymer-DNA complex were found to have 10⁶-fold upregulation HIF-1α expression, an approximately 2-fold increase in secreted VEGF, and caused the formation of vessel tubules compared to an untransfected control. These gene therapy biomaterials may be useful for regenerative medicine.

Statement of Significance

Not only is the formation of stable vasculature a challenge for engineering human tissues in vitro, but it is also of valuable interest to clinical applications such as peripheral artery disease. Previous studies using HIF-1α to induce vascular formation have been limited by the necessity of hypoxic chambers. It would be advantageous to simulate endogenous responses to hypoxia without the need for physical hypoxia. In this study, 3D vascular formation was shown to be inducible through non-viral gene delivery of HIF-1α with new polymeric nanocomplexes. A biodegradable polymer N'-(3-aminopropyl)-N,N-dimethylpropane-1,3-diamine end-modified poly(1,4-butanediol diacrylate-co-4-amino-1-butanol) demonstrates improved transfection of human adipose-derived stem cells. This nanobiotechnology could be a promising strategy for the creation of vasculature for tissue engineering and clinical applications.

中文翻译：

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HIF-1α 的非病毒基因递送促进人类脂肪干细胞的血管生成。

稳定和成熟的血管形成是工程功能组织当前面临的挑战。瞬时、非病毒基因传递提供了一个独特的平台，用于将遗传信息传递给细胞以用于组织工程目的并恢复缺血组织的血流。除其他因素外，缺氧诱导因子-1α (HIF-1) 的上调可诱导新血管的形成。我们假设可生物降解聚合物可用于有效地将 HIF-1α 基因递送至人类脂肪来源的基质/干细胞 (hASC)，并且这种治疗可以募集现有的内源性内皮细胞群以在体外诱导 3D 细胞构建体中的血管生成. 在这项研究中，末端改性的聚（β-氨基酯）（PBAE）纳米复合物首先被优化用于转染 hASCs 和一种具有增加的疏水性和仲胺结构的新型可生物降解聚合物，N'-（3-氨基丙基）-N,N发现-二甲基丙烷-1,3-二胺末端改性的聚（1,4-丁二醇二丙烯酸酯-co-4-氨基-1-丁醇）是最有效的。最佳 PBAE 纳米复合物的流体动力学直径约为 140 nm，zeta 电位为 30 mV。PBAE 聚合物与 HIF-1α 质粒 DNA 自组装并用这些纳米复合物处理 hASC 诱导 3D 血管化。发现用这种聚合物-DNA 复合物转染的细胞有 10 ⁶与未转染的对照相比，HIF-1α 表达的上调倍数增加，分泌的 VEGF 增加约 2 倍，并导致血管小管的形成。这些基因治疗生物材料可能对再生医学有用。

重要性声明

不仅稳定脉管系统的形成是体外工程人体组织的挑战，但它对于外周动脉疾病等临床应用也具有重要意义。以前使用 HIF-1α 诱导血管形成的研究受到缺氧室必要性的限制。在不需要物理缺氧的情况下模拟对缺氧的内源性反应将是有利的。在这项研究中，3D 血管形成被证明可以通过 HIF-1α 与新的聚合物纳米复合物的非病毒基因递送来诱导。一种可生物降解的聚合物 N'-(3-氨基丙基)-N,N-二甲基丙烷-1,3-二胺末端改性的聚(1,4-丁二醇二丙烯酸酯-co-4-氨基-1-丁醇) 证明了人类转染的改善脂肪来源的干细胞。这种纳米生物技术可能是创建用于组织工程和临床应用的脉管系统的有前途的策略。