内皮细胞促进无支架三维血管组织中间充质干细胞的分化。

Endothelial Cells Increase Mesenchymal Stem Cell Differentiation in Scaffold-Free 3D Vascular Tissue.

作者信息

DeMaria William G, Figueroa-Milla Andre E, Kaija Abigail, Harrington Anne E, Tero Benjamin, Ryzhova Larisa, Liaw Lucy, Rolle Marsha W

机构信息

Department of Biomedical Engineering, Worcester Polytechnic Institute, Worcester, Massachusetts, USA.

MaineHealth Institute for Research, Scarborough, Maine, USA.

出版信息

Tissue Eng Part A. 2025 Jun;31(11-12):456-470. doi: 10.1089/ten.TEA.2024.0122. Epub 2024 Sep 12.

Abstract

In this study, we present a versatile, scaffold-free approach to create ring-shaped engineered vascular tissue segments using human mesenchymal stem cell-derived smooth muscle cells (hMSC-SMCs) and endothelial cells (ECs). We hypothesized that incorporation of ECs would increase hMSC-SMC differentiation without compromising tissue ring strength or fusion to form tissue tubes. Undifferentiated hMSCs and ECs were co-seeded into custom ring-shaped agarose wells using four different concentrations of ECs: 0%, 10%, 20%, and 30%. Co-seeded EC and hMSC rings were cultured in SMC differentiation medium for a total of 22 days. Tissue rings were then harvested for histology, Western blotting, wire myography, and uniaxial tensile testing to examine their structural and functional properties. Differentiated hMSC tissue rings comprising 20% and 30% ECs exhibited significantly greater SMC contractile protein expression, endothelin-1 (ET-1)-meditated contraction, and force at failure compared with the 0% EC rings. On average, the 0%, 10%, 20%, and 30% EC rings exhibited a contractile force of 0.745 ± 0.117, 0.830 ± 0.358, 1.31 ± 0.353, and 1.67 ± 0.351 mN (mean ± standard deviation [SD]) in response to ET-1, respectively. Additionally, the mean maximum force at failure for the 0%, 10%, 20%, and 30% EC rings was 88.5 ± 36. , 121 ± 59.1, 147 ± 43.1, and 206 ±  0.8 mN (mean ± SD), respectively. Based on these results, 30% EC rings were fused together to form tissue-engineered blood vessels (TEBVs) and compared with 0% EC TEBV controls. The addition of 30% ECs in TEBVs did not affect ring fusion but did result in significantly greater SMC protein expression (calponin and smoothelin). In summary, co-seeding hMSCs with ECs to form tissue rings resulted in greater contraction, strength, and hMSC-SMC differentiation compared with hMSCs alone and indicates a method to create a functional 3D human vascular cell coculture model. Impact Statement The goal of this work is to create an vascular model that exhibits structural and functional properties similar to those of native vascular tissue. For the first time, we demonstrated that human mesenchymal stem cells cocultured with endothelial cells as 3D cell aggregates, differentiated into smooth muscle cells, exhibited contractile protein expression, and contracted in response to endothelin-1. These tissue rings could be fused together to form cohesive tubular constructs to mimic the geometry of native vasculature. Overall, this study demonstrated a novel method to create and assess 3D human vascular tissue constructs using quantitative metrics.

摘要

在本研究中,我们提出了一种通用的、无支架的方法,利用人骨髓间充质干细胞来源的平滑肌细胞(hMSC-SMCs)和内皮细胞(ECs)创建环形工程化血管组织片段。我们假设加入ECs会增加hMSC-SMC的分化,同时不影响组织环的强度或融合形成组织管的能力。未分化的hMSCs和ECs以四种不同浓度的ECs(0%、10%、20%和30%)共接种到定制的环形琼脂糖孔中。共接种的EC和hMSC环在SMC分化培养基中总共培养22天。然后收获组织环进行组织学、蛋白质免疫印迹、线肌描记法和单轴拉伸试验,以检查其结构和功能特性。与0% EC环相比,包含20%和30% ECs的分化hMSC组织环表现出显著更高的SMC收缩蛋白表达、内皮素-1(ET-1)介导的收缩以及破坏时的力。平均而言,0%、10%、20%和30% EC环在ET-1刺激下的收缩力分别为0.745±0.117、0.830±0.358、1.31±0.353和1.67±0.351 mN(平均值±标准差[SD])。此外,0%、10%、20%和30% EC环的平均破坏时最大力分别为88.5±36.、121±59.1、147±43.1和206±0.8 mN(平均值±SD)。基于这些结果,将30% EC环融合在一起形成组织工程血管(TEBVs),并与0% EC TEBV对照进行比较。在TEBVs中加入30% ECs不影响环的融合,但确实导致SMC蛋白表达(钙调蛋白和平滑肌蛋白)显著增加。总之,与单独的hMSCs相比,将hMSCs与ECs共接种形成组织环会导致更大的收缩、强度和hMSC-SMC分化,并表明了一种创建功能性3D人血管细胞共培养模型的方法。影响声明这项工作的目标是创建一个具有与天然血管组织相似的结构和功能特性的血管模型。我们首次证明,作为3D细胞聚集体与内皮细胞共培养的人骨髓间充质干细胞分化为平滑肌细胞,表现出收缩蛋白表达,并对内皮素-1产生收缩反应。这些组织环可以融合在一起形成有粘性的管状结构,以模拟天然脉管系统的几何形状。总体而言,本研究展示了一种使用定量指标创建和评估3D人血管组织构建体的新方法。

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