Weldon School of Biomedical Engineering, Purdue University, United States.
Department of Biomedical Engineering, University of California, Davis, United States.
Acta Biomater. 2019 Mar 15;87:97-107. doi: 10.1016/j.actbio.2019.01.058. Epub 2019 Jan 30.
Vocal fold scarring is the fibrotic manifestation of a variety of voice disorders, and is difficult to treat. Tissue engineering therapies provide a potential strategy to regenerate the native tissue microenvironment in order to restore vocal fold functionality. However, major challenges remain in capturing the complexity of the native tissue and sustaining regeneration. We hypothesized that hydrogels with tunable viscoelastic properties that present relevant biological cues to cells might be better suited as therapeutics. Herein, we characterized the response of human vocal fold fibroblasts to four different biomimetic hydrogels: thiolated hyaluronan (HA) crosslinked with poly(ethylene glycol) diacrylate (PEGDA), HA-PEGDA with type I collagen (HA-Col I), HA-PEGDA with type III collagen (HA-Col III) and HA-PEGDA with type I and III collagen (HA-Col I-Col III). Collagen incorporation allowed for interpenetrating fibrils of collagen within the non-fibrillar HA network, which increased the mechanical properties of the hydrogels. The addition of collagen fibrils also reduced hyaluronidase degradation of HA and hydrogel swelling ratio. Fibroblasts encapsulated in the HA-Col gels adopted a spindle shaped fibroblastic morphology by day 7 and exhibited extensive cytoskeletal networks by day 21, suggesting that the incorporation of collagen was essential for cell adhesion and spreading. Cells remained viable and synthesized new DNA throughout 21 days of culture. Gene expression levels significantly differed between the cells encapsulated in the different hydrogels. Relative fold changes in gene expression of MMP1, COL1A1, fibronectin and decorin suggest higher degrees of remodeling in HA-Col I-Col III gels in comparison to HA-Col I or HA-Col III hydrogels, suggesting that the former may better serve as a natural biomimetic hydrogel for tissue engineering applications. STATEMENT OF SIGNIFICANCE: Voice disorders affect about 1/3rd of the US population and significantly reduce quality of life. Patients with vocal fold fibrosis have few treatment options. Tissue engineering therapies provide a potential strategy to regenerate the native tissue microenvironment in order to restore vocal fold functionality. Various studies have used collagen or thiolated hyaluronan (HA) with gelatin as potential tissue engineering therapies. However, there is room for improvement in providing cells with more relevant biological cues that mimic the native tissue microenvironment and sustain regeneration. The present study introduces the use of type I collagen and type III collagen along with thiolated HA as a natural biomimetic hydrogel for vocal fold tissue engineering applications.
声带瘢痕是多种嗓音障碍的纤维表现，难以治疗。组织工程疗法提供了一种潜在的策略，可以再生天然组织微环境，以恢复声带功能。然而，在捕捉天然组织的复杂性和维持再生方面仍然存在重大挑战。我们假设具有可调节粘弹性的水凝胶，为细胞提供相关的生物学线索，可能更适合作为治疗方法。在此，我们研究了人声带成纤维细胞对四种不同仿生水凝胶的反应：巯基化透明质酸（HA）与聚乙二醇二丙烯酸酯（PEGDA）交联、HA-PEGDA 与 I 型胶原蛋白（HA-Col I）、HA-PEGDA 与 III 型胶原蛋白（HA-Col III）和 HA-PEGDA 与 I 型和 III 型胶原蛋白（HA-Col I-Col III）。胶原蛋白的加入允许胶原蛋白纤维在非纤维状 HA 网络内相互贯穿，从而增加了水凝胶的机械性能。胶原蛋白纤维的加入还降低了透明质酸酶对 HA 的降解和水凝胶的溶胀比。第 7 天，包封在 HA-Col 凝胶中的成纤维细胞呈梭形成纤维细胞形态，第 21 天表现出广泛的细胞骨架网络，表明胶原蛋白的加入对于细胞黏附和铺展是必不可少的。细胞在 21 天的培养过程中保持存活并合成新的 DNA。不同水凝胶中包封的细胞的基因表达水平有显著差异。MMP1、COL1A1、纤连蛋白和饰胶蛋白基因表达的相对倍数变化表明，与 HA-Col I 或 HA-Col III 水凝胶相比，HA-Col I-Col III 凝胶中具有更高程度的重塑，这表明前者可能更适合作为组织工程应用的天然仿生水凝胶。
约有 1/3 的美国人口受到嗓音障碍的影响，这大大降低了他们的生活质量。声带纤维化患者的治疗选择有限。组织工程疗法提供了一种潜在的策略，可以再生天然组织微环境，以恢复声带功能。各种研究已经使用胶原蛋白或巯基化透明质酸（HA）与明胶作为潜在的组织工程疗法。然而，在为细胞提供更相关的生物学线索以模拟天然组织微环境并维持再生方面仍有改进的空间。本研究介绍了使用 I 型胶原蛋白和 III 型胶原蛋白以及巯基化 HA 作为声带组织工程应用的天然仿生水凝胶。
