Fazel Anvari Yazdi Abbas, Tahermanesh Kobra, Ejlali Maryam, Babaei-Ghazvini Amin, Acharya Bishnu, Badea Ildiko, MacPhee Daniel J, Chen Xiongbiao
Division of Biomedical Engineering, University of Saskatchewan, Saskatoon, Canada.
Department of Obstetrics and Gynecology, School of Medicine, Iran University of Medical Sciences (IUMS), Tehran, Iran.
Front Bioeng Biotechnol. 2024 Oct 2;12:1418034. doi: 10.3389/fbioe.2024.1418034. eCollection 2024.
Decellularized uterine extracellular matrix has emerged as a pivotal focus in the realm of biomaterials, offering a promising source in uterine tissue regeneration, research on disease diagnosis and treatments, and ultimately uterine transplantation. In this study, we examined various protocols for decellularizing porcine uterine tissues, aimed to unravel the intricate dynamics of DNA removal, bioactive molecules preservation, and microstructural alterations.
Porcine uterine tissues were treated with 6 different, yet rigorously selected and designed, protocols with sodium dodecyl sulfate (SDS), Triton X-100, peracetic acid + ethanol, and DNase I. After decellularization, we examined DNA quantification, histological staining (H&E and DAPI), glycosaminoglycans (GAG) assay, scanning electron microscopy (SEM), Fourier-transform infrared spectroscopy (FT-IR), X-ray photoelectron spectroscopy (XPS), and Thermogravimetric Analysis (TGA).
A comparative analysis among all 6 protocols was conducted with the results demonstrating that all protocols achieved decellularization; while 0.1% SDS + 1% Triton X-100, coupled with agitation, demonstrated the highest efficiency in DNA removal. Also, it was found that DNase I played a key role in enhancing the efficiency of the decellularization process by underscoring its significance in digesting cellular contents and eliminating cell debris by 99.79% (19.63 ± 3.92 ng/mg dry weight).
Our findings enhance the nuanced understanding of DNA removal, GAG preservation, microstructural alteration, and protein decomposition in decellularized uterine extracellular matrix, while highlighting the importance of decellularization protocols designed for intended applications. This study along with our findings represents meaningful progress for advancing the field of uterine transplantation and related tissue engineering/regenerative medicine.
去细胞化的子宫细胞外基质已成为生物材料领域的一个关键焦点，为子宫组织再生、疾病诊断与治疗研究乃至子宫移植提供了一个有前景的来源。在本研究中，我们研究了多种去细胞化猪子宫组织的方案，旨在揭示DNA去除、生物活性分子保留以及微观结构改变的复杂动态过程。
用十二烷基硫酸钠（SDS）、 Triton X-100、过氧乙酸+乙醇和脱氧核糖核酸酶I，对猪子宫组织采用6种经过严格筛选和设计的不同方案进行处理。去细胞化后，我们检测了DNA定量、组织学染色（苏木精-伊红染色和4',6-二脒基-2-苯基吲哚染色）、糖胺聚糖（GAG）测定、扫描电子显微镜（SEM）、傅里叶变换红外光谱（FT-IR）、X射线光电子能谱（XPS）和热重分析（TGA）。
对所有6种方案进行了比较分析，结果表明所有方案均实现了去细胞化；而0.1% SDS + 1% Triton X-100并结合搅拌，在DNA去除方面表现出最高效率。此外，发现脱氧核糖核酸酶I在提高去细胞化过程效率中起关键作用，强调了其在消化细胞内容物和清除99.79%（19.63±3.92 ng/mg干重）细胞碎片中的重要性。
我们的研究结果加深了对去细胞化子宫细胞外基质中DNA去除、GAG保留、微观结构改变和蛋白质分解的细微理解，同时突出了为特定应用设计的去细胞化方案的重要性。这项研究以及我们的发现代表了子宫移植及相关组织工程/再生医学领域取得的有意义进展。
