Cell and Molecular Biology Program, University of Arkansas, 850 W Dickson St., Rm. 601, Fayetteville, AR 72701, United States of America.
Biomed Mater. 2017 Dec 28;13(1):015023. doi: 10.1088/1748-605X/aa895c.
The therapeutic potential of biological scaffolds as adjuncts to synthetic polymers motivates the engineering of fibers formed using the extracellular matrix (ECM) secreted by cells. To capture the ECM secreted by cells during in vitro culture, a solvent degradable hollow fiber membrane (HFM) was created and utilized as a cell culture platform. NIH/3T3 fibroblasts were injected into the narrow (0.986 ± 0.042 mm) lumina of mesoporous polysulfone HFMs and maintained in culture for up to 3 weeks. Following cell culture, HFMs were dissolved using N-methyl-2-pyrrolidone and the accumulated ECM was collected. The ECM retained the filamentous dimensions of the HFM lumen. The process yielded up to 0.89 ± 0.20 mg of ECM for every mm of HFM dissolved. Immunofluorescence, second-harmonic generation microscopy, and tandem mass spectrometry indicated the presence of an array of ECM constituents, including collagen, fibronectin, and proteoglycans, while FTIR spectra suggested thorough HFM material dissolution. Isolated ECM fibers, although fragile, were amenable to handling and exhibited an average elastic modulus of 34.6 ± 15.3 kPa, ultimate tensile strength of 5.2 ± 2.2 kPa, and elongation-at-break of 29% ± 18%. ECM fibers consisted of an interconnected yet porous (32.7% ± 5.8% open space) network which supported the attachment and in vitro proliferation of mammalian cells. ECM fibers were similarly synthesized using muscle and astrocyte cells, suggesting process robustness across different cell types. Ultimately, these ECM fibers could be utilized as an alternative to synthetics for the manufacture of woven meshes targeting wound healing or regenerative medicine applications.
生物支架作为合成聚合物的辅助物具有治疗潜力,这促使人们利用细胞分泌的细胞外基质(ECM)来工程化纤维。为了在体外培养过程中捕获细胞分泌的 ECM,我们制备了一种溶剂可降解的中空纤维膜(HFM),并将其用作细胞培养平台。将 NIH/3T3 成纤维细胞注入中孔聚砜 HFMs 的狭窄(0.986±0.042 mm)内腔中,并在培养物中维持长达 3 周。细胞培养后,用 N-甲基-2-吡咯烷酮溶解 HFMs,并收集积累的 ECM。ECM 保留了 HFM 内腔的丝状尺寸。该过程每溶解 1mm 的 HFM 可得到 0.89±0.20mg 的 ECM。免疫荧光、二次谐波产生显微镜和串联质谱分析表明存在一系列 ECM 成分,包括胶原蛋白、纤维连接蛋白和糖胺聚糖,而傅里叶变换红外光谱表明 HFM 材料得到了彻底的溶解。尽管分离的 ECM 纤维很脆弱,但易于处理,表现出平均弹性模量为 34.6±15.3kPa、极限拉伸强度为 5.2±2.2kPa 和断裂伸长率为 29%±18%。ECM 纤维由相互连接但多孔(32.7%±5.8%的开放空间)的网络组成,该网络支持哺乳动物细胞的附着和体外增殖。使用肌肉和星形胶质细胞同样可以合成 ECM 纤维,表明该过程在不同细胞类型中具有稳健性。最终,这些 ECM 纤维可以替代合成纤维用于制造针对伤口愈合或再生医学应用的编织网。