Nocera Aden Díaz, Comín Romina, Salvatierra Nancy Alicia, Cid Mariana Paula
Departamento de Química, Facultad de Ciencias Exactas, Físicas y Naturales, Universidad Nacional de Córdoba, Córdoba, Argentina.
Instituto de Investigaciones Biológicas y Tecnológicas, (IIByT-CONICET-UNC), Córdoba, Argentina.
Biomed Microdevices. 2018 Feb 27;20(2):26. doi: 10.1007/s10544-018-0270-z.
Collagen is widely used in tissue engineering because it can be extracted in large quantities, and has excellent biocompatibility, good biodegradability, and weak antigenicity. In the present study, we isolated printable collagen from bovine Achilles tendon and examined the purity of the isolated collagen using sodium dodecyl sulfate polyacrylamide gel electrophoresis. The bands obtained corresponded to α, α and β chains with little contamination from other small proteins. Furthermore, rheological measurements of collagen dispersions (60 mg per ml of PBS) at pH 7 revealed values of viscosity of 35.62 ± 1.42 Pa s at shear rate of 10 s - 1 and a shear thinning behavior. Collagen gels and solutions can be used for building scaffolds by three-dimensional (3D) printing. After designing and fabricating a low-cost 3D printer we assayed the collagen printing and obtaining 3D printed scaffolds of collagen at pH 7. The porosity of the scaffold was 90.22% ± 0.88% and the swelling ratio was 1437% ± 146%. The microstructure of the scaffolds was studied using scanning electron microscopy, and a porous mesh of fibrillar collagen was observed. In addition, the 3D printed collagen scaffold was not cytotoxic with cell viability higher than 70% using Vero and NIH 3 T3 cells. In vitro evaluation using both cells lines demonstrated that the collagen scaffolds had the ability to support cell attachment and proliferation. Also a fibrillar collagen mesh was observed after two weeks of culture at 37 °C. Overall, these results are promising since they show the capability of the presented protocol to obtain printable fibrillar collagen at pH 7 and the potential of the printing technique for building low-cost biocompatible 3D plotted structures which maintained the fibrillar collagen structure after incubation in culture media without using additional strategies as crosslinking.
胶原蛋白在组织工程中被广泛应用，因为它可以大量提取，具有优异的生物相容性、良好的生物降解性和较弱的抗原性。在本研究中，我们从牛跟腱中分离出可打印的胶原蛋白，并使用十二烷基硫酸钠聚丙烯酰胺凝胶电泳检测分离出的胶原蛋白的纯度。获得的条带对应于α、α和β链，几乎没有其他小蛋白质的污染。此外，在pH值为7时对胶原蛋白分散液（每毫升PBS含60毫克）进行流变学测量，发现在剪切速率为10 s⁻¹时粘度值为35.62±1.42 Pa·s，且具有剪切变稀行为。胶原蛋白凝胶和溶液可用于通过三维（3D）打印构建支架。在设计并制造了一台低成本3D打印机后，我们对胶原蛋白打印进行了检测，并在pH值为7时获得了3D打印的胶原蛋白支架。支架的孔隙率为90.22%±0.88%，溶胀率为1437%±146%。使用扫描电子显微镜研究了支架的微观结构，观察到了纤维状胶原蛋白的多孔网络。此外，3D打印的胶原蛋白支架对Vero细胞和NIH 3T3细胞无细胞毒性，细胞活力高于70%。使用这两种细胞系进行的体外评估表明，胶原蛋白支架具有支持细胞附着和增殖的能力。在37℃培养两周后也观察到了纤维状胶原蛋白网络。总体而言，这些结果很有前景，因为它们表明了所提出的方案能够在pH值为7时获得可打印的纤维状胶原蛋白，以及打印技术在构建低成本生物相容性3D绘制结构方面的潜力，该结构在培养基中孵育后保持了纤维状胶原蛋白结构，无需使用额外的交联策略。
