Oliveira J T, Crawford A, Mundy J M, Moreira A R, Gomes M E, Hatton P V, Reis R L
3B's Research Group-Biomaterials, Biodegradables, and Biomimetics, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.
J Mater Sci Mater Med. 2007 Feb;18(2):295-302. doi: 10.1007/s10856-006-0692-7.
In the present work we originally tested the suitability of corn starch-polycaprolactone (SPCL) scaffolds for pursuing a cartilage tissue engineering approach. Bovine articular chondrocytes were seeded on SPCL scaffolds under dynamic conditions using spinner flasks (total of 4 scaffolds per spinner flask using cell suspensions of 0.5 x 10(6) cells/ml) and cultured under orbital agitation for a total of 6 weeks. Poly(glycolic acid) (PGA) non-woven scaffolds and bovine native articular cartilage were used as standard controls for the conducted experiments. PGA is a kind of standard in tissue engineering approaches and it was used as a control in that sense. The tissue engineered constructs were characterized at different time periods by scanning electron microscopy (SEM), hematoxylin-eosin (H&E) and toluidine blue stainings, immunolocalisation of collagen types I and II, and dimethylmethylene blue (DMB) assay for glycosaminoglycans (GAG) quantification assay. SEM results for SPCL constructs showed that the chondrocytes presented normal morphological features, with extensive cells presence at the surface of the support structures, and penetrating the scaffolds pores. These observations were further corroborated by H&E staining. Toluidine blue and immunohistochemistry exhibited extracellular matrix deposition throughout the 3D structure. Glycosaminoglycans, and collagen types I and II were detected. However, stronger staining for collagen type II was observed when compared to collagen type I. The PGA constructs presented similar features to SPCL at the end of the 6 weeks. PGA constructs exhibited higher amounts of matrix glycosaminoglycans when compared to the SPCL scaffolds. However, we also observed a lack of tissue in the central area of the PGA scaffolds. Reasons for these occurrences may include inefficient cells penetration, necrosis due to high cell densities, or necrosis related with acidic by-products degradation. Such situation was not detected in the SPCL scaffolds, indicating the much better biocompatibility of the starch based scaffolds.
在本研究中，我们最初测试了玉米淀粉 - 聚己内酯（SPCL）支架用于软骨组织工程方法的适用性。使用转瓶在动态条件下将牛关节软骨细胞接种到SPCL支架上（每个转瓶使用0.5×10⁶个细胞/毫升的细胞悬液共接种4个支架），并在轨道振荡下培养6周。聚乙醇酸（PGA）无纺布支架和牛天然关节软骨用作所进行实验的标准对照。PGA是组织工程方法中的一种标准材料，在这个意义上用作对照。在不同时间段通过扫描电子显微镜（SEM）、苏木精 - 伊红（H&E）和甲苯胺蓝染色、I型和II型胶原的免疫定位以及用于糖胺聚糖（GAG）定量分析的二甲基亚甲基蓝（DMB）分析对组织工程构建体进行表征。SPCL构建体的SEM结果表明，软骨细胞呈现正常形态特征，在支撑结构表面有大量细胞存在，并穿透支架孔隙。H&E染色进一步证实了这些观察结果。甲苯胺蓝和免疫组织化学显示在整个三维结构中都有细胞外基质沉积。检测到了糖胺聚糖以及I型和II型胶原。然而，与I型胶原相比，观察到II型胶原的染色更强。在6周结束时，PGA构建体呈现出与SPCL相似的特征。与SPCL支架相比，PGA构建体表现出更高量的基质糖胺聚糖。然而，我们还观察到PGA支架的中央区域缺乏组织。这些情况的原因可能包括细胞穿透效率低下、由于高细胞密度导致的坏死或与酸性副产物降解相关的坏死。在SPCL支架中未检测到这种情况，表明基于淀粉的支架具有更好的生物相容性。
