Objective To investigate the feasibility of fabricating an oriented scaffold combined with chondrogenic-induced bone marrow mesenchymal stem cells (BMSCs) for enhancement of the biomechanical property of tissue engineered cartilage in vivo. Methods Temperature gradient-guided thermal-induced phase separation was used to fabricate an oriented cartilage extracellular matrix-derived scaffold composed of microtubules arranged in parallel in vertical section. No-oriented scaffold was fabricated by simple freeze-drying. Mechanical property of oriented and non-oriented scaffold was determined by measurement of compressive modulus. Oriented and non-oriented scaffolds were seeded with chondrogenic-induced BMSCs, which were obtained from the New Zealand white rabbits. Proliferation, morphological characteristics, and the distribution of the cells on the scaffolds were analyzed by MTT assay and scanning electron microscope. Then cell-scaffold composites were implanted subcutaneously in the dorsa of nude mice. At 2 and 4 weeks after implantation, the samples were harvested for evaluating biochemical, histological, and biomechanical properties. Results The compressive modulus of oriented scaffold was significantly higher than that of non-oriented scaffold (t=201.099, P=0.000). The cell proliferation on the oriented scaffold was significantly higher than that on the non-oriented scaffold from 3 to 9 days (P lt; 0.05). At 4 weeks, collagen type II immunohistochemical staining, safranin O staining, and toluidine blue staining showed positive results in all samples, but negative for collagen type I. There were numerous parallel giant bundles of densely packed collagen fibers with chondrocyte-like cells on the oriented-structure constructs. Total DNA, glycosaminoglycan (GAG), and collagen contents increased with time, and no significant difference was found between 2 groups (P gt; 0.05). The compressive modulus of the oriented tissue engineered cartilage was significantly higher than that of the non-oriented tissue engineered cartilage at 2 and 4 weeks after implantation (P lt; 0.05). Total DNA, GAG, collagen contents, and compressive modulus in the 2 tissue engineered cartilages were significantly lower than those in normal cartilage (P lt; 0.05). Conclusion Oriented extracellular matrix-derived scaffold can enhance the biomechanical property of tissue engineered cartilage and thus it represents a promising approach to cartilage tissue engineering.
ObjectiveTo assess the role and effect of Wharton's jelly of human umbilical cord oriented scaffold on chondrocytes co-cultured in vitro. MethodsChondrocytes from shoulder cartilage of adult New Zealand rabbits were isolated,cultured,amplified,and labelled using fluorescent dye PKH26.Cells were extracted from human umbilical cord tissue using wet-grinding chemical technology to prepare the Wharton's jelly of human umbilical cord oriented scaffold by freeze-drying and cross-linking technology.Second generation of chondrocytes were cultured with Wharton's jelly of human umbilical cord oriented scaffold.Inverted microscope and scanning electron microscope (SEM) were used to observe the cell distribution and adhesion on the scaffold; extracellular matrix secretion of the chondrocytes were observed by toluidine blue and safranin O staining.Cells distribution and proliferation on the scaffold were assessed by fluorescein diacetate-propidium iodide (FDA-PI) and Hoechst33258 staining.The viability of the in vitro cultured and PKH26 fluorescence labelled chondrocytes on the scaffold were assessed via fluorescence microscope. ResultsInverted microscope showed that the cells cultured on the scaffold for 3 days were round or oval shaped and evenly distributed into space of the scaffold.SEM observation showed that large number of cultured cells adhered to the pores between the scaffolds and were round or oval shape,which aggregated,proliferated,and arranged vertically on longitudinally oriented scaffold at 7 days after culture.Histological observation showed that cells distributed and proliferated on the scaffold,and secreted large amount of extracellular matrix at 7 days.Scaffold could guide cell migration and proliferation,and could effectively preserve and promote the secretion of extracellular matrix.Cell viability assessments at 3 days after culture showed most of the adhered cells were living and the viability was more than 90%.PKH26 labelled chondrocytes were seen,which distributed uniformly along the pore of oriented scaffold,and exuberantly proliferated. ConclusionWharton's jelly of human umbilical cord oriented scaffold favors adhesion,proliferation,and survival of chondrocytes.It possesses a favorable affinity and cell compatibility.Thus,it is an ideal scaffold for cartilage tissue engineering.