Lopez Serafina G, Estroff Lara A, Bonassar Lawrence J
Meinig of Biomedical Engineering, Cornell University, Ithaca, NY 14853, USA.
Department of Materials Science and Engineering, Cornell University, Ithaca, NY 14853, USA.
Bioengineering (Basel). 2024 Dec 23;11(12):1308. doi: 10.3390/bioengineering11121308.
The complex collagen network of the native meniscus and the gradient of the density and alignment of this network through the meniscal enthesis is essential for the proper mechanical function of these tissues. This architecture is difficult to recapitulate in tissue-engineered replacement strategies. Prenatally, the organization of the collagen fiber network is established and aggrecan content is minimal. In vitro, fibrochondrocytes (FCCs) produce proteoglycans and associated glycosaminoglycan (GAG) chains early in culture, which can inhibit collagen fiber formation during the maturation of tissue-engineered menisci. Thus, it would be beneficial to both specifically and temporarily block deposition of proteoglycans early in culture. In this study, we transiently inhibited aggrecan production by meniscal fibrochondrocytes using siRNA in collagen gel-based tissue-engineered constructs. We evaluated the effect of siRNA treatment on the formation of collagen fibrils and bulk and microscale tensile properties. Specific inhibition of aggrecan production by fibrochondrocytes via siRNA was successful both in 2D monolayer cell culture and 3D tissue culture. This inhibition during early maturation of these in vitro constructs increased collagen fibril diameter by more than 2-fold. This increase in fibril diameter allowed these tissues to distribute strains more effectively at the local level, particularly at the interface of the bone and soft tissue. These data show that siRNA can be used to modulate the ECM to improve collagen fiber formation and mechanical properties in tissue-engineered constructs, and that a transient decrease in aggrecan promotes the formation of a more robust fiber network.
天然半月板复杂的胶原网络以及该网络通过半月板附着点处的密度和排列梯度对于这些组织的正常力学功能至关重要。这种结构在组织工程替代策略中很难重现。在胎儿期，胶原纤维网络的组织就已形成，而聚集蛋白聚糖的含量极少。在体外，纤维软骨细胞（FCCs）在培养早期会产生蛋白聚糖和相关的糖胺聚糖（GAG）链，这会在组织工程半月板成熟过程中抑制胶原纤维的形成。因此，在培养早期特异性且暂时地阻断蛋白聚糖的沉积将是有益的。在本研究中，我们在基于胶原凝胶的组织工程构建物中使用小干扰RNA（siRNA）瞬时抑制半月板纤维软骨细胞的聚集蛋白聚糖产生。我们评估了siRNA处理对胶原纤维形成以及整体和微观拉伸性能的影响。通过siRNA对纤维软骨细胞的聚集蛋白聚糖产生进行特异性抑制在二维单层细胞培养和三维组织培养中均取得成功。在这些体外构建物早期成熟过程中的这种抑制使胶原纤维直径增加了两倍多。纤维直径的这种增加使这些组织能够在局部水平更有效地分散应变，特别是在骨与软组织的界面处。这些数据表明，siRNA可用于调节细胞外基质，以改善组织工程构建物中的胶原纤维形成和力学性能，并且聚集蛋白聚糖的短暂减少会促进形成更坚固的纤维网络。
