Department of Biomedical Engineering, Tufts University, Medford, MA, USA.
Department of Molecular Medicine, University of Pavia, Pavia, Italy; Biotechnology Research Laboratories, IRCCS San Matteo Foundation, Pavia, Italy.
Biomaterials. 2018 Sep;178:122-133. doi: 10.1016/j.biomaterials.2018.06.018. Epub 2018 Jun 17.
In the bone marrow, the interaction of progenitor cells with the vasculature is fundamental for the release of blood cells into circulation. Silk fibroin, derived from Bombyx mori silkworm cocoons, is a promising protein biomaterial for bone marrow tissue engineering, because of its tunable architecture and mechanical properties, the capacity to incorporate labile compounds without loss of bioactivity and the demonstrated ability to support blood cell formation without premature activation. In this study, we fabricated a custom perfusion chamber to contain a multi-channel lyophilized silk sponge mimicking the vascular network in the bone marrow niche. The perfusion system consisted in an inlet and an outlet and 2 splitters that allowed funneling flow in each single channel of the silk sponge. Computational Fluid Dynamic analysis demonstrated that this design permitted confined flow inside the vascular channels. The silk channeled sponge supported efficient platelet release from megakaryocytes (Mks). After seeding, the Mks localized along SDF-1α functionalized vascular channels in the sponge. Perfusion of the channels allowed the recovery of functional platelets as demonstrated by increased PAC-1 binding upon thrombin stimulation. Further, increasing the number of channels in the silk sponge resulted in a proportional increase in the numbers of platelets recovered, suggesting applicability to scale-up for platelet production. In conclusion, we have developed a scalable system consisting of a multi-channeled silk sponge incorporated in a perfusion chamber that can provide useful technology for functional platelet production ex vivo.
在骨髓中，祖细胞与脉管系统的相互作用对于将血细胞释放到循环中至关重要。丝素蛋白来源于家蚕茧，是一种很有前途的用于骨髓组织工程的蛋白质生物材料，因为它具有可调节的结构和机械性能、能够在不损失生物活性的情况下掺入不稳定化合物的能力，以及能够支持血细胞形成而不会过早激活的能力。在这项研究中，我们制造了一个定制的灌注室来容纳多通道冻干丝海绵，以模拟骨髓龛中的血管网络。该灌注系统由入口和出口以及 2 个分流器组成，允许在丝海绵的每个单通道中引导流动。计算流体动力学分析表明，这种设计允许在血管通道内进行受限流动。带有通道的丝海绵能够有效地从巨核细胞（Mks）中释放血小板。接种后，Mks 沿着 SDF-1α 功能化的血管通道在海绵中定位。通道的灌注允许通过凝血酶刺激增加 PAC-1 结合来恢复功能性血小板。此外，增加丝海绵中的通道数量会导致回收的血小板数量成比例增加，这表明该系统可适用于规模化生产血小板。总之，我们开发了一种由多通道丝海绵和灌注室组成的可扩展系统，可为体外功能性血小板生产提供有用的技术。
