Bader Andrew R, Li Tina, Wang Weiping, Kohane Daniel S, Loscalzo Joseph, Zhang Ying-Yi
Department of Medicine, Brigham and Women's Hospital and Harvard Medical School.
Laboratory for Biomaterials and Drug Delivery, Department of Anesthesiology, Division of Critical Care, Boston Children's Hospital.
J Vis Exp. 2015 Jan 22(95):52323. doi: 10.3791/52323.
Chitosan (CS) and dextran sulfate (DS) are charged polysaccharides (glycans), which form polyelectrolyte complex-based nanoparticles when mixed under appropriate conditions. The glycan nanoparticles are useful carriers for protein factors, which facilitate the in vivo delivery of the proteins and sustain their retention in the targeted tissue. The glycan polyelectrolyte complexes are also ideal for protein delivery, as the incorporation is carried out in aqueous solution, which reduces the likelihood of inactivation of the proteins. Proteins with a heparin-binding site adhere to dextran sulfate readily, and are, in turn, stabilized by the binding. These particles are also less inflammatory and toxic when delivered in vivo. In the protocol described below, SDF-1α (Stromal cell-derived factor-1α), a stem cell homing factor, is first mixed and incubated with dextran sulfate. Chitosan is added to the mixture to form polyelectrolyte complexes, followed by zinc sulfate to stabilize the complexes with zinc bridges. The resultant SDF-1α-DS-CS particles are measured for size (diameter) and surface charge (zeta potential). The amount of the incorporated SDF-1α is determined, followed by measurements of its in vitro release rate and its chemotactic activity in a particle-bound form.
壳聚糖（CS）和硫酸葡聚糖（DS）是带电荷的多糖（聚糖），在适当条件下混合时会形成基于聚电解质复合物的纳米颗粒。聚糖纳米颗粒是蛋白质因子的有用载体，有助于蛋白质在体内的递送并使其在靶组织中保持。聚糖聚电解质复合物对于蛋白质递送也是理想的，因为掺入是在水溶液中进行的，这降低了蛋白质失活的可能性。具有肝素结合位点的蛋白质很容易与硫酸葡聚糖结合，进而通过结合而稳定。这些颗粒在体内递送时炎症性和毒性也较小。在下面所述的方案中，首先将干细胞归巢因子基质细胞衍生因子-1α（SDF-1α）与硫酸葡聚糖混合并孵育。向混合物中加入壳聚糖以形成聚电解质复合物，然后加入硫酸锌以通过锌桥稳定复合物。对所得的SDF-1α-DS-CS颗粒进行尺寸（直径）和表面电荷（zeta电位）测量。测定掺入的SDF-1α的量，然后测量其体外释放速率及其以颗粒结合形式的趋化活性。
