3B's Research Group-Biomaterials, Biodegradables and Biomimetics, Univ. Minho, Headquarters of the European Institute of Excellence on Tissue Engineering and Regenerative Medicine, AvePark, S. Cláudio de Barco, Taipas, Guimarães, Portugal.
Bone. 2010 May;46(5):1424-35. doi: 10.1016/j.bone.2010.02.007. Epub 2010 Feb 10.
Recently, our group has proposed a combinatorial strategy in tissue engineering principles employing carboxymethylchitosan/poly(amidoamine) dendrimer nanoparticles (CMCht/PAMAM) towards the intracellular release and regimented supply of dexamethasone (Dex) aimed at controlling stem cell osteogenic differentiation in the absence of typical osteogenic inducers, in vivo. In this work, we have investigated if the Dex-loaded CMCht/PAMAM dendrimer nanoparticles could play a crucial role in the regulation of osteogenesis, in vivo. Macroporous hydroxyapatite (HA) scaffolds were seeded with rat bone marrow stromal cells (RBMSCs), whose cells were expanded in MEM medium supplemented with 0.01 mg ml(-1) Dex-loaded CMCht/PAMAM dendrimer nanoparticles and implanted subcutaneously on the back of rats for 2 and 4 weeks. HA porous ceramics without RBMSCs and RBMSCs/HA scaffold constructs seeded with cells expanded in the presence and absence of 10(-8) M Dex were used as controls. The effect of initial cell number seeded in the HA scaffolds on the bone-forming ability of the constructs was also investigated. Qualitative and quantitative new bone formation was evaluated in a non-destructive manner using micro-computed tomography analyses of the explants. Haematoxylin and Eosin stained implant sections were also used for the histomorphometrical analysis. Toluidine blue staining was carried out to investigate the synthesis of proteoglycan extracellular matrix. In addition, alkaline phosphatase and osteocalcin levels in the explants were also quantified, since these markers denote osteogenic differentiation. At 4 weeks post-implantation results have shown that the novel Dex-loaded carboxymethylchitosan/poly(amidoamine) dendrimer nanoparticles may be beneficial as an intracellular nanocarrier, supplying Dex in a regimented manner and promoting superior ectopic de novo bone formation.
最近，我们小组提出了一种组合策略，即在组织工程学原理中使用羧甲基壳聚糖/聚（酰胺-胺）树枝状大分子纳米粒子（CMCht/PAMAM），以实现细胞内释放和有规律地供应地塞米松（Dex），从而在缺乏典型成骨诱导剂的情况下控制干细胞成骨分化，体内。在这项工作中，我们研究了负载 Dex 的 CMCht/PAMAM 树枝状大分子纳米粒子是否可以在体内调节成骨作用中发挥关键作用。大孔羟基磷灰石（HA）支架接种了大鼠骨髓基质细胞（RBMSCs），将细胞在补充有 0.01mg/ml Dex 负载的 CMCht/PAMAM 树枝状大分子纳米粒子的 MEM 培养基中扩增，并将其皮下植入大鼠背部 2 和 4 周。未接种 RBMSCs 的多孔 HA 陶瓷和接种细胞的 RBMSCs/HA 支架构建体，细胞在存在和不存在 10(-8)M Dex 的情况下扩增，作为对照。还研究了在 HA 支架中接种初始细胞数对构建体成骨能力的影响。通过对标本进行微计算机断层扫描分析，以非破坏性方式评估定性和定量的新骨形成。苏木精和伊红染色的植入物切片也用于组织形态计量学分析。甲苯胺蓝染色用于研究蛋白多糖细胞外基质的合成。此外，还定量测定了标本中的碱性磷酸酶和骨钙素水平，因为这些标志物表示成骨分化。植入后 4 周的结果表明，新型负载 Dex 的羧甲基壳聚糖/聚（酰胺-胺）树枝状大分子纳米粒子可用作细胞内纳米载体，以有规律的方式供应 Dex，并促进异位新骨形成。
