Centre for Biomaterial Development and Berlin-Brandenburg Centre for Regenerative Therapies, Institute of Polymer Research, Helmholtz-Zentrum Geesthacht, Teltow, Germany.
Artif Organs. 2012 Mar;36(3):E28-38. doi: 10.1111/j.1525-1594.2011.01410.x. Epub 2012 Feb 23.
The cell population of peripheral blood monocytes/macrophages (MO) is heterogeneous: The majority of the MO are CD14++ CD16- and named "classical" (= MO1). Furthermore, two other subpopulations were described: CD14++ CD16+ ("intermediate" = MO2) and CD14+ CD16++ ("non-classical" = MO3). It is reported that MO2 possess anti-inflammatory properties and express the MO lineage marker CD163. On a hydrophilic neutrally charged acrylamide-based hydrogel human intermediate (CD14++ CD16+ ), angiogenically stimulated CD163++ monocytes/macrophages (aMO2) maintained a proangiogenic and noninflammatory status for at least 14 days. Here, we explored whether this aMO2 subset adhered to hydrophobic poly(n-butyl acrylate) networks (cPnBA) and also remained in its proangiogenic and noninflammatory status. Because substrate elasticity can impact adherence, morphology, and function of cells, cPnBAs with different Young's modulus (250 and 1100 kPa) were investigated, whereby their elasticity was tailored by variation of the cross-linker content and matched to the elasticity of human arteries. The cPnBAs exhibited similar surface properties (e.g., surface roughness), which were maintained after ethylene oxide sterilization and exposure in serum-free cell culture medium for 18 h at 37°C. aMO2 were seeded on cPnBA samples (1.7 × 10(5) cells/1.33 cm(2) ) in Dulbecco's modified Eagle medium (DMEM high glucose) supplemented with vascular endothelial growth factor 165 (VEGF-A(165) , 10 ng/mL) and fetal calf serum (10 vol%) for 3 and 72 h. On both polymeric samples (n = 3 each), the numbers of adherent cells per unit area were significantly higher (P < 0.01; cPnBA0250: 3 h 13 ± 5 cells/mm(2) , 72 h 234 ± 106 cells/mm(2) ; cPnBA1100: 3 h 14 ± 3 cells/mm(2) , 72 h 198 ± 113 cells/mm(2) ) compared to control cultures (glass, 3 h: 6 ± 3 cells/mm(2) , 72 h: 130 ± 83 cells/mm(2) ) and showed a typically spread morphology. The mRNA expression profile of the aMO2 was not influenced by the substrate elasticity. In the supernatant of aMO2 on cPnBA0250, significantly less VEGF-A(165) product was found than expected based on the mRNA level measured (P < 0.01). Tests with recombinant VEGF-A(165) then demonstrated that significantly more VEGF-A(165) was adhered on cPnBA0250 than on cPnBA1100 (P < 0.01). Seeded on cPnBA, aMO2-unaffected by the elastic moduli of both substrates-seemed to remain in their subset status and secreted VEGF-A(165) without release of proinflammatory cytokines. These in vitro results might indicate that this MO subset can be used as cellular delivery system for proangiogenic and noninflammatory mediators to support the endothelialization of cPnBA.
外周血单核细胞/巨噬细胞（MO）的细胞群体是异质的：大多数 MO 是 CD14++ CD16-，称为“经典”（= MO1）。此外，还描述了另外两个亚群：CD14++ CD16+（“中间”= MO2）和 CD14+ CD16++（“非经典”= MO3）。据报道，MO2 具有抗炎特性，并表达 MO 谱系标志物 CD163。在基于亲水性中性带电丙烯酰胺的水凝胶上，人中间（CD14++ CD16+），被血管生成刺激的 CD163++单核细胞/巨噬细胞（aMO2）至少保持 14 天的促血管生成和非炎症状态。在这里，我们探索了这种 aMO2 亚群是否粘附在疏水性聚（正丁基丙烯酰胺）网络（cPnBA）上，并且仍然保持其促血管生成和非炎症状态。因为基质弹性可以影响细胞的粘附、形态和功能，所以研究了不同杨氏模量（250 和 1100 kPa）的 cPnBAs，通过改变交联剂含量来调整它们的弹性，并与人体动脉的弹性相匹配。cPnBAs 表现出相似的表面性质（例如表面粗糙度），在环氧乙烷灭菌和在 37°C 的无血清细胞培养基中暴露 18 小时后仍保持不变。aMO2 被接种在 cPnBA 样品（1.7×10（5）个细胞/1.33 cm（2））中，在补充有血管内皮生长因子 165（VEGF-A（165），10ng/ml）和胎牛血清（10vol%）的 Dulbecco 改良 Eagle 培养基（DMEM 高葡萄糖）中孵育 3 和 72 小时。在两种聚合物样品上（n=3 个每个），单位面积的粘附细胞数量明显更高（P<0.01；cPnBA0250：3 小时 13±5 个细胞/mm（2），72 小时 234±106 个细胞/mm（2）；cPnBA1100：3 小时 14±3 个细胞/mm（2），72 小时 198±113 个细胞/mm（2））与对照培养物（玻璃，3 小时：6±3 个细胞/mm（2），72 小时：130±83 个细胞/mm（2））相比，并且表现出典型的伸展形态。aMO2 的 mRNA 表达谱不受基质弹性的影响。在 cPnBA0250 上 aMO2 的上清液中，发现的 VEGF-A（165）产物明显少于根据测量的 mRNA 水平预期的产物（P<0.01）。然后用重组 VEGF-A（165）进行的测试表明，cPnBA0250 上粘附的 VEGF-A（165）明显多于 cPnBA1100（P<0.01）。接种在 cPnBA 上的 aMO2 不受两种基质弹性模量的影响，似乎保持其亚群状态，并分泌 VEGF-A（165）而不释放促炎细胞因子。这些体外结果可能表明，这种 MO 亚群可用作促血管生成和非炎症介质的细胞递送系统，以支持 cPnBA 的内皮化。
